Low and Intermediate risk protocol - BSPHO...2017/01/02 · European Low and Intermediate Risk Neuroblastoma Protocol: A SIOPEN Study Version 5.0; 1 October2015 Page 3 of 237 1. LIST

European Low and Intermediate Risk Neuroblastoma Protocol: A SIOPEN Study

Version 5.0; 1 October2015 Page 1 of 237

European Low and

Intermediate Risk

Neuroblastoma

A SIOPEN Study

Study Group 1: A phase III randomised prospective multi-centre trial

Study Groups 2-10 and Neonatal Suprarenal Masses: Non-blinded, one-armed, multi-centre prospective trials

Sponsor: FUNDACION PARA LA INVESTIGACION HOSPITAL UNIVERSITARIO LA FE.

6ª PLANTA ESCUELA DE ENFERMERIA

AVDA DE CAMPANAR 21

46009 VALENCIA

ESPAÑA

Status: Version 5.0; 1 October 2015

EudraCT number: 2010-021396-81



IMPORTANT INFORMATION ABOUT THIS CLINICAL TRIAL

This trial does not intend to use new drugs in neuroblastoma treatment. This is based on

previously and commonly used drugs for this disease. Therefore, the chemotherapy agents and

other substances will not be provided freely by any pharmaceutical company or the sponsor. The

pharmacy Unit of the participating institutions must ensure and perform the traceability of the

medication according to current legislation

Diagnosis, treatment and follow-up of patients with neuroblastoma requires a high-level of

competence and infrastructure. Therefore, these patients only can be treated in highly-specialized

paediatric oncology units. Treatment must be administered according to GCP, European and

National Clinical Trial Directives.

The trial committee does not intend it to be used as an aide-memoire or guide for treatment

of NON-registered patients.Amendments may be necessary; these will be circulated to known

participants of the trial, but institutions entering patients are advised to contact the appropriate

study centres to confirm the correctness of theprotocol in their possession. Before entering

patients clinicians must ensure that the study protocol has been cleared by their ethical committee.



1. LIST OF RESPONSIBILITIES

Structure of SIOPEN

Low and Intermediate Risk Neuroblastoma Protocol

Principal Investigators (PI)

Low Risk (LR) Study: Gudrun Schleiermacher PI- Kate Wheeler coPI

Intermediate Risk (IR) Study: Andrea di Cataldo PI- Adela Cañete coPI

Neonatal Suprarenal Mass

Study (NSM): Adela Cañete PI, Vassilios Papadakis coPI

Study Committee

Adela Cañete, Victoria Castel, Andrea di Cataldo, Ruth Ladenstein, Jean Michon, Vassilios Papadakis,

Gudrun Schleiermacher, Kate Wheeler, Veronique Mosseri, José Bermudez

National Coordinators

Australia Toby Trahair

Austria Ruth Ladenstein

Belgium Bénédicte Brichard

Czech Republic Josef Malis

Denmark Henrik Schroeder

France LR: Gudrun Schleiermacher; IR: Anne-Sophie Defachelles;

NSM: Dominique Plantaz

Greece LR, NSM:Vassilios Papadakis;IR: Margarita Baka

Hungary Miklós Garami

Ireland Anne O’Meara and Michael Capra

Israel Shifra Ash

Italy LR: Massimo Conte; IR: Andrea di Cataldo;

NSM: Massimo Conte

Norway LR: Maria W Gunnes; IR: Ellen Ruud; NSM: Maria W Gunnes

Poland Walentyna Balwierz

Portugal Ana Forjaz de Lacerda

Serbia Dragana Vujic

Slovakia Pavel Bician

Spain LR: Adela Cañete; IR: Victoria Castel; NSM: Adela Cañete

Sweden (including Iceland) Per Kogner

Switzerland Maja Beck-Popovic

United Kingdom LR: Kate Wheeler; IR:Deborah Tweddle; NSM: Kate Wheeler

Specialty Committee Coordinators

Biology Gudrun Schleiermacher, Gian Paolo Tonini, Peter Ambros

Bone Marrow Klaus Beiske

Nuclear Medicine & Physics Ariane Boubaker

Pathology Michel Peuchmaur

Radiology Claudio Granata

Radiotherapy Mark Gaze, Tom Boterberg

Statistics Veronique Mosseri, Jose D. Bermudez

Surgery Sabine Sarnacki, Roly Squire



2. SYNOPSES

2.1 Synopsis - Low Risk Neuroblastoma

Sponsor FUNDACION PARA LA INVESTIGACION HOSPITAL

UNIVERSITARIO LA FE.

Principal investigator Dr Gudrun Schleiermacher/ Dr Kate Wheeler

Title of the study Low Risk Neuroblastoma European Study

EudraCT number 2010-021396-81

Participating centres Paediatric Haematology and Oncology Units in participating

European Countries

Time schedule Inclusion first patient: 2011

Inclusion last patient: 2016

Study type Study group 1: Phase III randomised prospective multi-centre

trial

Study groups 2-6: Multi-centre, non-blinded, one armed

prospective trials

Aims Primary aim:

To demonstrate through a randomisation between

observation and chemotherapy that you can safely reduce

treatment in a subgroup of L2 low risk patients (those

without life threatening symptoms (LTS) and without any

segmental chromosomal changes (SCA), i.e. study group

1) by giving less treatment than has been given historically

while maintaining an excellent OS of 100%.

Secondary aims:

To maintain a 2 year EFS of at least 90% and an OS of at

least 95% in L2 patients with LTS without SCA (study

group 2)

To maintain the 2 year EFS of 85% and an OS of at least

98% in Ms patients without SCA (study groups 4 and 5)

To improve the 2 year EFS to at least 90% and maintain

the OS of close to 100% in L2 patients with SCA (Study

Group 3) and improve the 2 year EFS to over 70% in Ms

patients with SCA (study group 6)

To evaluate adherence to the protocol recommendations

regarding LTS

To reduce surgical morbidity by promoting strict

adherence to IDRFs to determine surgical resectability

To define the long term follow-up and natural history of

the Stage L2 non-resected masses that have remained

IDRF positive at the end of treatment (study groups 1-3).



Inclusion and exclusion

criteria

Inclusion criteria for the whole low risk group:

informed consent and follow-up warranted; group

assignment completed within 6 weeks from diagnosis;

no prior chemotherapy or radiotherapy

Biopsy proven neuroblastoma

Tumour genomic profile obtained in a NRL according

to guidelines

MYCN non-amplified

Exclusion criteria for the whole low risk group:

Diagnosis of ganglioneuroma or ganglioneuroblastoma

intermixed

INRG Stage L2

Inclusion criteria:

age ≤ 18 months

Exclusion criteria:

any metastatic site

MYCN amplification

age > 18 months

INRG Stage Ms

Inclusion criteria:

age ≤ 12 months

Exclusion criteria:

bone, pleura/lung and/or CNS metastasis

MYCN amplification

age > 12 months

Risk group allocation These patients are treated within 6 separate study groups:

Group 1: Stage L2, ≤ 18 months, MYCN non-amplified with a

NCA genomic profile, with no life threatening symptoms

Group 2: Stage L2, ≤ 18 months, MYCN non-amplified, NCA

genomic profile, with life threatening symptoms

Group 3: Stage L2, ≤ 18 months, MYCN non-amplified, SCA

genomic profile, without or with life threatening symptoms.

Group 4: Stage Ms, ≤ 12 months, MYCN non-amplified, NCA

genomic profile, without life threatening symptoms

Group 5: Stage Ms, ≤ 12 months, MYCN non-amplified, NCA

genomic profile, with life threatening symptoms

Group 6: Stage Ms, ≤ 12 months, MYCN non-amplified SCA

genomic profile, without or with life threatening symptoms

Patients without a genomic profile should be treated according

to the guidelines outlined in section 9.4.7 (page 72). Data will

be collected on these patients.



NCA : numerical chromosome alterations

SCA : segmental chromosome alterations

Treatment Study group 1: Randomisation between observation and two-

six courses of chemotherapy (CO x 2-4 ± VP/Carbo x 2) ±

surgery

Study group 2: Two-four courses of chemotherapy (VP/Carbo

x 2 ± CADO x 2) ± surgery

Study group 3: Four courses of chemotherapy (VP/Carbo x 2-4

± CADO x 2) ± surgery

Study group 4: Observation only

Study group 5: Two-four courses of chemotherapy (VP/Carbo

x 2 ± CADO x 2)

Study group 6: Four courses of chemotherapy (VP/Carbo x 2-4

± CADO x 2) ± surgery

Patient recruitment

expected over 5 years

Total: 410

Group 1: 150

Group 2: 60

Group 3: 30

Group 4: 90

Group 5: 50

Group 6: 30

Statistical methods Study group 1: Randomised phase 3 trial comparing

chemotherapy versus observation with chemotherapy initiated

if progression occurs. The primary endpoint is the time of

initiation of VP/Carbo during the first year following

diagnosis: the 1-year rate will be approximately 55% in the

chemotherapy group, with a two-sided test of 5% and power

85%, a trial of 150 patients will be sufficient to detect a

reduction of 25% (Freedman).

The trial will be formally reviewed by an independent Data

Monitoring Committee (DMC) should the 1-year death rate

exceed 1% at any time during the course of the trial in either

arm.

Survival, event free survival (EFS) and time of initiation of

chemotherapy will be calculated, from date of randomisation,

by Kaplan-Meier, and compared by the logrank test.

Study groups 2-6: Single arm strategies which will be judged

by reference to historical information for each study.

The number of patients within the various subgroups will not

be sufficient to base sample size calculations on conventional

methods of test size and power. Hence the general objective is

to obtain as many patients as possible within 5 years, and



thereby to estimate the endpoint statistics (2 year-EFS), with a

two-sided 95% confidence interval. In addition, adopting a

Bayesian approach, the probability that the endpoint of

concern is worse than the specified target will be calculated.

EFS will be estimated by Kaplan-Meier, from the date of

presentation to the date of the first event, that is, progressive

disease, recurrence or death.

The study groups will be monitored with respect to safety so

guidelines, based on the 1 year-event rate (and, for study

groups 4-5, the 1-year-death rate), are provided for each study

for the DMC, who will review the data on an annual basis.

GCP conformity This trial will be performed according to the international

accepted guidelines of Good Clinical Practice (ICH-GCP)

including archival storage of essential documents.

Financial support Financial support will be realised on a national basis. For

details see country specific appendix.

2.2 Synopsis - Intermediate Risk Neuroblastoma



Principal investigator Prof. Andrea Di Cataldo/Dr Adela Canete

Title of the study Intermediate Risk Neuroblastoma European Study



European Countries



Study type Multi-centre, non-blinded, one armed prospective clinical trial

Aims Primary aim:

To improve the EFS to 70% with an OS of 90% of INRG

stage L2 patients over the age of 18 months, with poorly

differentiated or undifferentiated tumour histology (INPC

criteria), by the addition of radiotherapy and 13-cis RA

compared to historical conventional treatment (study group

8).

Secondary aims:

To confirm in a larger patient cohort the excellent OS of

95% in stage M neuroblastoma without MYCN

amplification, less than 12 months of age, when treated



with moderate therapy (study group 10).

To maintain the excellent results of 3 year EFS of 90% and

3 year OS of 100% in stage L2 patients over the age of 18

months, with differentiating neuroblastoma or

differentiating ganglioneuroblastoma nodular, despite a

treatment reduction (study group 7).

To improve the 3 year EFS to at least 50% and the 3 year

OS to 80% in INSS stage I patients with MYCN amplified

neuroblastoma by the addition of adjuvant treatment (study

group 9).

To evaluate the impact of the tumour genomic profile on

patient outcome, in order to consider its role in the

treatment stratification of these intermediate risk patients

(all study groups).


criteria

Inclusion criteria for the whole intermediate risk group:

informed consent and follow-up warranted; group

assignment completed within 6 weeks from diagnosis;

no prior chemotherapy or radiotherapy

Tumour material available for biological studies

according to guidelines

Biopsy proven neuroblastoma confirmed in a NRL

Exclusion criteria for the whole intermediate risk group:

Diagnosis of ganglioneuroma or ganglioneuroblastoma

intermixed INRG Stage L1 and INSS stage 1:

Inclusion criteria:

MYCN amplified

Exclusion criteria:

MYCN non-amplified

INSS stages 2, 3, 4, 4s

INRG Stage L2:

Inclusion criteria:

Histology: differentiating, poorly differentiated,

undifferentiated neuroblastoma or

ganglioneuroblastoma nodular

MYCN non-amplified

age >18 months

Exclusion criteria:

neuroblastoma NOS

MYCN amplification.

age ≤ 18 months

INRG Stage M:

Inclusion criteria:

Any histology

MYCN non-amplified

age ≤ 12 months



Exclusion criteria:

MYCN amplification

age > 12 months

Risk group allocation These patients are treated within 4 separate study groups:

Group 7: INRG stage L2 neuroblastoma or

ganglioneuroblastoma nodular, differentiating histology,

MYCN non-amplified, age >18 months

Group 8: INRG stage L2 neuroblastoma or

ganglioneuroblastoma nodular, poorly differentiated or

undifferentiated histology, MYCN non-amplified, age >18

months

Group 9: INSS stage 1 neuroblastoma, MYCN amplified, any

age

Group 10: INRG stage M neuroblastoma, MYCN non-

amplified, age <12 months

L2 patients without a defined histotype should be treated

according to the guidelines outlined in section 10.4.5 (page

91). Data will be collected on these patients.

Treatment Group 7: Four courses of chemotherapy (VP/Carbo x 2 +

CADO x 2 or VP/Carbo x 2) ± surgery

Group 8: Six courses of chemotherapy (VP/Carbo x 2, CADO

x 2, VP/Carbo x 1 + CADO x 1 or CADO x 2), ± surgery,

radiotherapy, and 6 courses of 13-cis-RA

Group 9: Six courses of chemotherapy (VP/Carbo x 2, CADO

x 2, VP/Carbo x 1, CADO x1) followed by radiotherapy and 6

courses of 13-cis-RA

Group 10: Four-eight courses of chemotherapy (VP/Carbo x 2-

4 ± CADO x 2-4), ± surgery

Patient recruitment


Total: 175

Group 7: 25

Group 8: 75

Group 9: 15

Group 10: 60

Statistical methods All study groups: Single arm strategies which will be judged

by reference to historical information for each study.

The number of patients within the various subgroups will not

be sufficient to base sample size calculations on conventional

methods of test size and power. Hence the general objective is

to obtain as many patients as possible within 5 years, and

thereby to estimate the endpoint statistics (3 year-EFS), with a



two-sided 95% confidence interval. In addition, adopting a

Bayesian approach, the probability that the endpoint of concern

is worse than the specified target will be calculated.

EFS will be estimated by Kaplan-Meier, from the date of

presentation to the date of the first event, that is, progressive

disease, recurrence or death.

The study groups will be monitored with respect to safety so

guidelines, based on the 1 year-event rate and 1-year-death

rate, are provided for each study for the DMC, who will review

the data on an annual basis.



including archival storage of essential documents



2.3 Synopsis – Neonatal Suprarenal Masses



Principal investigator Dr Adela Canete/Dr Vassilios Papadakis

Title of the study Neonatal Suprarenal Masses: An Observational Approach



European Countries



Study type Multi-centre, non-blinded, one armed prospective trial

Aims Primary aim:

To maintain a 3-year event free survival over 80% with a

non-operative therapeutic approach (serial monitoring,

surgery if warranted) in infants with a localised suprarenal

mass discovered ante or neonatally.

Secondary aims:

To manage infants with suprarenal masses discovered ante

or neonatally with a uniform approach in Europe in a

multicentre setting.

To maintain an excellent overall survival with a non-

operative therapeutic approach (serial monitoring, surgery



if warranted) in infants with a localised suprarenal mass

discovered ante or neonatally.

To determine the 3-year surgery-free survival in infants

with suprarenal masses discovered ante or neonatally and

managed conservatively (non initial surgery).

To find out the natural history of perinatal suprarenal

masses, according to the definitions set up for the study.

To study the kinetics of regression in those suspected

suprarenal neuroblastomas in infants with suprarenal

masses discovered ante or neonatally and managed

conservatively (non initial surgery).

To collect tissue from those suprarenal masses excised in

order to perform standard and investigational pathological

and biological studies (INPC, MYCN, 1p, 11).

To collect frozen plasma from all patients included in the

study in order to perform research.


criteria

Inclusion criteria:

Age less than or equal to 90 days when the suprarenal

mass is discovered.

Suprarenal mass detected by ultrasound and/or MRI.

The suprarenal mass may be cystic and/or solid, but IT

CANNOT REACH THE MIDLINE AND should

MEASURE ≤ 5 CM AT THE LARGEST DIAMETER.

No regional involvement: MRI scan does not show

evidence of positive ipsi/contralateral lymph nodes or

other spread outside the suprarenal gland.

No metastatic involvement.

Frozen plasma available.

Informed consent.

Availability to do the adequate follow-up

Exclusion criteria:

Age older than 90 days.

Suprarenal mass bigger than 5 cm.

Regional involvement.

Metastatic involvement.

Inability to undertake mandatory diagnostic studies

(biological markers, US, MRI, MIBG).

Follow-up not guaranteed by parents/guardians.

Treatment Observation

Patient recruitment


Total: 100

Statistical methods This is a single arm strategy the success or otherwise of which

will be judged by reference to historical information

(summarised in the background). It is also expected that the

numbers of patients within this subgroup will not be sufficient

to base sample size calculations on conventional methods of



test size and power. Hence the general objective is to obtain as

many patients as possible within a reasonable time frame

(initially set at 5 years) and thereby to obtain a two-sided 95%

confidence interval for the endpoint measure (3-year EFS) and

use this as the basis for conclusions.

Monitoring: In order to monitor the possibility of an increased

number of events the trial will be formally reviewed by an

independent Data Monitoring Committee should the

cumulative 1-year event rate exceed 15% or the 1-year death

rate exceed 1% at any time during the course of the study.



including archival storage of essential documents





3. ABBREVIATIONS

Abbreviations used in this protocol in alphabetical order

13-cis RA 13-cis-retinoic acid

ABMT autologous bone marrow transplant

ADL activities of daily living

AH adrenal haemorrhage

ANC absolute neutrophil count

AP anteroposterior

BM bone marrow

BUMEL busulphan and melphalan (MAT regimen)

CADO cyclophosphamide, adriamycin, vincristine

CARBO carboplatin

CDDP cisplatin

CGH comparative genomic hybridisation

CI confidence interval

CNS central nervous system

CO vincristine and cyclophosphamide

COG Children’s Oncology Group

CR complete response

CRF case report form

CT computer tomography

CTDI computer tomography dose index

CXR chest x-ray

CYC cyclophosphamide

DIC disseminated intravascular coagulation

DMC data monitoring committee

DMSA dimercaptosuccinic acid

DNA deoxyribonucleic acid

DTPA diethylene triamine pentaacetic acid

Dx diagnosis

EC ethics committee

EDTA ethylenediaminetetraacetic acid

EFS event-free survival

ENQUA European Neuroblastoma Quality Assessment

EUNS European Unresectable Neuroblastoma Study

FISH Fluorescence in situ hybridization

GFR glomerular filtration rate

GI gastro-intestinal tract

Gy Gray

HVA homovanillic acid

IDRF image defined risk factors

IL2 interleukin-2

INES Infant Neuroblastoma European Study

INPC International Neuroblastoma Pathology Classification

INRG International Neuroblastoma Risk Group

INRGSS International Neuroblastoma Risk Group Staging System

INSS International Neuroblastoma Staging System

IRB institutional review board

IV intravenous

IVC inferior vena cava

LDH lactate dehydrogenase

LI local irradiation

LINES Low and Intermediate risk Neuroblastoma European Study

LLN lower limit of normal

LNESG-2 Localised Neuroblastoma European Study Group 2

LTS life threatening symptoms

MAT myeloablative therapy

MDCT multi-detector row computer tomography



MDP methylene diphosphate

MDT multidisciplinary team

MIBG meta-iodobenzylguanidine

MKI mitosis-karyorrhexis index

MLPA multiplex ligation-dependent probe amplification

MNC mononuclear cell

MPR multiplanar reconstructions

MRI magnetic resonance imaging

MTD maximum tolerated dose

MYCN V-Myc myelocytomatosis viral related oncogene, neuroblastoma derived (avian)

NSM neonatal suprarenal masses

NB neuroblastoma

NCA numerical chromosomal abnormalities

NRL national reference laboratory

NSE neuron-specific enolase

OS overall survival

PB peripheral blood

PBSCR peripheral blood stem cell rescue

PCR polymerase chain reaction

PD progressive disease

PNT peripheral neuroblastic tumour

POG Pediatric Oncology Group

RA 13-cis retinoic acid

RDE remote data entry

RHA regulatory health authorities

RL reference laboratory

RNA ribonucleic acid

RUQ right upper quadrant

SAR specific absorption rate

SCA segmental chromosomal abnormalities

SCC spinal cord compression

SD stable disease

SEPS subdiaphragmatic extralobar pulmonary sequestration

SIOP Société Internationale d’Oncologie Pédiatrique

SIOPEN Société Internationale d’Oncologie Pédiatrique European Neuroblastoma

SMZ sulfamethoxazol

SNP single nuleotide polymorphism

STIR short-tau inversion recovery

TBI total body irradiation

TMP trimethoprim

TVD topotecan, vincristine and doxorubicin

US ultrasonography

ULN upper limit normal

VCR vincristine

VMA vanillyl mandelic acid

VOD veno-occlusive disease

VP16 etoposide

W weeks

WBC white blood cells

WBMRI whole body magnetic resonance imaging WI weighted images



TABLE OF CONTENTS

1. LIST OF RESPONSIBILITIES ........................................................................................... 3

2. SYNOPSES ............................................................................................................................ 4

2.1 SYNOPSIS - LOW RISK NEUROBLASTOMA .............................................................................. 4

2.2 SYNOPSIS - INTERMEDIATE RISK NEUROBLASTOMA ......................................................... 7

2.3 SYNOPSIS – NEONATAL SUPRARENAL MASSES .................................................................. 10

3. ABBREVIATIONS.............................................................................................................. 13

4. BACKGROUND .................................................................................................................. 26

4.1 LOW RISK STUDY ......................................................................................................................... 26

4.2 INTERMEDIATE RISK STUDY .................................................................................................... 26

4.3 CLINICAL BACKGROUND .......................................................................................................... 27 4.3.1 Staging ...................................................................................................................................... 27 4.3.2 Stratification of patients: differences from the INRG staging system (INRGSS) ..................... 27

4.3.2.1 Stage L1 – confined to one body compartment ................................................................ 27 4.3.2.2 Age cut-off........................................................................................................................ 27 4.3.2.3 Histopathology ................................................................................................................. 27 4.3.2.4 Definition of bone disease ................................................................................................ 28 4.3.2.5 Definition of a “segmental” genomic profile ................................................................... 28

4.3.3 Results of previous neuroblastoma trials and studies .............................................................. 28 4.3.3.1 Localised resectable disease (LNESG1) ........................................................................... 28 4.3.3.2 Neuroblastoma in infants < 12 months at diagnosis (INES99) ........................................ 29 4.3.3.3 Localised unresectable neuroblastoma in children > 12 months (EUNS) ........................ 30

4.3.4 Chemotherapy........................................................................................................................... 30

4.4 PATHOLOGY .................................................................................................................................. 30

4.5 RADIOTHERAPY ........................................................................................................................... 30

4.6 13-CIS-RETINOIC ACID (13-CIS-RA) .......................................................................................... 31

4.7 BIOLOGY BACKGROUND ........................................................................................................... 32 4.7.1 Recurrent genetic alterations in neuroblastoma ...................................................................... 32 4.7.2 Genetic subtypes in neuroblastoma .......................................................................................... 32

5. AIMS ..................................................................................................................................... 34

5.1 LOW RISK STUDY ......................................................................................................................... 34 5.1.1 Primary aim .............................................................................................................................. 34 5.1.2 Secondary aims ......................................................................................................................... 34

5.2 INTERMEDIATE RISK STUDY .................................................................................................... 34 5.2.1 Primary aim .............................................................................................................................. 34 5.2.2 Secondary aims ......................................................................................................................... 34

6. TREATMENT STRATIFICATION CRITERIA ............................................................. 35

6.1 CLINICAL STAGING ..................................................................................................................... 35 6.1.1 Non-metastatic neuroblastoma ................................................................................................. 35 6.1.2 Metastatic neuroblastoma ........................................................................................................ 35

6.2 NEW CONCEPTS FOR TREATMENT STRATIFICATION ........................................................ 35 6.2.1 Genotyping ............................................................................................................................... 35



6.2.2 International Neuroblastoma Pathology Classification (INPC) .............................................. 36 6.2.3 International Neuroblastoma Risk Group Staging System (INRGSS) ...................................... 36 6.2.4 Image Defined Risk Factors (IDRF) ........................................................................................ 36 6.2.5 Life Threatening Symptoms (LTS) ............................................................................................ 36

7. ELIGIBILITY ..................................................................................................................... 37

8. DIAGNOSTIC EVALUTATION ....................................................................................... 39

8.1 DEFINITION OF DATE OF DIAGNOSIS ..................................................................................... 39

8.2 MANDATORY PRE-TREATMENT STEPS .................................................................................. 39

8.3 EVALUATION AT DIAGNOSIS FOR LOW AND INTERMEDIATE RISK PATIENTS .......... 39 8.3.1 Full history ............................................................................................................................... 39 8.3.2 Full clinical examination (including blood pressure) .............................................................. 39 8.3.3 Haematology/Biochemistry ...................................................................................................... 39 8.3.4 Imaging ..................................................................................................................................... 39 8.3.5 Bone marrow ............................................................................................................................ 40 8.3.6 Pathology and biology of the primary tumour (see Appendix 5 and Appendix 6) .................... 40 8.3.7 Organ function.......................................................................................................................... 40

8.4 LOW RISK STUDY - PRE-TREATMENT INVESTIGATIONS .................................................. 41

8.5 INTERMEDIATE RISK STUDY - PRE-TREATMENT INVESTIGATIONS .............................. 42

9. LOW RISK STUDY: TREATMENT STRATEGY ......................................................... 43

9.1 SUMMARY ..................................................................................................................................... 43 9.1.1 Study groups 1-3, Stage L2, MYCN non-amplified and < 18 months ...................................... 43

9.1.1.1 Study group 1 ................................................................................................................... 43 9.1.1.2 Study group 2 ................................................................................................................... 43 9.1.1.3 Study group 3 ................................................................................................................... 44

9.1.2 Study groups 4-6, Stage Ms, MYCN non-amplified and < 12 months ...................................... 44 9.1.1.4 Study group 4 ................................................................................................................... 44 9.1.1.5 Study group 5 ................................................................................................................... 44 9.1.1.6 Study group 6 ................................................................................................................... 44

9.2 ELIGIBILITY................................................................................................................................... 44

9.3 STUDY DESIGN OVERVIEW ....................................................................................................... 45 9.3.1 Low Risk Study: L2 Patients (≤ 18 months).............................................................................. 45 9.3.2 Low Risk Study: Ms Patients (≤ 12 months) ............................................................................. 46

9.4 TREATMENT STRATEGY FOR LOW RISK PATIENTS ........................................................... 47 9.4.1 Study group 1: randomised study ............................................................................................. 47

9.4.1.1 Study group 1: Treatment Flow Chart – Randomised to Observation ............................. 47 9.4.1.2 Study group 1: Treatment Flow Chart – Randomised to Chemotherapy ......................... 48 9.4.1.3 Study group 1: Road Map ................................................................................................. 49 9.4.1.4 Study group 1: Treatment Strategy – Randomised to Observation .................................. 50 9.4.1.5 Study group 1: Treatment Strategy – Randomised to Chemotherapy .............................. 51

9.4.2 Study group 2 ............................................................................................................................ 54 9.4.2.1 Study group 2: Treatment Flow Chart .............................................................................. 54 9.4.2.2 Study group 2: Road Map ................................................................................................. 55 9.4.2.3 Study group 2: Treatment Strategy ................................................................................... 56


9.4.4 Study group 4 ............................................................................................................................ 62 9.4.4.1 Study group 4: Treatment Flow Chart .............................................................................. 62



9.4.4.2 Study group 4: Road Map ................................................................................................. 63 9.4.4.3 Study group 4: Treatment Strategy ................................................................................... 64



9.4.7 Treatment recommendations for patients without genomic profile result ................................ 72 9.4.8 Chemotherapy and/or surgical procedures outside the recommendations of this protocol ..... 72

10. INTERMEDIATE RISK STUDY: TREATMENT STRATEGY ............................... 73

10.1 SUMMARY ..................................................................................................................................... 73 10.1.1 Study groups 7-8 ....................................................................................................................... 73 10.1.2 Study group 9 ............................................................................................................................ 73 10.1.3 Study group 10 .......................................................................................................................... 73

10.2 ELIGIBILITY................................................................................................................................... 73

10.3 STUDY DESIGN ............................................................................................................................. 75 10.3.1 Intermediate Risk Study: L2 Patients (>18 months)................................................................. 75 10.3.2 Intermediate Risk Study: (L1) INSS 1 ....................................................................................... 76 10.3.3 Intermediate Risk Study: M (≤ 12 months) ............................................................................... 77

10.4 TREATMENT STRATEGY FOR INTERMEDIATE RISK PATIENTS ....................................... 78 10.4.1 Study group 7 ............................................................................................................................ 78

10.4.1.1 Study group 7: Treatment Flow Chart .......................................................................... 78 10.4.1.2 Study group 7: Road Map ............................................................................................. 79 10.4.1.3 Study group 7: Treatment Strategy ............................................................................... 80

Study group 8 ............................................................................................................................................ 81 10.4.2.1 Study group 8: Treatment Flow Chart .......................................................................... 81 10.4.2.2 Study group 8: Road Map ............................................................................................. 82 10.4.2.3 Study group 8: Treatment Strategy ............................................................................... 83

10.4.3 Study group 9 ............................................................................................................................ 85 10.4.3.1 Study group 9: Treatment Flow Chart .......................................................................... 85 10.4.3.2 Study group 9: Road Map ............................................................................................. 86 10.4.3.3 Study group 9: Treatment Strategy ............................................................................... 87

10.4.4 Study group 10 .......................................................................................................................... 88 10.4.4.1 Study group 10: Treatment Flow Chart ........................................................................ 88 10.4.4.2 Study group 10: Road Map ........................................................................................... 89 10.4.4.3 Study group 10: Treatment Strategy ............................................................................. 90

10.4.5 Treatment recommendations for patients with stage L2, age >18 months, without a defined

histotype (i.e. neuroblastoma NOS) .......................................................................................................... 91 10.4.6 Chemotherapy and/or surgical procedures outside the recommendations of this protocol ..... 91

11. INVESTIGATIONS DURING AND AFTER TREATMENT .................................... 92

11.1 MINIMAL MANDATORY INVESTIGATIONS DURING AND AT THE END OF

TREATMENT .............................................................................................................................................. 92

11.2 INVESTIGATIONS AFTER COMPLETION OF TREATMENT ................................................. 93 11.2.1 Tumour Assessment/Detection .................................................................................................. 93 11.2.2 Toxicity Assessment .................................................................................................................. 93

12. LONG TERM FOLLOW UP ......................................................................................... 94

12.1 TUMOUR ASSESSMENT .............................................................................................................. 94



12.2 TOXICITY ASSESSMENT ............................................................................................................. 94 12.2.1 Renal Follow-Up ...................................................................................................................... 94 12.2.2 Auditory Follow-Up .................................................................................................................. 94 12.2.3 Cardiac Follow-Up .................................................................................................................. 94 12.2.4 Etoposide: second malignancy follow up ................................................................................. 94

13. STATISTICAL CONSIDERATIONS ........................................................................... 95

13.1 LOW RISK STUDY ......................................................................................................................... 95 13.1.1 Stage L2 .................................................................................................................................... 95

13.1.1.1 Group 1 ......................................................................................................................... 95 13.1.1.2 Group 2 ......................................................................................................................... 96 13.1.1.3 Group 3 ......................................................................................................................... 96

13.1.2 Stage Ms ................................................................................................................................... 96 13.1.2.1 Group 4 ......................................................................................................................... 96 13.1.2.2 Group 5 ......................................................................................................................... 97 13.1.2.3 Group 6 ......................................................................................................................... 97

13.2 INTERMEDIATE RISK STUDY .................................................................................................... 97 13.2.1 Stage L2 .................................................................................................................................... 97

13.2.1.1 Group 7 ......................................................................................................................... 97 13.2.1.2 Group 8 ......................................................................................................................... 98

13.2.2 INSS stage 1 .............................................................................................................................. 98 13.2.2.1 Group 9 ......................................................................................................................... 98

13.2.3 Stage M ..................................................................................................................................... 98 13.2.3.1 Group 10 ....................................................................................................................... 98

14. NEONATAL SUPRARENAL MASSES: AN OBSERVATIONAL APPROACH.... 99

14.1 BACKGROUND .............................................................................................................................. 99 14.1.1 Review of anatomy and embryology [51] ................................................................................. 99 14.1.2 Perinatal tumours and foetal diagnostic imaging .................................................................... 99 14.1.3 Differential diagnosis ............................................................................................................. 100 14.1.4 Neuroblastoma biology .......................................................................................................... 101

14.1.4.1 Benign behaviour ........................................................................................................ 101 14.1.4.2 Neuroblastoma in the pre and neonatal period [60-65] .............................................. 101 14.1.4.3 Utility of imaging studies and biological markers in the neonatal period .................. 102

14.1.5 Summary ................................................................................................................................. 103 14.1.6 Definitions .............................................................................................................................. 103

14.2 HYPOTHESIS ................................................................................................................................ 104

14.3 TYPE OF STUDY .......................................................................................................................... 104

14.4 AIMS OF THE STUDY ................................................................................................................. 104 14.4.1 Primary aim ............................................................................................................................ 104 14.4.2 Secondary aims ....................................................................................................................... 104

14.5 ELIGIBILITY................................................................................................................................. 104 14.5.1 Inclusion criteria .................................................................................................................... 104 14.5.2 Exclusion criteria ................................................................................................................... 105

14.6 DIAGNOSTIC PROCEDURES ..................................................................................................... 105

14.7 THERAPEUTIC STRATEGIES .................................................................................................... 105

14.8 REGISTRATION AND STUDY MONITORING ........................................................................ 108

14.9 DEFINITION OF AN EVENT....................................................................................................... 109

14.10 STATISTICAL CONSIDERATIONS ....................................................................................... 109

14.11 COLLECTION OF SAMPLES TO STUDY MYCN AMPLIFICATION IN PLASMA [74-76]109



APPENDIX 1. THE INTERNATIONAL NEUROBLASTOMA RISK GROUP (INRG)

STAGING SYSTEM ................................................................................................................. 111

A1.1 MODIFIED INRGSS AS USED IN LINES .............................................................................. 111

A1.2 ORIGINAL INRGSS .................................................................................................................. 111

APPENDIX 2. SUMMARY OF SIOPEN NEUROBLASTOMA TREATMENT STRATEGIES

112

APPENDIX 3. LIFE THREATENING SYMPTOMS ..................................................... 113

APPENDIX 4. LIST OF IMAGE DEFINED SURGICAL RISK FACTORS [7] ......... 114

APPENDIX 5. RESPONSE EVALUATION .................................................................... 115

APPENDIX 6. PATHOLOGY GUIDELINES ................................................................. 116

A6.1 NEEDLE BIOPSIES .................................................................................................................. 116

A6.2 SURGICAL OPEN BIOPSIES AND SURGICAL RESECTIONS ........................................... 116

A6.3 REVIEW PROCESS .................................................................................................................. 117 A6.3.1 Material to be sent .................................................................................................................. 117 A6.3.2 Timeline .................................................................................................................................. 117 A6.3.3 Pathology reviewers ............................................................................................................... 117 A6.3.4 Logistical issues...................................................................................................................... 118

A6.4 PATHOLOGY FORM ............................................................................................................... 119

A6.5 LOCAL PATHOLOGIST AGREEMENT FORM .................................................................... 120

APPENDIX 7. BIOLOGY GUIDELINES ........................................................................ 121

A7.1 MANDATORY INVESTIGATIONS ........................................................................................ 121

A7.2 SCHEDULE FOR RESULTS .................................................................................................... 121

A7.3 TUMOUR MATERIAL FOR BIOLOGICAL STUDIES OF NEUROBLASTOMA ............... 121

A7.4 TUMOUR HANDLING ............................................................................................................. 122

A7.5 RECOMMENDATIONS FOR DETERMINATION OF MYCN STATUS .............................. 122

A7.6 RECOMMENDATIONS FOR MLPA ....................................................................................... 122

A7.7 RECOMMENDATIONS FOR ARRAY-CGH .......................................................................... 123

A7.8 RECOMMENDATIONS FOR SNP-ARRAY ........................................................................... 123

A7.9 CENTRAL REVIEW ................................................................................................................. 124

A7.10 GENOMIC CLASSIFICATION ................................................................................................ 124

APPENDIX 8. IMAGING GUIDELINES ........................................................................ 126

A8.1 INTRODUCTION ...................................................................................................................... 126

A8.2 METHODS OF INVESTIGATION ........................................................................................... 126

A8.3 CT-SCAN TECHNICAL RECOMMENDATIONS .................................................................. 126 A8.3.1 Preparation............................................................................................................................. 126 A8.3.2 CT protocol ............................................................................................................................. 126

A8.4 MRI ............................................................................................................................................ 127 A8.4.1 Preparation, sedation and monitoring ................................................................................... 127 A8.4.2 MRI protocol .......................................................................................................................... 127



A8.5 TUMOUR MEASUREMENTS ................................................................................................. 128

A8.6 RELATIONSHIP BETWEEN THE PRIMARY TUMOUR AND NEIGHBOURING VITAL

STRUCTURES .......................................................................................................................................... 129 A8.6.1 “Separation” .......................................................................................................................... 129 A8.6.2 “Contact” and related terms .................................................................................................. 129 A8.6.3 “Encasement” and related terms ........................................................................................... 129 A8.6.4 “Infiltration” .......................................................................................................................... 129 A8.6.5 Compartments, uni- and multifocality .................................................................................... 129 A8.6.6 Effusions ................................................................................................................................. 130

A8.7 IDRF ASSESSMENT ACCORDING TO ANATOMIC LOCATION ...................................... 130 A8.7.1 Neck ........................................................................................................................................ 130 A8.7.2 Thorax .................................................................................................................................... 130 A8.7.3 Abdomen ................................................................................................................................. 131 A8.7.4 Pelvis ...................................................................................................................................... 131

APPENDIX 9. GUIDELINE FOR RADIOIODINATED MIBG SCINTIGRAPHY IN

CHILDREN 132

A9.1 PURPOSE .................................................................................................................................. 132

A9.2 BACKGROUND INFORMATION AND DEFINITIONS ........................................................ 132

A9.3 COMMON INDICATIONS ....................................................................................................... 132 A9.3.1 Indications .............................................................................................................................. 132 A9.3.2 Contra indications .................................................................................................................. 133

A9.4 PROCEDURE ............................................................................................................................ 133 A9.4.1 Information about previous examinations relevant to this procedure .................................... 133 A9.4.2 Patient preparation ................................................................................................................ 133

A9.4.2.1 Information with appointment letter: .......................................................................... 133 A9.4.2.2 Thyroid blockade ........................................................................................................ 133 A9.4.2.3 Drugs interactions ....................................................................................................... 133

A9.4.3 Precautions ............................................................................................................................. 134 A9.4.4 Radiopharmaceutical ............................................................................................................. 134

A9.4.4.1 Radionuclide ............................................................................................................... 134 A9.4.4.2 Pharmaceutical ........................................................................................................... 134 A9.4.4.3 Dose Schedule (see A9.5) ........................................................................................... 134 A9.4.4.4 Injection technique ..................................................................................................... 134 A9.4.4.5 Radiation Burden ........................................................................................................ 134

A9.4.5 Image Acquisition ................................................................................................................... 134 A9.4.5.1 Timing for imaging ..................................................................................................... 135 A9.4.5.2 Collimator ................................................................................................................... 135 A9.4.5.3 Positioning of the child ............................................................................................... 135 A9.4.5.4 Views .......................................................................................................................... 135 A9.4.5.5 Computer acquisition set up ....................................................................................... 135 A9.4.5.6 Optional images .......................................................................................................... 135

A9.4.6 Intervention............................................................................................................................. 136 A9.4.7 Processing .............................................................................................................................. 136 A9.4.8 Hard copy output .................................................................................................................... 136 A9.4.9 Interpretation, reporting and pitfalls ...................................................................................... 136

A9.4.9.1 Normal biodistribution ............................................................................................... 136 A9.4.9.2 Pathology .................................................................................................................... 136 A9.4.9.3 False negative scans.................................................................................................... 136

A9.4.10 Quality control.................................................................................................................... 136

A9.5 ISSUES REQUIRING FURTHER CLARIFICATION ............................................................. 137

APPENDIX 10. SURGICAL GUIDELINES ...................................................................... 138



A10.1 INTRODUCTION ...................................................................................................................... 138

A10.2 AIMS OF SURGERY ................................................................................................................ 138

A10.3 SURGICAL PROCEDURES AT PRESENTATION ................................................................ 138

A10.4 SUBSEQUENT/SECONDARY SURGICAL PROCEDURES ................................................. 139 A10.4.1 Excision of primary mass should be attempted for: ........................................................... 139 A10.4.2 Excision of primary mass should be discussed with the surgical coordinators for: .......... 139 A10.4.3 Excision of primary mass is not recommended for: ........................................................... 140

A10.5 TIMING OF SUBSEQUENT PROCEDURES (WHERE APPLICABLE) ............................... 140 A10.5.1 Neonatal suprarenal masses (NSM): an observational approach...................................... 140 A10.5.2 Low risk study ..................................................................................................................... 140 A10.5.3 Intermediate risk study ....................................................................................................... 140

A10.6 DEFINITIONS ........................................................................................................................... 140 A10.6.1 Complete Excision .............................................................................................................. 140 A10.6.2 Excision with minimal residual disease (<5% of original or <5ml volume) ..................... 140 A10.6.3 Incomplete gross excision ................................................................................................... 140 A10.6.4 Biopsy ................................................................................................................................. 140

A10.7 SURGICAL PROCEDURES ..................................................................................................... 141 A10.7.1 Biopsy (please refer also to the pathology guidelines) ....................................................... 141 A10.7.2 Surgical approach for tumour resection ............................................................................ 141

A10.7.2.1 Abdominal tumours .................................................................................................... 141 A10.7.2.2 Pelvic tumours ............................................................................................................ 141 A10.7.2.3 Thoracic/cervical tumours .......................................................................................... 141

A10.7.3 Lymph Node Evaluation ..................................................................................................... 142 A10.7.4 Intraspinal Extension .......................................................................................................... 142 A10.7.5 Intraspinal disease with neurological symptoms. ............................................................... 142 A10.7.6 Nephrectomy ....................................................................................................................... 142 A10.7.7 Tumour Relation with Great Vessels. ................................................................................. 142 A10.7.8 Risk factors encountered at operation ................................................................................ 142 A10.7.9 Clips .................................................................................................................................... 142

A10.8 DEFINITION OF MAJOR SURGICAL COMPLICATIONS ................................................... 142

APPENDIX 11. CHEMOTHERAPY GUIDELINES ........................................................ 143

A11.1 CO (CYCLOPHOSPHAMIDE AND VINCRISTINE) ............................................................. 143

A11.2 VP/CARBO (ETOPOSIDE {VP16} AND CARBOPLATIN) .................................................. 143

A11.3 CADO (CYCLOPHOSPHAMIDE, DOXORUBICIN AND VINCRISTINE) .......................... 144

A11.4 DOSE MODIFICATION ........................................................................................................... 144

APPENDIX 12. RADIOTHERAPY GUIDELINES .......................................................... 145

A12.1 INDICATIONS .......................................................................................................................... 145 A12.1.1 Indication ............................................................................................................................ 145 A12.1.2 No indication ...................................................................................................................... 145

A12.2 TREATMENT CENTRES ......................................................................................................... 145

A12.3 TIMING OF RADIOTHERAPY................................................................................................ 145

A12.4 ASSESSMENT........................................................................................................................... 145

A12.5 INFORMED CONSENT ............................................................................................................ 146

A12.6 IMMOBILISATION AND POSITIONING .............................................................................. 146

A12.7 IMAGING, PLANNING AND TARGET VOLUME DEFINITION ........................................ 146



A12.8 PROSPECTIVE QUALITY CONTROL ................................................................................... 148

A12.9 ENERGY, DOSE AND FRACTIONATION ............................................................................ 148

A12.10 NORMAL TISSUE TOLERANCE........................................................................................ 149 A12.10.1 Kidneys ............................................................................................................................... 149 A12.10.2 Liver .................................................................................................................................... 149 A12.10.3 Lungs .................................................................................................................................. 149 A12.10.4 Bone .................................................................................................................................... 149 A12.10.5 Spinal cord.......................................................................................................................... 149 A12.10.6 Bowel .................................................................................................................................. 150 A12.10.7 Bladder ............................................................................................................................... 150 A12.10.8 Heart ................................................................................................................................... 150 A12.10.9 Gonads ................................................................................................................................ 150

A12.11 SUPPORTIVE CARE ............................................................................................................ 150

A12.12 GEOMETRIC VERIFICATION ............................................................................................ 150

A12.13 DOCUMENTATION ............................................................................................................. 151

A12.14 FOLLOW-UP ......................................................................................................................... 152

APPENDIX 13. SUPPORTIVE CARE ............................................................................... 153

A13.1 ANTI-EMETICS ........................................................................................................................ 153

A13.2 HYDRATION ............................................................................................................................ 153

A13.3 BLOOD COMPONENT THERAPY ......................................................................................... 153

A13.4 RED BLOOD CELLS ................................................................................................................ 153

A13.5 PLATELETS .............................................................................................................................. 153

A13.6 CENTRAL LINES ..................................................................................................................... 153

A13.7 TREATMENT GUIDELINES FOR FEBRILE NEUTROPENIA ............................................ 153

A13.8 PNEUMOCYSTIS PNEUMONITIS PROPHYLAXIS ............................................................. 154

A13.9 NUTRITION .............................................................................................................................. 154

A13.10 RENAL FUNCTION MONITORING ................................................................................... 154

APPENDIX 14. SPINAL CORD COMPRESSION ........................................................... 155

A14.1 SPINAL CORD COMPRESSION – BACKGROUND ............................................................. 155

A14.2 DIAGNOSIS AND EVALUATION OF SPINAL CORD COMPRESSION ............................ 157

A14.3 TREATMENT OF SPINAL CORD COMPRESSION .............................................................. 157

A14.4 ASYMPTOMATIC INTRASPINAL RESIDUAL TUMOUR FOLLOWING

CHEMOTHERAPY ................................................................................................................................... 158

APPENDIX 15. TOXICITY ................................................................................................. 159

APPENDIX 16. DRUG INFORMATION ........................................................................... 162

A16.1 CARBOPLATIN ........................................................................................................................ 162

A16.2 CYCLOPHOSPHAMIDE .......................................................................................................... 163

A16.3 DOXORUBICIN ........................................................................................................................ 164

A16.4 ETOPOSIDE (VP16) .................................................................................................................. 165

A16.5 VINCRISTINE ........................................................................................................................... 166



A16.6 13-CIS-RETINOIC ACID (ISOTRETINOIN®, ROACCUTANE®) ....................................... 167

APPENDIX 17. DIFFERENTIATION THERAPY WITH 13-CIS RETINOIC ACID . 169

A17.1 INDICATION............................................................................................................................. 169

A17.2 TREATMENT SCHEDULE FOR 13-CIS-RA .......................................................................... 169

A17.3 SUGGESTED SUPPORTIVE CARE ........................................................................................ 169

A17.4 CRITERIA PRIOR TO EACH CYCLE OF 13-CIS-RA ........................................................... 170

A17.5 DOSE MODIFICATIONS ......................................................................................................... 170

APPENDIX 18. ORGANISATIONAL AND ADMINISTRATIVE ISSUES ................... 171

A18.1 ETHICAL AND LEGAL ASPECTS AND AGREEMENTS .................................................... 171 A18.1.1 Good Clinical Practice/Declaration of Helsinki ................................................................ 171 A18.1.2 Sponsorship ........................................................................................................................ 171 A18.1.3 Insurance ............................................................................................................................ 171 A18.1.4 Ethics committee (EC)/Institutional Review Board (IRB) approval ................................... 172 A18.1.5 Competent Health Authority approval ............................................................................... 172 A18.1.6 Financing ............................................................................................................................ 172 A18.1.7 Informed consent ................................................................................................................ 172

A18.2 DATA MANAGEMENT AND PROTECTION/CONFIDENTIALITY ................................... 173 A18.2.1 Data Collection/Submission ............................................................................................... 173 A18.2.2 Registration of a new patient .............................................................................................. 173 A18.2.3 Randomisation .................................................................................................................... 173 A18.2.4 Data protection/confidentiality........................................................................................... 174 A18.2.5 Data storage ....................................................................................................................... 174 A18.2.6 Data transfer ...................................................................................................................... 174

A18.3 SAFETY PHARMACOVIGILANCE INSTRUCTIONS .......................................................... 174

A18.4 QUALITY CONTROL AND QUALITY ASSURANCE ......................................................... 174 A18.4.1 Data monitoring committee ................................................................................................ 174 A18.4.2 Monitoring .......................................................................................................................... 175 A18.4.3 Audits .................................................................................................................................. 175 A18.4.4 Inspections .......................................................................................................................... 175

A18.5 ADMINISTRATIVE ISSUES .................................................................................................... 175 A18.5.1 Protocol compliance ........................................................................................................... 175 A18.5.2 Protocol amendments ......................................................................................................... 175 A18.5.3 End of the study .................................................................................................................. 176 A18.5.4 Publication rules ................................................................................................................ 176

A18.6 CENTRAL REVIEW ................................................................................................................. 176 A18.6.1 Pathology ............................................................................................................................ 176 A18.6.2 Biology ................................................................................................................................ 176

A18.7 REMOVAL FROM TRIAL TREATMENT .............................................................................. 176

A18.8 FOLLOW-UP ............................................................................................................................. 176

APPENDIX 19. GUIDELINES FOR REPORTING TOXICITY AND SERIOUS ADVERSE

EVENTS (SAES) 178

A19.1 DEFINITIONS ........................................................................................................................... 178 A19.1.1 Adverse Event (AE) ............................................................................................................. 178 A19.1.2 Adverse Reaction (AR) ....................................................................................................... 178 A19.1.3 Unexpected Adverse Reaction (UAR) ................................................................................. 178 A19.1.4 Serious Adverse Events (SAE) or Reaction (SAR) .............................................................. 179 A19.1.5 Suspected Expected Serious Adverse Reaction (SESAR) .................................................... 179



A19.1.6 Suspected Unexpected Serious Adverse Reaction (SUSAR) ............................................... 179

A19.2 DOCUMENTATION AND REPORTING OF EVENTS .......................................................... 179

A19.3 EXPECTED SAES WHICH ARE EXEMPT FROM EXPEDITED REPORTING .................. 179

A19.4 REPORTING PROCEDURES AND TIME LIMITS ................................................................ 179 A19.4.1 Local investigator responsibilities...................................................................................... 179 A19.4.2 Sponsor responsibilities ...................................................................................................... 180

A19.5 CAUSALITY ASSESSMENT ................................................................................................... 181

APPENDIX 20. SERIOUS ADVERSE EVENT REPORT FORM .................................. 182

APPENDIX 21. PARENT AND PATIENT INFORMATION .......................................... 186

A21.1 REGISTRATION INTO LINES FOR LOW AND INTERMEDIATE RISK STUDY

GROUPS 186 A21.1.1 Information for parents ...................................................................................................... 186 A21.1.2 Informed consent for parents (all study groups) ................................................................ 188

A21.2 LOW RISK STUDY ................................................................................................................... 190 A21.2.1 Information for parents ...................................................................................................... 190 A21.2.2 Informed consent for parents (all low risk study groups with a genomic profile) .............. 197 A21.2.3 Informed consent for parents (all low risk study groups without a genomic profile) ........ 198 A21.2.4 Informed consent for parents (additional for study group 1) ............................................. 199

A21.3 INTERMEDIATE RISK STUDY .............................................................................................. 200 A21.3.1 Information for Parents ...................................................................................................... 200 A21.3.2 Informed consent for parents (all IR study groups)............................................................ 205 A21.3.3 Informed consent for parents (IR patients without a defined histotype)............................. 206 A21.3.4 Information for patients over 6 years ................................................................................. 207

A21.4 SAMPLE OF CONSENT FORM FOR RADIOTHERAPY ...................................................... 209

A21.5 NEONATAL SUPRARENAL MASSES: AN OBSERVATIONAL APPROACH .................. 212 A21.5.1 Information sheet for parents ............................................................................................. 212 A21.5.2 Informed consent for parents .............................................................................................. 216

APPENDIX 22. CONTACT INFORMATION .................................................................. 217

A22.1 PRINCIPAL INVESTIGATORS ............................................................................................... 217

A22.2 NATIONAL COORDINATORS ............................................................................................... 217

A22.3 DATA MONITORING COMMITTEE...................................................................................... 218

A22.4 BIOLOGY COMMITTEE ......................................................................................................... 219

A22.5 NUCLEAR MEDICINE AND PHYSIC COMMITTEE COORDINATOR ............................. 223

A22.6 BONE MARROW COMMITTEE COORDINATOR ............................................................... 223

A22.7 PATHOLOGY COMMITTEE ................................................................................................... 223

A22.8 RADIOLOGY COMMITTEE .................................................................................................... 224

A22.9 RADIOTHERAPY COMMITTEE ............................................................................................ 224

A22.10 STATISTICS COMMITTEE ................................................................................................. 225

A22.11 SURGERY COMMITTEE ..................................................................................................... 225

APPENDIX 23. DECLARATION OF HELSINKI ............................................................ 226

A23.1 INTRODUCTION ...................................................................................................................... 226

A23.2 PRINCIPLES FOR ALL MEDICAL RESEARCH ................................................................... 227



A23.3 ADDITIONAL PRINCIPLES FOR MEDICAL RESEARCH COMBINED WITH MEDICAL

CARE 229

APPENDIX 24. REFERENCE LIST .................................................................................. 231



4. BACKGROUND

The European study, LINES 2009 (Low and Intermediate Risk Neuroblastoma European Study),

groups together in a single protocol the treatment of all patients with “non high risk”

neuroblastoma (NB), with stratification into two groups: low risk and intermediate risk. These

two separate cohorts are included in one single protocol to enable patient data from these two

groups to be entered into a common database, as the current prognostic classifications determining

treatment may evolve further with subsequent more detailed molecular analysis of the tumours.

4.1 Low Risk Study

The low risk group of patients includes NB patients without MYCN amplification with or without

life threatening symptoms in the following clinical situations:

Children aged ≤ 18 months with localised neuroblastoma associated with image defined

risk factors precluding upfront surgery (stage INRG L2).

Children aged ≤ 12 months with disseminated neuroblastoma without bone, pleura, lung or

CNS disease (stage INRG Ms)

The Low Risk Study is proposed in order to:

minimise the amount of treatment (chemotherapy and surgery) for all appropriate low risk

patients, who in previous studies have been shown to have an excellent long-term outcome

(as in the SIOPEN 99.1-2 infant neuroblastoma studies where the overall survival was

greater than 97%, personal communication H. Rubie, JCO in press).

improve the EFS and maintain the OS in L2 and Ms patients with a SCA genomic profile

tumour (presence of any segmental chromosomal change (SCA)) by electively treating

these patients with chemotherapy despite the absence of symptoms.

4.2 Intermediate Risk Study

The intermediate risk group of patients includes NB patients in the following clinical situations:

Children aged >18 months with localised neuroblastoma without MYCN amplification,

associated with image defined risk factors precluding upfront surgery (stage INRG L2).

Children aged ≤12 months with disseminated neuroblastoma involving bone, pleura, lung

and/or CNS (stage INRG M), without MYCN amplification.

Children with localised resected NB (stage INSS I) with MYCN amplification.

The Intermediate Risk Study is proposed in order to:

reduce the amount of chemotherapy for differentiating histology INRG stage L2 NB and

ganglioneuroblastoma nodular patients who in previous SIOPEN study have been shown

to have an excellent long-term outcome;

increase the amount of treatment (radiotherapy and 13-cis-RA) for poorly differentiated or

undifferentiated histology INRG stage L2 NB or ganglioneuroblastoma nodular patients in

order to improve the EFS registered in the previous SIOPEN study;

improve the EFS of MYCN amplified INSS stage 1 NB patients with the introduction of

adjuvant treatment;

maintain the very good results obtained in previous SIOPEN study for INRG stage M

infants with a moderate treatment.



4.3 Clinical Background

4.3.1 Staging

Neuroblastoma (NB) is the most frequent extracranial solid tumour of childhood, representing 8 to

10 % of all cases of childhood cancer. This cancer is characterised by a heterogeneous clinical

presentation and outcome. In the past, in Europe, treatment was stratified according to tumour

stage defined by the International Neuroblastoma Staging System [1], an age cut-off at twelve

months and the MYCN status. Recently, the new international classification for these tumours

(International Neuroblastoma Risk Group or INRG) takes into account the clinical heterogeneity

and provides the clinicians with a powerful and robust tool for treatment stratification based on

the clinical parameters age (≤ or > 18 months) and stage (localised tumour: L or metastatic

tumour: M), the presence or absence of image defined risk factors for a surgical removal of a

tumour (no image defined risk: L1; image defined risk: L2), as well as the molecular data

resulting from the analysis of tumour tissue including the status of the proto-oncogene MYCN and

the presence or absence of segmental chromosomal abnormalities (SCA) while presently taking

into account only a small number of the alterations recurrently observed in NB (deletion of

chromosome arm 1p or 11q) [2].

4.3.2 Stratification of patients: differences from the INRG staging system (INRGSS)

The previous SIOPEN studies for patients with low or intermediate risk NB, particularly the INES

and EUNB studies, have provided clinicians with important data which has been taken into

account in the design of this new protocol. Thus the stratification of patients in this protocol

differs slightly from the INRGSS.

The main differences between the INRGSS and the stratification of patients in LINES are:

4.3.2.1 Stage L1 – confined to one body compartment

In the staging of L1 tumours the INRGSS states that a localised tumour must not involve the vital

structures as defined by the list of image defined risk factors (IDRFs) and be confined to one body

compartment [2]. However, it has since been acknowledged that localised tumours without

IDRFs are L1 tumours regardless of regional location (personal communication Dr. A. Pearson,

2010).

4.3.2.2 Age cut-off

Age cut-off at 12 months instead of 18 months for patients with metastatic disease as proposed by

the INRGSS [2]. Within SIOPEN, patients with metastatic NB aged 12-18 months are currently

recruited into the SIOPEN protocol for high risk NB. In this ongoing trial, data is blinded and,

consequently, we cannot yet verify if the patients included in the high risk protocol aged 12 – 18

months (with favourable biology i.e. absent MYCN amplification) have a better outcome than

older patients. Furthermore, detailed biological analyses are ongoing to enable a critical inspection

of the tumour biological profile of these patients. Therefore, it is felt that currently existing data

does not allow a down-staging of these patients into the intermediate risk group. Once the

complete data is available there will be further discussions about this age cut-off.

4.3.2.3 Histopathology

The INRGSS considers patients with L2 NB over 18 months with poorly differentiated and

undifferentiated histology as a “high-risk” group. We will treat them as “intermediate risk”, as in

the past, but with a more intensive treatment than before. Preliminary results from SIOPEN

EUNB show an EFS of 62% in cases with poorly differentiated or undifferentiated histology. The

majority of children were >18 months with poorly differentiated and undifferentiated NB. There

were also a very small number of children <18 months with undifferentiated NB or high MKI. For



this reason we are keeping these patients in an “intermediate risk” protocol instead of giving them

a more aggressive treatment such as high dose chemotherapy.

4.3.2.4 Definition of bone disease

According to INRGSS, MIBG uptake in the skeleton is a condition for considering a patient to be

a stage M NB, even in infants < 12 months. However, SIOPEN has recently demonstrated, that

this uptake should be confirmed by X-ray or CT in order to consider it as a true bone metastasis in

infants < 12 months. If a bony metastasis is not confirmed by radiology the child should be

observed, as in stage Ms, with an excellent chance of cure (>95%) [3].

4.3.2.5 Definition of a “segmental” genomic profile

INRG criteria enable the classification of nearly all neuroblastic tumours, but do not yet take into

account data from recent pan-genomic studies based on array-CGH indicating that any of the

segmental chromosomal alterations recurrently observed in NB are associated with a poorer

prognosis, in addition to the alterations cited in the INRGSS [4]. Indeed, INRGSS takes into

account the status at chromosome 1p or 11q, but not segmental alterations occurring at other

recurrently altered chromosome regions in neuroblastoma. However, additional analysis of INRG

data clearly demonstrates that an occurrence of segmental alterations at 1p, 11q and/or 17q can be

combined to define a “segmental” genomic profile with higher prognostic impact than 1p or 11q

status only (Schleiermacher, Couturier, oral presentation ANR2008 Congress; Tokyo, Japan, May

2008). Furthermore, recent publications have clearly demonstrated the prognostic impact of any

segmental alteration in NB [4, 5]. Until recently no reliable, robust and easily accessible technique

was available to define the status in all recurrently involved chromosome arms in a clinical

setting. The advent of the MLPA technique now enables an analysis of all neuroblastic tumours to

define a multi-genomic profile whenever array-CGH is not available. Thus it is justified to take

into account all recurrently involved chromosome regions in the definition of a “segmental”

genomic type in NB, which are associated with this higher risk of relapse. This “multi or pan-

genomic profile” defined as above will be used for the stratification of low risk patients in this

study. The prognostic impact of alterations of chromosome regions not recurrently involved in NB

will be studied prospectively.

Information that is relevant to the deviations of staging used in this protocol when compared to

the INRGSS staging system (age cut-off, definition of bone disease in infants < 12 months,

definition of a “segmental” genomic profile) will all be recorded in the CRFs to enable an analysis

of the data according to the INRG criteria.

4.3.3 Results of previous neuroblastoma trials and studies

4.3.3.1 Localised resectable disease (LNESG1)

Over the past 15 years, the European group SIOPEN has developed a series of clinical treatment

trials covering the whole clinical spectrum of neuroblastoma patients. The LNESG1 study has

shown the feasibility of treatment by surgical resection only in patients of any age with localised

resectable disease [6-8]. Guidelines were established indicating which patients should be

operated on immediately and which should undergo surgery after tumour reduction with

chemotherapy by defining surgical risk factors based on the imaging characteristics, with a

validation of surgical risk factors as predictors of adverse surgical outcome in this series of 719

patients.

However, a recently completed analysis from the International Neuroblastoma Risk Group

(INRG) database showed less favourable EFS and OS in patients with MYCN amplified INSS

stage 1 and 2 tumours than in patients with non-amplified tumours (53%+8% and 72%+7% vs.



90%+1% and 98%+1%, p<0.0001 respectively). Within this cohort of patients with MYCN

amplification, both EFS and OS were statistically significantly higher for patients whose tumours

were hyperdiploid rather than diploid (EFS: 82%+20% vs. 37%+21% (p=0.0069) and OS:

94%+11% vs. 54%+15% (p=0.0056), respectively). No other clinical, pathological or biological

variable evaluated or initial treatment approach had prognostic significance in this cohort.

However, the retrospective analysis and the very small patient number concerned doesn’t allow

consideration of a treatment reduction for the subgroup of patients with INSS stage 1 MYCN

amplified tumours [9].

4.3.3.2 Neuroblastoma in infants < 12 months at diagnosis (INES99)

The following studies concerned infants aged less than 12 months at diagnosis.

Infants with localised unresectable disease without MYCN amplification were eligible for the

INES99.1 study. Among 120 patients included in the study, 79 without threatening symptoms

received up front chemotherapy consisting of Cyclophosphamide/Vincristine, 48 infants of whom

then required second line chemotherapy because of no response, progression or surgical risk

remained. Surgery was attempted in 105 patients including 22 after CV alone. Five years survival

and 5 years event-free survival were 99% +/- 1% and 90% +/- 3% with a median follow up of 6.1

years (range 1.6-9.1). Primary low-dose chemotherapy without anthracyclines is efficient in 61%

of infants presenting with an unresectable NB and no MYCN amplification, allowing excellent

survival rates without jeopardizing their long-term outcome (personal communication H. Rubie,

manuscript in preparation).

On the assumption that most infants with disseminated neuroblastoma without MYCN

amplification have a favourable prognosis, two concomitant prospective trials (INES99.2 and

INES99.3) were started in which up-front chemotherapy by VP/Carbo was given to patients

presenting life- or organ-threatening symptoms or overt metastases to skeleton, lung, or CNS.

Surgery was to be performed only in the absence of surgical risk factors. One hundred seventy

infants with disseminated neuroblastoma without MYCNA, diagnosed between June 1999 and

June 2004 in nine European countries were eligible for either of the two studies. The 125 infants

treated on trial 99.2 had a 2-year overall survival (OS) of 97.6% with no difference between

asymptomatic and symptomatic patients (97.7% v 97.3%), patients without or with unresectable

primary tumours (96.8% v 100%), and patients without or with positive skeletal scintigraphy

without radiologic abnormalities (97.2% v 100%). The 45 infants treated on trial 99.3 had a 2-year

OS of 95.6%. No patients died of surgery- or chemotherapy-related complications. Infants with

disseminated disease without MYCN amplification have excellent survival with minimal or no

treatment. Asymptomatic infants with an unresectable primary tumour or positive skeletal

scintigraphy without radiologic abnormalities may undergo observation alone [3].

Finally, among infants with MYCN amplification enrolled in the INES99.4 study, induction

chemotherapy using conventional drugs resulted in non response or disease progression in 30% of

patients, and event free and overall survival were poor despite megatherapy, justifying new

therapeutic approaches in this high risk population [10].

We now aim to demonstrate the feasibility of a reduction in treatment for those infants with the

best prognosis defined by clinical and pangenomic criteria who present with image defined risk

factors and therefore cannot benefit from initial surgical resection. This hypothesis is strongly

supported by the excellent results reported from the German group who in a non-randomised

study reduced chemotherapy in certain situations, thus showing that these selected patients do not

require chemotherapy [11].



4.3.3.3 Localised unresectable neuroblastoma in children > 12 months (EUNS)

The SIOPEN European Unresectable Neuroblastoma Study (EUNS) was open from 2001-2006,

and recruited 164 eligible patients aged more than 12 months with MYCN non-amplified

unresectable NB (presence of Image-Defined-Risk-Factors). All patients were to receive 6 courses

of chemotherapy (3 of carboplatin-etoposide and 3 of CADO) and a surgical resection of the

primary tumour. The 5-year Overall Survival (OS) and Event-Free Survival (EFS) of the whole

group were respectively 79% and 68%. Age was confirmed to be the most important prognostic

factor for OS and EFS. Both OS and EFS significantly improved in younger patients (age 12-18

months) vs. older patients (OS 100% vs. 70%, p=.006, and EFS 78% vs. 62%, p=0.001).

Histology also (central review in 127/164 cases) proved to be of significant prognostic value.

Histological subtyping, obtained in 99 patients, identified 17 patients with differentiating NB and

82 with poorly differentiated or undifferentiated NB. OS was 100% in differentiating NB cases vs.

72% in poorly differentiated or undifferentiated NB cases (p=0.13). 104 patients were classified

according to INPC histoprognostic categories, and 46% had differentiating histology while 54%

had poorly differentiated or undifferentiated histology. OS in the “differentiating” histology group

reached 100% vs. 63% in the “poorly differentiated or undifferentiated” cases (p=0.004), and EFS

was 91% vs. 62% (p=0.002). These results comply with the INRG conclusions [2].

Accordingly, patients with INRG stage L2 NB aged less than or equal 18 months, in this protocol

will be considered as having a “low risk” NB and will be treated with less chemotherapy than in

the past or will be randomised for observation only. While stage L2 NB older than 18 months

patients will be considered as harbouring an “intermediate risk” NB. Those patients with

differentiating NB will be treated according to the histological subtype with less chemotherapy (4

courses). Patients with poorly differentiated/undifferentiated NB will receive a more aggressive

schedule containing 6 courses of chemotherapy followed by local radiotherapy and 6 courses of

13-cis-RA.

4.3.4 Chemotherapy

For the present LINES protocol, known chemotherapy associations will be administered as first

line treatment. Cyclophosphamide/Vincristine (CO) has been shown to be effective in infants with

localised unresectable disease without life threatening symptoms, with a very good clinical

tolerance even in small infants. Carboplatin/etoposide (VP/Carbo) has been shown to be effective

in infants both with a localised unresectable NB with symptoms, or in disseminated disease

including metastasis to skeleton, lung or CNS. Finally, cyclophosphamide-doxorubicin-

vincristine (CADO) has been given in combination with VP/Carbo to older children with localised

unresectable disease with an acceptable toxicity and a high response rate.

4.4 Pathology

The SIOPEN Pathology Speciality Committee focused on the histopathological review of tumours

enrolled in the protocol for European unresectable neuroblastoma (EUNS). By the end of August

2008, 127 tumours had been reviewed. Of these, 122 were histologically evaluable and classified

morphologically and prognostically according to the International Neuroblastoma Pathology

Classification (INPC). Data collected during the review included not only the INPC

histoprognosis (Differentiating/Poorly differentiated or undifferentiated), but also INPC

histological category and subtype, because these are elements of the recently developed INRG

pre-treatment risk grouping scheme. This justifies a prospective application of INPC for

treatment stratificaiton of patients with localised unresectable disease (INRG stage L2) over 18

months of age.

4.5 Radiotherapy

Before the start of SIOPEN EUNS protocol, the use of radiotherapy in patients with unresectable

NB in European countries was very limited.



The French Group reported a series of patients with unresectable NB, of whom 34 were over 1

year of age and MYCN non-amplified [12]. They were treated with primary chemotherapy,

surgery and additional chemotherapy in case of post-operative residue. Only 4 patients with

persisting macroscopic residue received subsequent local irradiation. OS and EFS of those 34

patients were respectively 94% 8% and 81% 14%.

The Spanish Group reported the treatment of 33 children with Evans Stage III neuroblastoma

over the age of 1 year, consisting of intensive induction chemotherapy, surgery and maintenance

chemotherapy for 14 months [13]. Radiotherapy was given to patients with residual tumour.

Actuarial survival at 48 months was 60%. In the following study (1992-1998) radiotherapy was

not included.

The Italian Group[14] reported 145 cases of localised unresectable NB treated with initial

chemotherapy, surgical resection and consolidation therapy. They concluded that children with

residual tumour after surgery did not gain an advantage from the addition of radiation therapy. In

a subsequent paper[15] describing a similar group of patients, they reported that only 2/43 patients

received local radiotherapy.

US experience: Castleberry et al reported improved local control and overall survival data from a

randomised trial evaluating radiotherapy in children aged over 1 year with POG stage C disease

[16]. Modak et al. (Memorial Sloan-Kettering) published a monoinstitutional series of INSS stage

3 NB treated with surgery only or surgery and standard chemotherapy obtaining an EFS of 74.9%

and an OS of 92.6% [17]. Park et al. published a series of COG high risk INSS stage 3 NB

including MYCN amplified cases but also unfavourable histology cases without MYCN

amplification, all treated with a high risk regimen including radiotherapy [18]. Their 5-year EFS

and OS were respectively 55% and 59%.

Preliminary data of SIOPEN EUNS, with surgery only as local treatment, report 30 cases of

relapse (18%), involving the primary site in 28 cases (19 isolated and 9 combined with metastatic

sites).

In conclusion, radiotherapy for patients with unresectable NB has not consistently demonstrated a

clear improvement in survival. However, taking into account the rate of local failure in EUNS and

the fact that there are not enough patients to do a randomised study, the “intermediate risk” study

considers radiotherapy as part of the scheduled regimen for patients with poorly differentiated or

undifferentiated histotypes.

4.6 13-cis-retinoic acid (13-cis-RA)

The vitamin A metabolite, 13-cis-RA, has been shown to have therapeutic activity in

neuroblastoma by inducing differentiation and/or growth arrest. Several phase 3 studies have

demonstrated an improvement in OS and EFS for high-risk neuroblastoma, especially stage 4.

Nevertheless, its role is not so clearly demonstrated for stage 3 patients. 72 INSS stage 3 patients

were included in the randomised study with 13-cis-RA versus non-therapy for minimal residual

disease treatment [19]: 1/3 of them were MYCN amplified cases and the other non-amplified cases

that were included in the study either had unfavourable histology and/or increased ferritin. The

results of this heterogeneous group has been recently conveyed [18], showing a 5y-OS and EFS of

55% and 59%, respectively. The 23 patients randomised to 13-cis-RA therapy show a 5y OS and

EFS of 78% and 70% respectively, better than those with no treatment (5y OS and EFS: 67% and

63% (p=0.55). For the group randomised to both ABMT and retinoic therapy, which represents

the best responding patients to the designed therapeutic strategic, the 5y OS and EFS reached



100% and 80% respectively. The pitfalls of this study are the scarce number of patients and its

heterogeneity, making it difficult to find out the real benefit of megatherapy and 13-cis-RA

therapy in this population.

The preliminary results of the last European Study for Unresectable neuroblastoma (EUNS) have

shown a 5-year OS and EFS of 79% and 68%, with conventional chemotherapy and delayed

surgery. Age (p=0.001) and histology (p=0.002) proved to be of significant prognostic value, in

identifying a subgroup of patients (older than 18 months and unfavourable histology) with a worse

outcome. The fact that 11 out of the 30 relapses were metastatic (2 isolated and 9 both local and

metastatic) could mean that there is a need not only to intensify local treatment but also to

introduce a maintenance treatment for minimal residual disease that could persist after

chemotherapy, surgery and radiotherapy. Our hypothesis is that 13-cis-RA could prevent

metastatic relapses, avoiding the intensity and toxicity of megatherapy in a non-amplified MYCN

population.

4.7 Biology Background

4.7.1 Recurrent genetic alterations in neuroblastoma

A large number of recurrent genetic alterations have been identified in NB [20-23]. Amplification

of the MYCN oncogene is observed in approximately 20% of cases and is clearly associated with a

poor outcome [24]. Variations of ploidy status have also been described in NB, with near-triploidy

associated with an excellent outcome, and diploidy/tetraploidy correlating with a poorer outcome

[25, 26]. Several other chromosomal structural abnormalities occur recurrently in NB, such as

deletions at chromosome 1p, 3p and 11q, all thought to harbour yet unidentified tumour

suppressor genes [27-30]. Gain of chromosome 17q, harbouring hypothetical oncogenes which

might play a role in the oncogenesis of NB, represents the most frequent genetic abnormality in

NB, and is thought to be a powerful independent predictor of poor outcome [31-34].

4.7.2 Genetic subtypes in neuroblastoma

Taking into account the different genetic alterations mentioned above, distinct genetic subtypes of

neuroblastoma have been described with specific prognostic characteristics [32, 35-39]. A

genomic profile characterised by whole chromosome gains or losses is observed more frequently

in localised tumours and in children less than 1 year of age, with a good prognosis [32]. Whereas,

unbalanced chromosome translocations leading to segmental chromosome gains or losses,

including in particular chromosome 17q gain, are observed more frequently in advanced stages of

disease or in older children. It is well recognised that tumours presenting with MYCN

amplification in addition to segmental chromosome alterations show highly aggressive clinical

behaviour and are associated with a poor outcome.

Until recently, the various genetic markers were analysed separately by conventional karyotyping,

24-colour karyotyping, FISH and/or LOH studies, making these analyses time consuming and

rendering their interpretation difficult as not all markers could be studied at one time. The

techniques of Comparative Genomic Hybridisation (CGH) and multiplex ligation-dependent

probe amplification (MLPA) enable the analysis of the genetic markers in a single step.

MLPA is a recent PCR based technique that detects a large variety of segmental alterations and

gene amplifications in a robust manner. Simultaneous investigation of a large number of loci,

covering all currently known important aberrant regions found in neuroblastomas, can be

performed in a single assay. The reliable nature of the results and the relatively low cost of the

MLPA kits make this technique attractive for routine NB analysis. This technique is robust and

can be applied in virtually every molecular biology laboratory. After designing a neuroblastoma



specific test kit and internal validation, inter-laboratory testing was performed engaging 7

different European SIOPEN Biology laboratories. This study revealed an excellent concordance

of the results. As a consequence, the SIOPEN Biology Group recommends MLPA and array-CGH

as the technique to identify segmental/structural alterations in neuroblastic tumours.

Several CGH and array-CGH studies have been published recently [4, 5, 40]. Schleiermacher and

Janoueix-Lerosey both report that a genomic profile characterised by the presence of any of the

segmental alterations typically observed in NB is strongly associated with a higher risk of relapse

and metastatic relapse, as well as a poorer outcome in the overall population. Finally, an analysis

of 175 samples of patients included in the previous INES99.1 (infants with localised unresectable

NB) INES99.2 (infants with INSS stage 4s) and INES99.3 (infants with INSS stage 4 disease

without MYCN amplification) have confirmed these results in the infant population

(Schleiermacher, Couturier, oral presentation ANR2008 Congress; Tokyo, Japan, May 2008).

Indeed, this study shows that segmental chromosome alterations are observed in approximately

10%, 20% and > 50% of patients in the 99.1, 99.2 and 99.3 protocol arms respectively. In this

population, segmental changes are associated with poorer progression free survival in all 3

protocol arms. Furthermore, in INSS stage 4s patients, segmental changes are not associated with

a more severe clinical presentation at the time of diagnosis, but with a significantly higher rate of

relapse and a significantly higher treatment burden for salvage therapy. This indicates that in

these patients an upfront treatment more aggressive than suggested by the clinical presentation is

justified.

Preliminary results of array-CGH analysis of 165/300 patients included in INES 99.1, 99.2 and

99.3 protocols

Protocol Arm 4 year PFS Log-rank

INES 99.1 NCA only (n=66) 94.9% ± 2.8

P=0.03 Any SCA (n=8) 75% ± 15

INES 99.2 NCA only (n=57) 87% ± 4.4

P=0.0003 Any SCA (n=13) 36.9% ± 13.8

INES 99.3 NCA only (n=9) 100%

P=0.11 Any SCA (n=12) 75% ± 12

In conclusion, the genomic type is thought to reflect an underlying genomic abnormality. In

tumours with numerical alterations, an abnormality in the mitotic segregation of chromosomes is

thought to exist. On the other hand, the structural chromosome alterations in high and

intermediate risk tumours are most frequently due to unbalanced chromosome translocations

which in turn are thought to arise from DNA double-strand breaks repaired erroneously due to an

impaired DNA maintenance or repair pathway [41, 42].

Thus, in the current LINES protocol, MCYN status, ploidy and the genomic profile detremined by

MLPA or array-CGH will be determined, and the latter will be taken into account for therapeutic

stratification in the Low Risk protocol arm.

Independent of the technique used, a uniform nomenclature, standard operating procedures and

quality assessment studies are indispensable to achieve and maintain the high quality of the

genetic test results needed [43, 44].



5. AIMS

5.1 Low risk study

5.1.1 Primary aim

To demonstrate through a randomisation between observation and chemotherapy that you can

safely reduce treatment in a subgroup of L2 low risk patients (those without life threatening

symptoms (LTS) and without any segmental chromosomal changes (SCA), i.e. study group 1,

page 47) by giving less treatment than has been given historically while maintaining an excellent

OS of 100%.

5.1.2 Secondary aims

To maintain a 2 year EFS of at least 90% and an OS of at least 95% in L2 patients with LTS

without SCA (Study Group 2, page 54).

To maintain the 2 year EFS of 85% and an OS of at least 98% in Ms patients without SCA

(Study Groups 4 and 5, pages 62 and 65).

To improve the 2 year EFS to at least 90% and maintain the OS of close to 100% in L2

patients with SCA (Study Group 3, page 58) and improve the 2 year EFS to over 70% in Ms

patients with SCA (study group 6, page 69).

To evaluate adherence to the protocol recommendations regarding LTS.

To reduce surgical morbidity by promoting strict adherence to IDRFs to determine surgical

resectability.

To define the long term follow-up and natural history of the Stage L2 non-resected masses

that have remained IDRF positive at the end of treatment (study groups 1-3).

5.2 Intermediate risk study

5.2.1 Primary aim

To improve the EFS to 70% with an OS of 90% of INRG stage L2 patients over the age of 18

months, with poorly differentiated or undifferentiated tumour histology (INPC criteria), by the

addition of radiotherapy and 13-cis-RA compared to historical conventional treatment (Kohler J,

personal communication) (Study group 8, page 81).


To confirm in a larger patient cohort the excellent OS of 95% in stage M NB without MYCN

amplification, less than 12 months of age, when treated with moderate therapy (study group

10, page 88) (as achieved in INES 99.3 [3])

To maintain the excellent results of 3 year EFS of 90% and 3 year OS of 100% in stage L2

patients over the age of 18 months, with differentiating neuroblastoma or differentiating

ganglioneuroblastoma nodular, despite a treatment reduction (study group 7, page 78).

To improve 3 year EFS to at least 50% and the 3 year OS to 80% in INSS stage I patients with

MYCN amplified NB by the addition of adjuvant treatment (study group 9, page 85).

To evaluate the impact of the tumour genomic profile on patient outcome, in order to consider

its role in the treatment stratification of these intermediate risk patients (all study groups).



6. TREATMENT STRATIFICATION CRITERIA

This SIOPEN biological and clinical study for low and intermediate risk neuroblastoma patients

uses new criteria to stratify treatment.

6.1 Clinical Staging

The clinical staging definitions used in this protocol are as follows.

6.1.1 Non-metastatic neuroblastoma

For non-metastatic neuroblastoma, the INRG Staging System [1] will be used.

Stage L1: Locoregional tumour not involving vital structures as defined by the list of Image

Defined Risk Factors (IDRF; Appendix 4, page 114).

Stage L2: Locoregional tumour with presence of one or more IDRFs.

Within Stage L1 tumours with MYCN amplification there are very occasionally INSS stage 1 [45]

localised tumours with complete gross excision, with or without microscopic residual disease who

have representative ipsilateral lymph nodes microscopically negative for tumour (nodes attached

to and removed with the primary tumour may be positive) [1]. This group of tumours (in contrast

to any other stage of patient with MYCN amplification who are all treated as a high risk patients)

are included in this study as intermediate risk patients.

6.1.2 Metastatic neuroblastoma

For metastatic neuroblastoma, the INRG Staging system will be used but modified so that it is in

accordance with the staging used in the previous trial for this group of patients INES 99.2-99.3

[3].

Stage M: Distant metastatic disease with positive MIBG (except Stage Ms). In infants <12

months, MIBG or technetium scintigraphy uptake in the skeleton must be confirmed with a bone

abnormality demonstrated on Plain X-ray and/or CT scan.

Stage Ms: Metastatic disease confined to skin and/or liver and/or bone marrow (or even other

sites such as lymph nodes and/or testes), but NOT bone, lung, pleura or CNS, in infants <12

months. MIBG or technetium scintigraphy uptake to the skeleton may occur but there should be

NO X-Ray or CT evidence of bone involvement.

6.2 New concepts for treatment stratification

The new concepts prospectively incorporated into this study for treatment stratification are listed

below.

6.2.1 Genotyping

Genotyping of the tumour will be done by Comparative Genomic Hybridization (array-CGH) or

multiplex ligation-dependent probe amplification (MLPA).

Genotyping is mandatory for all the patients.

In the low risk study, this will enable specific patients to have an opportunity for

treatment reduction in a randomised manner

In the intermediate risk study for the first time in this category of patients, the

determined genotyping profile will be prospectively compared with the INPC



classification. However, in this patient group genotyping will not be used to determine

treatment.

Genotyping is classified as follows:

NCA GENOMIC TYPE

Presence of numerical chromosomal alterations (NCA) only

SCA GENOMIC TYPE

Presence of any segmental chromosomal alteration (SCA) observed recurrently in neuroblastoma

(deletion of chromosome 1p, 3p, 4p, or 11q; gain of 1q, 2p, or 17q) without or with numerical

chromosomal alterations

NO RESULT

No sample or sample containing an insufficient number of tumour cells

Technical failure

Prognostically non informative

presence of segmental alterations but none of those observed recurrently in NB;

no chromosomal abnormality (neither NCA nor SCA) despite sufficient tumour

cell content of the analysed sample

6.2.2 International Neuroblastoma Pathology Classification (INPC)

For patients included in the intermediate risk study, INPC classification [46, 47] of the tumour is

mandatory and will be used for treatment arm allocation.

6.2.3 International Neuroblastoma Risk Group Staging System (INRGSS)

See Appendix 1 (page 110).

6.2.4 Image Defined Risk Factors (IDRF)

These are based on cross sectional imaging prior to surgery. They are intended to guide surgeons

in the preoperative period and identify the patients in whom it would be a resection would be

hazardous and or incomplete (see Appendix 4, page 114).

The same system of risk factors can be applied during the course of treatment and follow-up,

although it is not designed for this purpose. Using IDRFs in tumours exposed to chemotherapy

provides a useful tool for assessing whether a surgical resection should be performed.

6.2.5 Life Threatening Symptoms (LTS)

This list has been revised (see Appendix 3, page 113).



7. ELIGIBILITY

Common Inclusion criteria for all the studies: INFORMED CONSENT AND FOLLOW-UP WARRANTED.

GROUP ASSIGNMENT COMPLETED WITHIN 6 WEEKS FROM DIAGNOSIS

NO PRIOR CHEMOTHERAPY* OR RADIOTHERAPY

Common exclusion criteria for all the studies: DIAGNOSIS OF GANGLIONEUROMA OR GANGLIONEUROBLASTOMA INTERMIXED

* For the purposes of this study, dexamethasone will not be considered chemotherapy and may be

administered prior to initiating trial therapy, at the discretion of the investigator.

LOW RISK STUDY INCLUSION CRITERIA EXCLUSION CRITERIA

L2 Locoregional tumour with presence of one

or more IDRFs


Tumour genomic profile

obtained in a NRL according

to guidelines (pages 122-123)

MYCN non-amplified

≤ 18 months

Any metastatic site

MYCN amplification

>18 months

Ms Metastatic disease confined to skin and/or

liver and/or bone marrow (or even other

sites such as lymph nodes and/or testes),

but NOT bone, lung, pleura or CNS, in

infants ≤12 months. MIBG or technetium

scintigraphy uptake to the skeleton can

occur but without bone lesions

documented by X-ray and/or CT scan





MYCN non-amplified

≤12 months

Bone, Pleura/lung,

and/or CNS metastasis

MYCN amplification

>12 months

NOTE: If the tumour biopsy is performed after the start of treatment the genomic profile result

will be considered non informative and cannot be used for treatment stratification. These patients

will not be eligible for the study. However, these patients should still be registered in the trial and

the required information submitted. See section 9.4.7 (page 72) for treatment recommendations

for this group of patients.



INTERMEDIATE RISK STUDY INCLUSION CRITERIA EXCLUSION CRITERIA

(L1) INSS1 Localised tumours with complete gross

excision, with or without microscopic

residual disease; representative

ipsilateral lymph nodes negative for

tumour microscopically (nodes attached

to and removed with the primary tumour

may be positive)


Tumour available for genomic

profiling according to

guidelines (pages 122-123)

MYCN amplified

MYCN non-amplified


L2 Locoregional tumour with presence of one

or more IDRFs


and confirmed in a NRL:

differentiating, poorly

differentiated, undifferentiated

NB or ganglioneuroblastoma

nodular




MYCN non-amplified

> 18 months

Neuroblastoma NOS.

MYCN amplification.

≤ 18 months

M Distant metastatic disease with positive

MIBG (except Stage Ms).In infants ≤12

months, MIBG or technetium scintigraphy

uptake to the skeleton to be confirmed

with a bone abnormality visible on X-ray

and/or CT scan.





MYCN non-amplified

≤ 12 months

MYCN amplification.

> 12 months

NOTE: L2 patients without a defined histotype will not be eligible for the study. However, these

patients should still be registered in the trial and the required information submitted. See section

10.4.5 (page 91) for treatment recommendations for this group of patients.



8. DIAGNOSTIC EVALUTATION

8.1 Definition of date of diagnosis

The date of diagnosis for the low and intermediate risk study groups is defined as being the date

of diagnostic biopsy/surgery.

8.2 Mandatory pre-treatment steps

There are three mandatory steps to be done before treatment starts:

Diagnosis of neuroblastoma according to INRGSS criteria (patients with ganglioneuroma and

ganglioneuroblastoma intermixed are excluded from this study and should be recruited into

the LNESG 2 trial).

Complete metastatic work-up with bone marrow evaluation (see section 8.3.5) and imaging:

CXR, abdominal US, CT or MRI of the primary tumour, MIBG scan (technetium bone scan if

primary tumour MIBG negative). Bone imaging is necessary to confirm symptomatic or

isotope positive areas of bone and a CT skull is required if bony skull disease is suspected.

Biology work-up completed at a SIOPEN/ENQUA National Reference Laboratory: Tumour

tissue studied for tumour cell content, MYCN status, ploidy status and genomic profile.

In case of emergency treatment, metastatic work up and biology studies should be carried out no

later than 7 days after initiation of emergency treatment.

8.3 Evaluation at diagnosis for low and intermediate risk patients

8.3.1 Full history

With attention to: weight loss, diarrhoea, pain, hypertension, mobility and bladder function. Note

duration of these symptoms.

8.3.2 Full clinical examination (including blood pressure)

Careful attention to the presence of:

any skin lesions.

absence of opsoclonus-myoclonus.

hepatomegaly or “masses” in different regions (cervical, axillae, inguinal).

signs of spinal cord compression.

life threatening symptoms (Appendix 3, page 113).

8.3.3 Haematology/Biochemistry

Full blood count.

Coagulation profile to rule out DIC.

Renal and liver function biochemistry (Na, K, Ca, Mg, PO4, urea, creatinine, glucose,

bilirubin, transaminases).

Serum lactic dehydrogenase (LDH).

Urine catecholamine metabolites (VMA and HVA minimum), measured in mol/mmol of

creatinine. It is recommended that dopamine is also measured.

8.3.4 Imaging

AP chest X-ray.

Abdominal ultrasound (US) for all primary masses. Baseline US to be used as reference for

further follow-up.

CT or MRI of primary tumour, with tumour volume calculated by elliptical approximation

(D1 x D2 x D3 x 0.52). Tumour volume can be calculated also by common software

algorithms which are implemented in most radiological workstations.

123I mIBG Isotope scintigraphy.



Tc MDP (Technetium) scintigraphy if primary tumour MIBG negative.

Bone radiographs of any symptomatic areas and of areas which are positive on isotope scan

to confirm bone disease. If bone disease of the skull is suspected clinically or radiologically

or in case of positive isotope scan of the skull, CT scan of the skull.

NOTE: MDP Tc 99 (Technetium) bone scan is mandatory for assessment of the skeleton if

the MIBG scan is negative at the primary tumour site or cannot be evaluated, and if there is

no clear evidence of enhanced MIBG uptake in the liver. If Tc scan is carried out then areas

appearing positive on the Tc scan should have bone radiographs carried out. In these cases

the definition of bone involvement is the same as for those with MIBG positive scans (i.e.

only with concurrent cortical bone abnormalities on skeletal radiographs (or CT scan)).

8.3.5 Bone marrow

For staging procedures, the bone marrow should be studied according to INSS recommendations:

Bone marrow aspirations (from two separate evaluable sites) for morphology and bone marrow

trephines (from two separate evaluable sites) are recommended where possible. They should be

performed by people with experience, as it is difficult to obtain satisfactory specimens in infants. If

trephines are not done then it is recommended that 4 aspirates are taken. All other minimal residual

disease studies performed by different national groups are NOT MANDATORY and they should

not influence STAGING.

8.3.6 Pathology and biology of the primary tumour (see Appendix 5 and Appendix 6)

It is essential that all children have a biopsy of the primary tumour, even in the presence of

metastatic disease (providing this can be obtained with minimal trauma to the child) so that

pathological and biological information can be obtained. If a biopsy of the primary would be

hazardous, for example in stage 4S with a small primary tumour and massively enlarged liver,

then a Tru-Cut biopsy of the liver would suffice. If skin nodules are present a biopsy of these may

be performed. If there is difficulty in obtaining a biopsy, please discuss with the co-

ordinators. For children with L2 tumours, an open biopsy is preferred, since the quality of

the pathological and biological studies can be higher. For those centres which use multiple

Tru-Cuts instead of open biopsy, sufficient amount of tumour material for pathological and

biological studies must be guaranteed.

Mandatory pathological analysis is histological grading according to INPC [46, 47]. Mandatory

biological analyses are percentage of tumour cell content of the tumour sample, ploidy, MYCN

status and genomic profile obtained in a National Reference Laboratory.

8.3.7 Organ function

Studies of cardiac function: Basal echocardiography prior to any anthracycline therapy is

mandatory.

Audiology is mandatory for patients > 1 year and highly recommended for patients < 1 year

prior to receiving carboplatin.



8.4 Low Risk Study - Pre-treatment investigations

PRE-TREATMENT EVALUATION

Date

FULL HISTORY *

PHYSICAL EXAMINATION *

PHOTOGRAPHS (of any signs) +

BLOOD PRESSURE *

LTS PRESENT? * YES NO

HAEMATOLOGY/BIOCHEMISTRY *

FULL BLOOD COUNT *

COAGULATION PROFILE *

RENAL AND LIVER FUNCTIONS *

LDH *

URINE CATECHOLAMINES *

IMAGING *

CHEST X-RAY *

US *

CT or MRI *

MIBG *

Tc BONE SCAN +

BONE X-RAY +

BONE MARROW

BIOPSIES *

ASPIRATES *

BIOPSY *

PATHOLOGY *

MYCN * MNA No MNA

PLOIDY *

GENOMIC TYPE * NCA SCA

CARDIAC FUNCTION - ECHO * prior to anthracycline therapy

HEARING + prior to carboplatin

* Mandatory evaluation

+ According to indications

For more details see section 8.3, page 39.



8.5 Intermediate Risk Study - Pre-treatment investigations

PRE-TREATMENT EVALUATION

Date

FULL HISTORY *

PHYSICAL EXAMINATION *

PHOTOGRAPHS (of any signs) +

BLOOD PRESSURE *

LTS PRESENT? + YES NO

HAEMATOLOGY/BIOCHEMISTRY *

FULL BLOOD COUNT *

COAGULATION PROFILE *

RENAL AND LIVER FUNCTIONS *

LDH *

URINE CATECHOLAMINES *

IMAGING *

CHEST X-RAY *

US *

CT or MRI *

MIBG *

Tc BONE SCAN +

BONE X-RAY +

BONE MARROW

BIOPSIES *

ASPIRATES *

BIOPSY *

PATHOLOGY *

MYCN * MNA No MNA

PLOIDY *

GENOMIC TYPE * NCA SCA

CARDIAC FUNCTION – ECHO * prior to anthracycline therapy

HEARING + prior to carboplatin

* Mandatory evaluation

+ According to indications

For more details see section 8.3, page 39.



9. LOW RISK STUDY: TREATMENT STRATEGY

9.1 Summary

The low risk groups are MYCN non-amplified neuroblastoma patients with or without life

threatening symptoms in the following clinical situations:

≤ 18 months with localised neuroblastoma associated with image defined risk factors

precluding upfront surgery (stage INRG L2).

≤ 12 months with disseminated neuroblastoma without bone, pleura, lung or CNS disease

(stage INRG Ms)

All the patients need to have a tumour genomic profile result. Treatment is stratified by stage,

presence or not of life threatening symptoms and the tumour genomic profile.

The purpose of this study is to:

1. Decrease the amount of treatment (chemotherapy and surgery) for appropriate low risk

unresectable patients, who in previous studies have been shown to have an excellent long-

term outcome (as in the SIOPEN 99.1-2 infant neuroblastoma studies where the overall

survival was greater than 97%, personal communication H. Rubie, manuscript in

preparation). There is a treatment reduction randomisation for L2 patients with no

symptoms and no segmental chromosomal change between initial observation and delayed

chemotherapy if there is local progression, and initial chemotherapy

2. Improve the EFS (and maintain the OS) in asymptomatic L2 and Ms patients who have

segmental chromosomal changes in their tumour genomic profile by electively treating

these patients with chemotherapy despite the absence of symptoms.

3. Reduce surgical morbidity by promoting strict adherence to Image Defined Risk Factors

(IDRFs) to determine surgical resectability. IDRFs have been scientifically validated in

tumours never exposed to chemotherapy. In tumours exposed to chemotherapy, IDRFs

(although not scientifically validated in this situation) are the only objective method of

assessing whether a surgical resection should be performed (Appendix 4, page 114 and

section 10.4, page 139).

The patients are treated within 6 separate study groups.

Patients eligible for this study will include those with dumbbell tumours with intraspinal

neuroblastoma. These patients may be either asymptomatic or symptomatic with symptoms

of spinal cord compression. In this protocol it is recommended that patients who have a

spinal cord component greater than 50% of the diameter of the spinal canal are treated with

chemotherapy even in the absence of signs or symptoms of spinal cord compression. These

patients should be treated as per the detailed recommendations in appendix 13 (page 157).

9.1.1 Study groups 1-3, Stage L2, MYCN non-amplified and < 18 months

9.1.1.1 Study group 1

L2 patients with a NCA genomic profile and with no life threatening symptoms. These patients

will be randomised between initial observation (chemotherapy is only given if there is subsequent

progression) or initial chemotherapy treatment.


L2 patients with a NCA genomic profile, with life threatening symptoms. Treatment:

Chemotherapy




L2 patients with a SCA genomic profile, with or without life threatening symptoms. Treatment:

Chemotherapy

9.1.2 Study groups 4-6, Stage Ms, MYCN non-amplified and < 12 months


Ms patients with a NCA genomic profile, without life threatening symptoms Treatment:

Observation


Ms patients with a NCA genomic profile, with life threatening symptoms Treatment:

Chemotherapy


Ms patients with a SCA genomic profile, with or without life threatening symptoms Treatment:

Chemotherapy

NOTE: Chemotherapy should never be delayed in a patient with life threatening symptoms in

order to obtain a pre-treatment biopsy. A biopsy should be undertaken only once the patient is

stable. For the purposes of this study a genomic profile result from a tumour biopsy taken after

chemotherapy has been given will be considered non informative. See section 9.4.7 (page 72) for

treatment recommendations.

9.2 Eligibility

Common Inclusion criteria for all the studies: INFORMED CONSENT AND FOLLOW-UP WARRANTED




* For the purposes of this study, dexamethasone will not be considered chemotherapy and may be administered prior

to initiating trial therapy, at the discretion of the investigator.

LOW RISK STUDY INCLUSION CRITERIA EXCLUSION CRITERIA

L2 Locoregional tumour with presence of

one or more IDRFs





MYCN non-amplified

≤ 18 months

Any metastatic site

MYCN amplification

>18 months

Ms Metastatic disease confined to skin and/or

liver and/or bone marrow (or even other

sites such as lymph nodes and/or testes),

but NOT bone, lung, pleura or CNS, in

infants ≤12 months. MIBG or technetium

scintigraphy uptake to the skeleton can

occur but without bone lesions

documented by X-ray and/or CT scan





MYCN non-amplified

≤12 months

Bone, Pleura/lung,

and/or CNS metastasis

MYCN amplification

>12 months

NOTE: If the tumour biopsy is performed after the start of treatment the genomic profile result

will be considered non informative and cannot be used for treatment stratification. These patients

will not be eligible for the study. However, these patients should still be registered in the trial and

the required information submitted. Treatment recommendations for this group of patients are in

section 9.4.7 (page 72).



9.3 Study Design Overview

For more details see the study group specific treatment flow charts.

9.3.1 Low Risk Study: L2 Patients (≤ 18 months)

NOTE: In tumours exposed to chemotherapy IDRFs (although not scientifically validated in this situation)

are the only objective method of assessing whether a surgical resection should be performed (Appendix 4,

page 114 and section 10.4, page 139).

GENOMIC PROFILES obtained in a National Reference Laboratory:

NCA genomic profile - presence of numerical chromosomal alterations (NCA)

only SCA genomic profile - presence of any segmental chromosomal alteration

(SCA) observed recurrently in neuroblastoma (deletion of chromosome 1p, 3p,

4p, or 11q; gain of 1q, 2p, or 17q) without or with numerical chromosomal alterations

No result - see recommendations section 9.4.7 (pg. 72)

L2 ≤ 18 months

No life threatening symptoms

Life threatening symptoms

NCA genomic profile RANDOMIZE Study Gp 1

SCA genomic profile NO

RANDOMIZATION Study Gp 3

Observation

Chemotherapy:

2-4 CO

Aim for surgical resection as soon as

IDRF negative

No LTS: 4 VP/Carbo LTS: 4 VP/Carbo; if LTS persist after 2 VP/Carbo then 2 CADO

Surgery if IDRF

negative. If IDRF positive

discuss with Surgical Coordinators

NCA genomic profile Study Gp 2

2VP/Carbo to resolve LTS. If LTS persist after 2 VP/Carbo then 2 CADO

Surgical resection only if IDRF negative

2 VP/Carbo as indicated for further

response

Surgical resection 1 year from diagnosis if

IDRF negative

In case of progression with

LTS 2 VP/Carbo ± CADO (see pg 50)



9.3.2 Low Risk Study: Ms Patients (≤ 12 months)

NOTE: In tumours exposed to chemotherapy, IDRFs (although not scientifically validated in this situation)



GENOMIC PROFILES obtained in a National Reference Laboratory:

NCA genomic profile - presence of numerical chromosomal alterations (NCA)

only SCA genomic profile - presence of any segmental chromosomal alteration

(SCA) observed recurrently in neuroblastoma (deletion of chromosome 1p, 3p,

4p, or 11q; gain of 1q, 2p, or 17q) without or with numerical chromosomal alterations

No result - see recommendations section 9.4.7 (pg. 72)

Ms ≤ 12 months

No life threatening symptoms

Life threatening symptoms


SCA genomic profile Study Gp 6

Observation

Surgical resection of primary not indicated

No LTS: 4 x VP/Carbo LTS: 4 x VP/Carbo but if LTS persist after 2 x VP/Carbo then 2 x CADO


2 VP/Carbo to resolve LTS. If LTS persist after 2 VP/Carbo then2 CADO

Surgical resection of primary not indicated

Surgery if IDRF negative.

If IDRF positive discuss with Surgical Coordinators



9.4 Treatment strategy for low risk patients

9.4.1 Study group 1: randomised study

9.4.1.1 Study group 1: Treatment Flow Chart – Randomised to Observation


are the only objective method of assessing whether a surgical resection should be preformed (Appendix 4,


Stage L2, MYCN non-amplified

≤ 18 months

NCA genomic profile

without life threatening symptoms (LTS)

Reassessment every 8 weeks

until there is evidence of

regression and then every 12

weeks for 1 year.

RANDOMISED

Observation

In case of progression with

LTS (see page 50 for

definition and treatment

details)

Surgical resection indicated

only if IDRF are negative 1

year from diagnosis.



9.4.1.2 Study group 1: Treatment Flow Chart – Randomised to Chemotherapy




RANDOMISED

Chemotherapy

Reassessment

2 x CO

IDRF -ve Surgical

resection

IDRF +ve

No response to

treatment

At least an

objective response

Reassessment

2 x CO

IDRF +ve

2 x VP/Carbo

Reassessment

IDRF +ve

IDRF -ve

Observation

No indication for

further chemotherapy

Surgical

resection


≤ 18 months

NCA genomic profile




9.4.1.3 Study group 1: Road Map

L2, ≤18 months, NCA genomic profile, Without LTS

Refer to the protocol text for more information and treatment details

WEEK Dx 1 3 5 7 9 12

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * * *

BIOPSY *

TOXICITY MONITORING

*

*

*

*

*

*

*

GENOMIC TYPE ASSIGNED *

RANDOMISED TREATMENT ARM: CHEMOTHERAPY

Cycle 1 2 3 4 5 6

CO • • • •

VP/Carbo • • • •

SURGERY if IDRF negative * * *

RANDOMISED TREATMENT ARM: OBSERVATION

SURGERY at 1yr, if IDRF negative *

* indicates that the test/monitoring/surgery should be prior to the start of the related course

RANDOMISE TO:

Chemotherapy Arm. 2-4 CO, 2 VP/Carbo (max).

Aim for surgical resection when IDRF negative (evaluation q2 cycles)

Observational Arm. Imaging q8 weeks

Surgical resection is indicated 1 year from diagnosis if IDRF negative




OR OR



9.4.1.4 Study group 1: Treatment Strategy – Randomised to Observation

Eligible patients

Stage L2, ≤ 18 months, MYCN non-amplified with a NCA genomic profile, with no life

threatening symptoms (see Appendix 3, page 113): These patients will be randomised between

initial observation and treatment with chemotherapy. Randomisation will be stratified depending

on the country and will be performed online through the RDE system, using the portal

https://www.siopen-r-net.org/. Randomisation shall be done within six weeks from the date of

diagnosis (see section 8.1, for the definition of the date of diagnosis and appendix 18, point

A18.2.3 for randomisation details).

NOTE: Patients not consenting to randomisation should be treated according to the chemotherapy

arm.

Treatment: Randomised to observation

These patients should be observed to allow spontaneous regression of their tumour. Regular

reassessment will be undertaken.

Surgical management: randomised to observation

These patients should only have a surgical resection of the primary mass if at one year from

diagnosis the primary mass is assessed as IDRF negative.

For further details of surgical management see surgical section: Appendix 9, page 138.

Investigations and assessment during treatment: randomised to observation

Regular reassessments will be undertaken with imaging of the tumour every 8 weeks until there is

evidence of regression and then every 12 weeks for one year. Imaging can be by USS or MRI

depending on the local unit. CT scanning should be avoided or kept to a minimum to avoid

unnecessary radiation. When assessing tumour response, tumour volume should be calculated as

outlined in Appendix 5 (page 115).

Disease progression and management: randomised to observation

In case of progression a full reassessment should be undertaken.

o Definition of progression requiring a change in clinical management (i.e. to be

considered an event)

1. Development of life threatening symptoms (Appendix 3, page 113)

NOTE: Local increase of tumour volume without the appearance of LTS is not an

indication for starting chemotherapy.

2. Development of metastatic disease (bone, lung and pleura and CNS). NOTE:

Liver, bone marrow and skin metastases in a patient ≤ 12 months implies Ms

disease and would only be treated if there were symptoms present.

o Reassessment (needed to re-establish stage)

1. Tumour biology – consider the need to reassess

2. 3D imaging of local tumour, MIBG scan and further imaging if indicated as per

pre-diagnosis investigations (section 8.3)

3. Bilateral bone marrow aspirates and trephines

o Management of progression

If MYCN amplification present treat on high risk protocol.

https://www.siopen-r-net.org/



Localised disease with LTS: If no MYCN amplification is present and disease

is localised treatment is with chemotherapy:

Chemotherapy should start with 2 courses of VP/Carbo. If the LTS

resolve after 2 courses of VP/Carbo stop treatment. However, if the

LTS persist after 2 courses of VP/Carbo continue treatment with 2

courses of CADO.

Surgical management: If the IDRF become negative after four courses

of chemotherapy then a surgical resection of the primary tumour should

be undertaken. If IDRF are present discuss with the Study Coordinator.

If a tumour remains IDRF positive after chemotherapy this is not an

absolute contraindication to surgery. Resection may still be

recommended if evaluation of the primary tumour suggests that the risk

to life, or of major functional loss, is less than the risk from leaving

residual disease.

Metastatic Disease: If no MYCN amplification is present and there is

metastatic disease in the bone, lung and/or CNS they need to be treated as stage

M disease. The treatment they should then receive depends on their age when

these metastases develop.

Patients ≤ 12 months go on to the intermediate risk protocol.

Patients > 12 months go on to the high risk protocol.

9.4.1.5 Study group 1: Treatment Strategy – Randomised to Chemotherapy

Eligible patients

Stage L2, ≤ 18 months, MYCN non-amplified with a NCA genomic profile, with no life

threatening symptoms (see Appendix 3, page 113): These patients will be randomised between

initial observation and treatment with chemotherapy.

Treatment: Randomised to chemotherapy

These patients will receive chemotherapy (as in a historical control arm) which is identical to

the treatment given to this group of patients in the First European Infant Neuroblastoma Study

(INES 99.1). The aim of this arm of treatment is to enable the tumour to become IDRF

negative with the minimum number of courses of chemotherapy so that a surgical resection

can proceed safely.

Two courses of Vincristine and Cyclophosphamide (CO x 2) followed by tumour

reassessment with USS or MRI. If no IDRF are then present, chemotherapy is discontinued

and a surgical resection performed.

If IDRF persist BUT there has been some objective response to treatment a further two

courses of Vincristine and Cyclophosphamide (CO x 2) should be given followed by

reassessment. If no IDRF are then present, stop chemotherapy and proceed to surgical

resection.

If IDRF still persist after 4 courses of CO then continue treatment with two courses of

Etoposide and Carboplatin (VP/Carbo x 2) followed by reassessment. If no IDRF present stop

chemotherapy and proceed to surgical resection.

If there is stable disease or an increase in tumour volume after the first two courses of CO,

treatment should switch to Etoposide and Carboplatin. Two courses should be given followed

by reassessment.

If no IDRF present after two course of VP/Carbo, proceed to surgical resection.

If IDRF still present after a total of four (CO x 2 and VP/Carbo x 2) or six (CO x 4 and

VP/Carbo x 2) courses of chemotherapy, the chemotherapy should be stopped and observation

continued.

There is NO recommendation for these patients to receive CADO chemotherapy.



Refer to Appendix 11 (page 143) for detailed information regarding chemotherapy and dose

modification.

Surgical management: randomised to chemotherapy

A surgical resection of the primary mass in low risk infants (L2) should not be undertaken if

IDRF are present during first-line treatment.

The patients randomised to the chemotherapy arm should have a surgical resection as soon as the

tumour becomes IDRF negative following the minimum number of chemotherapy courses.


Investigations and assessment during treatment: randomised to chemotherapy

Regular reassessments will be undertaken with imaging of the tumour after every 2 courses of

chemotherapy as above. When chemotherapy has been discontinued patients who still have an

unresectable IDRF positive tumour should have 12 weekly tumour reassessments and then if

stable these should be annual whereas patients who have had a surgical resection there should be

an annual assessment of disease status. Imaging can be by USS or MRI depending on the local

unit. CT scanning should be avoided or kept to a minimum to avoid unnecessary radiation. When

assessing tumour response, tumour volume should be calculated by elliptical approximation (D1 x

D2 x D3 x 0.52) or by common software algorithms which are implemented in most radiological

workstations.

Disease progression and management: randomised to chemotherapy



1. Development of life threatening symptoms (Appendix 3, page 113)

2. Development of metastatic disease (bone, lung and pleura and CNS). NOTE:

Liver, bone marrow and skin metastases in a patient ≤ 12 months implies Ms

disease.

NOTE: Local increase of tumour volume without the appearance of LTS should be

treated as indicated above and will not be considered as disease progression (page 51).


1. Tumour biology -consider the need to reassess


pre-diagnosis investigations (section 8.3).



Treatment of progression in these patients depends on re-establishing the

biological profile and on the previous treatment that they have received.

If MYCN amplification present follow high risk protocol treatment

recommendations.

Localised Disease with LTS: If no MYCN amplification then treatment

should be the next chemotherapy in the escalation of treatment:

if CO already given then give VP/Carbo x 2

if VP/Carbo given treat with CADO x 2

if no response to CADO further treatment is at the discretion of the trial

coordinator, consider TVD or Cyclophosphamide/Topotecan.

Surgical management: If the IDRF become negative then a surgical

resection of the primary tumour should be undertaken. If IDRF are

present discuss with the Study Coordinator. If a tumour remains IDRF

positive after chemotherapy this is not an absolute contraindication to

surgery. Resection may still be recommended if evaluation of the

primary tumour suggests that the risk to life, or of major functional

loss, is less than the risk from leaving residual disease.



Metastatic Disease: If no MYCN amplification is present and there is

metastatic disease in the bone, lung and/or CNS they need to be treated as stage

M disease. The treatment they should then receive depends on their age when

these metastases develop.

Patients ≤ 12 months should follow intermediate risk protocol treatment

recommendations.

Patients > 12 months should follow high risk protocol treatment

recommendations.



9.4.2 Study group 2

9.4.2.1 Study group 2: Treatment Flow Chart





≤ 18 months

NCA genomic profile

with life threatening symptoms (LTS)

Reassessment

2 x VP/Carbo

LTS -ve Surgical

resection

LTS +ve

Reassessment

2 x CADO

IDRF -ve

IDRF +ve

Observation

IDRF +ve

Observation

No indication for


Surgical

resection IDRF -ve




L2, ≤18 months, NCA genomic profile, With LTS

L2, ≤18 months, no genomic profile result, With LTS


WEEK Dx 1 4 7 10 13

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

GENOMIC TYPE

ASSIGNED

*

CHEMOTHERAPY

Cycle 1 2 3 4

VP/Carbo • •

CADO • •

SURGERY if IDRF

negative

* *


LTS: 2 VP/Carbo; If LTS persist after 2 VP/Carbo then 2 CADO

Surgical resection is indicated only if IDRF negative






9.4.2.3 Study group 2: Treatment Strategy

Eligible patients

Stage L2, ≤ 18 months, MYCN non-amplified, NCA genomic profile, with life threatening

symptoms (see Appendix 3, page 113)

Treatment

These patients will all receive chemotherapy with a minimum of 2 courses of Etoposide and

Carboplatin (VP/Carbo).

If the LTS persist after 2 courses of VP/Carbo they should receive CADO x 2 .The aim is to

resolve the life threatening symptoms with the minimum number of courses of

chemotherapy.

A surgical resection of the tumour will be performed only if the IDRF become negative.


modification.

Surgical management

A surgical resection of the primary mass in these low risk infants (L2) should not be undertaken if

IDRF are present during first-line treatment. The symptomatic patients who receive

chemotherapy should have a surgical resection only when the tumour becomes IDRF negative. If

the child remains symptomatic after chemotherapy discuss with the National or International

Surgical coordinators.

Dumbbell tumours: If the symptoms resolve and the extraspinal component becomes resectable

i.e. IDRF negative then a surgical resection of the extraspinal component should take place. There

is no indication to surgically resect the residual spinal canal component of a dumbbell tumour.


Investigations and assessment during treatment

The table on page 55 indicates when investigations should be carried out. Investigations are to be

carried out prior to the course referred to.

The courses are numbered to include the maximum number of courses of chemotherapy a patient

might receive (e.g. 2 VP/Carbo and 2 CADO). Assessments for tumour volume are repeated

after every two courses whilst on chemotherapy. When chemotherapy is completed assessments

for tumour volume should be 12 weekly for a year then annual for 5 years.

Disease progression and management

Definition of progression:

1. Development of new or different life threatening symptoms (Appendix 3, page 113).

2. Progression of localised disease (Appendix 5, page 115).

3. Development of metastatic disease (bone, lung and pleura and CNS). NOTE: Liver, bone

marrow and skin metastases in a patient ≤ 12 months implies Ms disease.

If any of the above occurs, this is an indication to undertake a full reassessment including:

1. Tumour biology – consider the need to reassess.

2. 3D imaging of local tumour, MIBG scan and further imaging if indicated as per pre-

diagnosis investigations (section 8.3)




Management of progression depends on which treatment was previously given.

If MYCN amplification present follow high risk protocol treatment recommendations.

Localised progression: If no MYCN amplification and no evidence of metastatic disease

then treatment should be next chemotherapy in escalation of treatment:

if only VP/Carbo has been given treat with CADO x 2.

if CADO x 2 given, treatment is at the discretion of the trial coordinator, consider

TVD or Cyclophosphamide/Topotecan.

Surgical management: If the IDRF become negative then a surgical resection of the

primary tumour should be undertaken. If IDRF are present discuss with the Study

Coordinator. If a tumour remains IDRF positive after chemotherapy this is not an

absolute contraindication to surgery. Resection may still be recommended if

evaluation of the primary tumour suggests that the risk to life, or of major

functional loss, is less than the risk from leaving residual disease.

Metastatic Disease: If no MYCN amplification is present and there is metastatic disease

in the bone, lung and/or CNS they need to be treated as stage M disease. The treatment

they should then receive depends on their age when these metastases develop.

Patients ≤ 12 months should follow the intermediate risk protocol treatment

recommendations.

Patients > 12 months follow the high risk protocol treatment recommendations.

If no MYCN amplification is present and there is metastatic disease in the liver, bone

marrow and/or skin this is Ms disease. The treatment they should then receive depends on

their age when these metastases develop.

Patients ≤ 12 months should be discussed with the National Coordinator.




9.4.3 Study group 3






≤ 18 months

SCA genomic profile

with or without life threatening symptoms

(LTS)

2 x VP/Carbo

Reassessment LTS -ve

LTS +ve

2 x CADO

LTS +ve

2 x VP/Carbo

LTS -ve

Reassessment

2 x VP/Carbo

2 x VP/Carbo

Reassessment

Reassessment

Reassessment

LTS -ve

SD

Surgery if IDRF –ve.

If IDRF +ve discuss with

Surgical Coordinators.


If IDRF +ve discuss with

Surgical Coordinators.




L2, ≤18 months, SCA genomic profile, With OR Without LTS


WEEK Dx 1 4 7 10 13

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

GENOMIC TYPE

ASSIGNED

*

CHEMOTHERAPY

Cycle 1 2 3 4


CADO • •

SURGERY if IDRF

negative†

*


No LTS: 4 VP/Carbo

LTS: 4 VP/Carbo; If LTS persist after 2 VP/Carbo then give 2 CADO (total 4 cycles) †Surgical resection is indicated if IDRF negative. If IDRF positive discuss the possibility of surgery with

the Surgical Coordinators.




OR OR




Eligible patients

Stage L2, ≤ 18 months, MYCN non-amplified, SCA genomic profile, without or with life

threatening symptoms (see Appendix 3, page 113).

Treatment

These patients will all receive chemotherapy.

Patients without life threatening symptoms will have four courses of VP/Carbo. The aim is

for the IDRF to become negative so that a surgical resection can be performed. If the IDRF are

still positive after 4 courses of VP/Carbo NO further courses of chemotherapy should be given

and the patient should be observed.

In patients with life threatening symptoms the aim is to resolve the life threatening

symptoms with four courses of chemotherapy, and for IDRF to become negative and to

proceed to surgical resection.

If the LTS persist and or are not resolving after 2 courses of VP/Carbo, 2 courses of CADO

should be given.


modification.

Surgical management

A surgical resection of the primary mass in low risk infants (L2) should not normally be

undertaken if IDRF are present during first-line treatment. However the balance of risk versus

benefit for these infants who have SCA genomic abnormalities may be influenced by the specific

details of the risk factors, and so tumours that remain IDRF positive after chemotherapy should be

discussed with the National or International Surgical coordinators. If a tumour remains IDRF

positive after chemotherapy this is not an absolute contraindication to surgery. Resection may still

be recommended if evaluation of the primary tumour suggests that the risk to life, or of major







might receive (e.g. 2 -4 VP/Carbo and 2 CADO). Assessments for tumour volume are repeated

after every two courses whilst on chemotherapy. When chemotherapy is completed assessments

for tumour volume should be 12 weekly for a year then annual for 5 years.





3. Development of metastatic disease (bone, lung and pleura and CNS). NOTE: Liver, bone

marrow and skin metastases in a patient ≤ 12 months implies Ms disease.



2. 3D imaging of local tumour, MIBG scan and further imaging if indicated as per pre-diagnosis

investigations (section 8.3)





If MYCN amplification present follow high risk protocol treatment recommendations

Localised progression: If no MYCN amplification and no evidence of metastatic disease

then treatment should be next chemotherapy in escalation of treatment:


if CADO x 2 given, treatment is at the discretion of the trial coordinator, consider

TVD or Cyclophosphamide/Topotecan.







Metastatic Disease: If no MYCN amplification is present and there is metastatic disease

in the bone, lung and/or CNS they need to be treated as stage M disease. The treatment

they should then receive depends on their age when these metastases develop.


recommendations

Patients > 12 months follow the high risk protocol treatment recommendations

If no MYCN amplification is present and there is metastatic disease in the liver, bone

marrow and/or skin this is Ms disease. The treatment they should then receive depends on

their age when these metastases develop.

Patients ≤ 12 months should be discussed with the National Coordinator.




9.4.4 Study group 4


Stage Ms, MYCN non-amplified

≤ 12 months

NCA genomic profile


Reassessment every 8 weeks

until there is evidence of

regression and then every 12

weeks for 1 year.

Observation

In case of progression (see

page 64 for definition) a full

reassessment must be

undertaken.

If progression is confirmed

and LTS develop treat

according to study group 5




Ms, ≤12 months, NCA genomic profile, Without LTS

Ms, ≤12 months, no genomic profile result, Without LTS


WEEK Dx 1 5 9 13 17

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

GENOMIC TYPE

ASSIGNED

*

OBSERVATION Surgical resection is not indicated

* indicates that the test/monitoring/surgery should be done during this week

Surgical resection of the primary is not indicated.




Eligible patients

Stage Ms, ≤ 12 months, MYCN non-amplified, NCA genomic profile, without life threatening

symptoms

Treatment

These patients should just be observed to allow spontaneous regression of their tumour. Regular

assessments will be undertaken.

Surgical management

Resection of the primary tumour in stage Ms is not indicated unless this is carried out as part of

the initial diagnostic workup. It should never be carried out if there are image defined risk factors

present. In patients with regressing tumours there is no evidence that a delayed resection is

necessary.


Imaging of the tumour (in most cases this will be of the involved liver) will be undertaken every 8

weeks until there is evidence of regression and then every 12 weeks for one year, followed by

annual assessments for 5 years.


In case of progression a full reassessment should be undertaken.



1. Development of life threatening symptoms (Appendix 3, page 113).

NOTE: Local increase of tumour volume without the appearance of LTS is not an

indication for starting chemotherapy.

2. Development of metastatic disease (bone, lung and pleura and CNS).


4. Tumour biology – consider the need to reassess


pre-diagnosis investigations (section 8.3).



If MYCN amplification present treat on the high risk protocol.

Progressive Ms disease: If no MYCN amplification is present, and LTS have

appeared, treatment is with chemotherapy:

Start with 2 courses of VP/Carbo, aiming to resolve the LTS with the

minimum number of courses of chemotherapy.

If the LTS persist after 2 courses of VP/Carbo give 2 courses of CADO.

A surgical resection of the primary tumour is not indicated.

Patients without LTS should continue to be observed.

Metastatic disease (Stage M): If no MYCN amplification is present and there

is metastatic disease in the bone, lung and/or CNS they need to be treated as

stage M disease. The treatment they should then receive depends on their age

when these metastases develop.

Patients ≤ 12 months should go onto the intermediate risk protocol.

Patients > 12 months go on to the high risk protocol.



9.4.5 Study group 5



≤ 12 months

NCA genomic profile

with life threatening symptoms (LTS)

Reassessment

2 x VP/Carbo

LTS -ve

LTS +ve

2 x CADO

Observation

No indication for


Observation

No indication for


Reassessment




Ms, ≤12 months, NCA genomic profile, With LTS

Ms, ≤12 months, no genomic profile result, With LTS


WEEK Dx 1 4 7 10 13

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

GENOMIC TYPE

ASSIGNED

*

CHEMOTHERAPY

Cycle 1 2 3 4

VP/Carbo • •

CADO • •

SURGERY Not indicated in this group


LTS: 2 VP/Carbo; If LTS persist after 2 VP/Carbo then 2 CADO

Surgical resection is not indicated in this group




Eligible patients

Stage Ms, ≤ 12 months, MYCN non-amplified, NCA genomic profile, with life threatening

symptoms (see Appendix 3, page 113).

Treatment

These patients will all receive chemotherapy with a minimum of 2 courses of Etoposide and

Carboplatin. The aim being to resolve the LTS with the minimum number of courses of

chemotherapy.

If the LTS persist after 2 courses of VP/Carbo, patients should receive 2 courses of CADO

chemotherapy.


modification.

Surgical management

Resection of the primary tumour in stage Ms is not indicated unless this is carried out as part of

the initial diagnostic workup. It should never be carried out if there are image defined risk factors

present. In patients with regressing tumours there is no evidence that a delayed resection is

necessary.





might receive (e.g. 2 VP/Carbo and 2 CADO). Assessments for tumour volume are repeated after

every two courses whilst chemotherapy is being administered.

When chemotherapy is completed assessments for tumour volume should be 12 weekly for a year

then annually for 5 years.




2. Progression of localised disease or previously known metastatic site (e.g. liver).

3. Development of new metastatic disease (bone, lung and pleura and CNS).



2. 3D imaging of local tumour, MIBG scan and further imaging if indicated as per pre-

diagnosis investigations (section 8.3)



If MYCN amplification is now present follow the high risk protocol treatment

recommendations

Progressive Ms disease: If there is no MYCN amplification and no evidence of metastatic

stage M disease then treatment should be the next chemotherapy in the escalation of

treatment:




if CADO x 2 has been given, treatment is at the discretion of the trial coordinator.

Consider TVD or Cyclophosphamide/Topotecan.



Coordinator. If the tumour remains IDRF positive after chemotherapy this is not an




Metastatic Disease (Stage M): If no MYCN amplification is present and there is

metastatic disease in the bone, lung and/or CNS the patient needs to be treated as stage M

disease. The treatment they should then receive depends on their age when these

metastases develop.


recommendations




9.4.6 Study group 6






≤ 12 months

SCA genomic profile

with or without life threatening symptoms

(LTS)

2 x VP/Carbo

Reassessment LTS -ve

LTS +ve

2 x CADO

LTS +ve

2 x VP/Carbo

LTS -ve

Reassessment

2 x VP/Carbo

2 x VP/Carbo

Reassessment


If IDRF +ve discuss

with Surgical

Coordinators.

Reassessment

Reassessment

LTS -ve


If IDRF +ve discuss

with Surgical

Coordinators.




Ms, ≤12 months, SCA genomic profile, With OR Without LTS


WEEK Dx 1 4 7 10 13

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

GENOMIC TYPE

ASSIGNED

*

CHEMOTHERAPY

Cycle 1 2 3 4


CADO • •

SURGERY if IDRF

negative†

*


No LTS: 4 VP/Carbo

LTS: 4 VP/Carbo; If LTS persist after 2 VP/Carbo then give 2 CADO (total 4 cycles) †Surgical resection is indicated if IDRF negative. If IDRF positive discuss the possibility of surgery with

the Surgical Coordinators.




OR OR




Eligible patients

Stage Ms, ≤ 12 months, MYCN non-amplified SCA genomic profile, without or with life

threatening symptoms (see Appendix 3, page 113)

Treatment

All of these patients will be treated with a total of four courses of chemotherapy.

When no LTS are present give VP/Carbo x 4

When LTS present give VP/Carbo x 2 followed by either 2 further courses of VP/Carbo if the

symptoms have responded or are resolving or 2 courses of CADO if the LTS symptoms are

not resolving with the initial 2 courses of VP/Carbo.


modification.

Surgical management

In this patient group, a resection of the primary tumour in stage Ms disease can be indicated.

Surgical resection should not normally be undertaken if IDRF are present. However the balance of

risk versus benefit for these infants who have SCA genomic abnormalities may be influenced by

the specific details of the risk factors, and so tumours that remain IDRF positive after

chemotherapy should be discussed with the National or International Surgical coordinators.

If a tumour remains IDRF positive after chemotherapy this is not an absolute contraindication to

surgery. Resection may still be recommended if evaluation of the primary tumour suggests

that the risk to life, or of major functional loss, is less than the risk from leaving residual disease.






might receive (e.g. 4 x VP/Carbo, or 4 x VP/Carbo x 2 and CADO x 2). Assessments for tumour

volume are repeated after every two courses whilst chemotherapy is being administered.

When chemotherapy is completed assessments for tumour volume should be 12 weekly for a year

then annually for 5 years.




2. Progression of localised disease or previously known metastatic site (e.g. liver).

3. Development of new metastatic disease (bone, lung and pleura and CNS).



2. 3D imaging of the local tumour, MIBG scan and further imaging if indicated as per pre-

diagnosis investigations (section 8.3).




Management of progression depends on what chemotherapy has been given previously.

If MYCN amplification is now present follow the high risk protocol treatment

recommendations

Progressive Ms disease: If there is no MYCN amplification and no evidence of metastatic

stage M disease then treatment should be the next chemotherapy in the escalation of

treatment:

If only VP/Carbo has been given treat with CADO x 2.

If CADO x 2 has been given treatment is at the discretion of the trial coordinator.

Consider TVD or Cyclophosphamide/Topotecan.







Metastatic Disease (Stage M): If no MYCN amplification is present and there is

metastatic disease in the bone, lung and/or CNS the patient needs to be treated as stage M

disease. The treatment they should then receive depends on their age when these

metastases develop.


recommendations


9.4.7 Treatment recommendations for patients without genomic profile result

In certain situations there will not be a genomic profile result (e.g. technical failure, missing

sample or the genomic profile result available will be considered non informative when it has

been obtained from a biopsy taken after beginning initial chemotherapy). In those situations the

patient cannot receive treatment stratified by genomic profile and will receive standard “historic”

treatment.

These patients should still be registered and will be study patients (rather than trial

patients), the full data set will be collected on them but they will be analysed separately.

These patients must be consented using the “without genomic profile” consent form (page

198).

Patients with Stage L2, ≤ 18 months, MYCN non-amplified without life threatening symptoms

with no genomic profile result treat according to the chemotherapy treatment arm in study group

1.

Stage L2, ≤ 18 months, MYCN non-amplified, no genomic profile result, with life threatening

symptoms treat as study group 2.

Stage Ms, ≤ 12 months, MYCN non-amplified, no genomic profile result, without life threatening


Stage Ms, ≤ 12 months, MYCN non-amplified, no genomic profile result, with life threatening


9.4.8 Chemotherapy and/or surgical procedures outside the recommendations of this protocol

In order for a complete treatment picture to be drawn for these patients, it is important that all

chemotherapy and/or surgical procedures are entered into the LINES database using the

appropriate chemotherapy and/or surgery CRFs via the SIOPEN-R-NET. This should be done

even if the chemotherapy and/or surgical procedure is not within the recommendations of this

protocol.



10. INTERMEDIATE RISK STUDY: TREATMENT STRATEGY

10.1 Summary

In the past these patients have been recruited into three different SIOPEN studies. This

Intermediate Risk Study includes patients in three separate risk groups.

10.1.1 Study groups 7-8

Children aged > 18 months, with INRG stage L2 neuroblastoma, MYCN non-amplified. In the

past these patients were treated according to the European Unresectable Neuroblastoma Study.

Preliminary results show that histology is a significant prognostic factor in this subpopulation,

and, consequently, patients will be stratified according to histology:

o study group 7:

- differentiating neuroblastoma

- differentiating ganglioneuroblastoma nodular

o study group 8:

- undifferentiated or poorly differentiated neuroblastoma

- undifferentiated or poorly differentiated ganglioneuroblastoma nodular

10.1.2 Study group 9

Children with INSS stage 1 neuroblastoma, MYCN amplified. Previously these patients when

recruited into the LNESG 1-2 protocols were only observed. The new recommendation is that

they will receive adjuvant treatment.


Infants (<12 months) with INRG stage M neuroblastoma, MYCN non-amplified. These patients

were treated according to the Infant Neuroblastoma European Study (INES) 99.3 (see page 35 for

a definition of stage M) obtaining excellent OS and EFS without megatherapy [3].

Histology is essential for determining the study group of the L2 patients. L2 patients without a

defined histotype should be treated according to the guidelines outlined in section 10.4.5 (page

91). All patients need a genomic profile result but their treatment will not be dependant on this.

The results of the genomic profile will be analysed to determine if it is a prognostic indicator in

any of these intermediate risk patients.

10.2 Eligibility

Common Inclusion criteria for all the studies: INFORMED CONSENT AND FOLLOW-UP WARRANTED




* For the purposes of this study, dexamethasone will not be considered chemotherapy and may be administered prior

to initiating trial therapy, at the discretion of the investigator.

INTERMEDIATE RISK STUDY INCLUSION CRITERIA EXCLUSION CRITERIA

(L1) INSS1 Localised tumours with complete gross

excision, with or without microscopic

residual disease; representative

ipsilateral lymph nodes negative for

tumour microscopically (nodes

attached to and removed with the

primary tumour may be positive)



profiling according to guidelines

(pages 122-123)

MYCN amplified

MYCN non-amplified




L2 Locoregional tumour with presence of

one or more IDRFs


and confirmed in a NRL:

differentiating, poorly

differentiated, undifferentiated

NB or ganglioneuroblastoma

nodular



(pages 122-123)

MYCN non-amplified

>18 months

Neuroblastoma NOS

MYCN amplification

≤ 18 months

M Distant metastatic disease with

positive MIBG (except Stage Ms).In

infants ≤12 months, MIBG or

technetium scintigraphy uptake to the

skeleton to be confirmed with a bone

abnormality visible on X-ray and/or CT

scan.




pages 122-123)

MYCN non-amplified

≤ 12 months

MYCN amplification

> 12 months

NOTE: L2 patients without a defined histotype will not be eligible for the study. However, these

patients should still be registered in the trial and the required information submitted. See section

10.4.5 (page 91) for treatment recommendations for this group of patients.



10.3 Study design

10.3.1 Intermediate Risk Study: L2 Patients (>18 months)




L2 > 18 months

Treatment Group 7 INRG Stage L2 differentiating NB or differentiating ganglioneuroblastoma

nodular*

Treatment Group 8 INRG stage L2 undifferentiated or

poorly differentiated NB*

2 VP/Carbo 2 VP/Carbo

Evidence of response: 2 VP/Carbo

No response: 2 CADO

2 CADO

IDFR positive: chemotherapy**

(VP/Carbo+CADO or 2 CADO), followed by surgery

IDFR negative: Surgery followed by

chemotherapy** (VP/Carbo+CADO or 2 CADO)

Local Radiotherapy Surgery if IDRF negative.

If IDRF positive discuss with Surgical Coordinators

6 courses of 13-cis-retinoic acid

* For patients without a defined histotype see section 10.4.5 (page 91) for treatment recommendations.

** Patients who responded to first VP-Carbo at this point receive 1

course of VP-Carbo and 1 of CADO

Patients who did not respond to first VP-Carbo at this point receive 2 courses of CADO



10.3.2 Intermediate Risk Study: (L1) INSS 1

Treatment Group 9

INSS Stage 1 MYCN amplified

2 VP/Carbo

2 CADO

2 courses of chemotherapy VP/Carbo+CADO

Local Radiotherapy

6 courses of 13-cis-retinoic acid



10.3.3 Intermediate Risk Study: M (≤ 12 months)

Contact trial coordinator for further

treatment options

2 VP/CARBO

Responding disease Progression or/and no response

2 VP/CARBO

2 CADO Metastatic CR (liver excluded)

Persistent metastases


Metastatic CR (liver excluded)

2 CADO

Metastatic CR (liver excluded)


Surgical resection of primary

Treatment Group 10 INRG Stage M ≤12 months



10.4 Treatment Strategy for intermediate risk patients







> 18 months

differentiating neuroblastoma

or

differentiating ganglioneuroblastoma nodular

Reassessment

2 x VP/Carbo

Response

2 x CADO 2 x VP/Carbo

No response

Reassessment

IDRF +ve

Discuss with National and

International Surgical

Coordinators.

IDRF -ve Surgical

resection




L2, >18 months, Differentiating NB or differentiating ganglioneuroblastoma nodular


WEEK Dx 1 4 7 10 13

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

PATHOLOGY

CONFIRMED

*

CHEMOTHERAPY

Cycle 1 2 3 4


CADO • •

SURGERY if IDRF

negative†

*


Following 2 VP/Carbo; If evidence of response give another 2 VP/Carbo (total 4 cycles)

Following 2 VP/Carbo; If NO evidence of response then give 2 CADO (total 4 cycles)

†Surgical resection is indicated if IDRF negative. If IDRF positive, discuss with Surgical Coordinators.




OR OR




Eligible patients

Stage L2, age >18 months, MYCN non-amplified, differentiating neuroblastoma or differentiating


Treatment

Treatment begins with two courses of VP/Carbo followed by clinical and imaging reassessment:

Patients with evidence of response (an objective reduction in tumour size): two more courses

of VP/Carbo.

Patients with no evidence of response (an objective reduction in tumour size): two courses of

CADO.

After four courses all the patients will be reassessed for resectability.

The total number of chemotherapy courses in this group is 4. Radiotherapy is not indicated for

patients in this group.


modification.

Surgical management

If after four courses IDRFs are not present treatment will be completed with tumour excision. If

IDRFs persist discuss the possibility of surgery despite IDRF with National or International

Surgical coordinators. If a tumour remains IDRF positive after chemotherapy this is not an

absolute contraindication to surgery. Resection may still be recommended if evaluation of the

primary tumour suggests that the risk to life, or of major functional loss, is less than the risk from

leaving residual disease. The decision making process needs to be documented in the study file.



After the first 2 courses of chemotherapy a reassessment is required, with clinical and ultrasound

evaluation.

After 4 courses of chemotherapy a CT scan or MRI is required to assess disease response.


Definition of relapse/progression:


2. Metastatic progression is defined by the appearance of distant disease (any localisation).

Clear documentation of progression and or relapse will be required in the clinical trial forms.

Management of progression

Localised progression: these patients should be discussed with the Study and Surgical

Coordinators in order to explore the feasibility of tumour resection.

Metastatic progression: these patients should be considered for high-risk treatment or for

a phase 2 treatment.



Study group 8






> 18 months

poorly differentiated and undifferentiated neuroblastoma

or poorly differentiated and undifferentiated ganglioneuroblastoma nodular

Reassessment

2 x VP/Carbo

2 x CADO

Reassessment

IDRF +ve

and response

IDRF –ve

OR

IDRF +ve and

no response

1 x VP/Carbo if there was a

response during the first two

VP/Carbo courses

otherwise

1 x CADO

1 x CADO

Reassessment

Radiotherapy

6 courses of

13-cis-RA

1 x VP/Carbo if there was a

response during the first two

VP/Carbo courses

otherwise

1 x CADO

1 x CADO

Surgical resection

Discuss with Surgical

Coordinators if +ve IDRF

cause concern.

Surgical resection

Discuss with Surgical

Coordinators if +ve IDRF

cause concern.

Reassessment




L2, >18 months, Undifferentiated or poorly differentiated NB or undifferentiated or poorly

differentiating ganglioneuroblastoma nodular


WEEK Dx 1 4 7 10 13 17 21

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

PATHOLOGY

CONFIRMED

*

CHEMOTHERAPY

Cycle 1 2 3 4 5 6

VP/Carbo • • •

CADO • • • •

SURGERY† * *

LOCAL

RADIOTHERAPY

•

13-CIS-RA x6 •


†Week 13, following 2 cycles VP/Carbo and 2 CADO:

If IDRF negative or positive IDRF with no response: surgery and then chemotherapy (2 cycles)**

If IDRF positive with response: chemotherapy (2 cycles)** and then surgery

** Patients who responded to initial VP/Carbo receive 1 course of VP/Carbo and 1of CADO

Patients who did not respond to initial VP/Carbo receive 2 courses of CADO




OR




Eligible patients

Stage L2, age >18 months, MYCN non-amplified, poorly differentiated and undifferentiated

neuroblastoma or poorly differentiated and undifferentiated ganglioneuroblastoma nodular.

Treatment

All patients will receive 6 courses of chemotherapy:

Courses 1 and 2: VP/Carbo

Courses 3 and 4: CADO

Course 5: VP/Carbo or CADO (VP/Carbo if the patient responded with courses 1 and 2.

CADO if the patient did not respond or progressed with courses 1 and 2).

Course 6: CADO in all patients.


modification.

Patients within this group will also receive radiotherapy followed by six courses of 13-cis-RA.

Radiotherapy will be given following completion of chemotherapy or surgical resection,

which ever occurs later. Radiotherapy to the pre-surgery tumour volume should be

administered according to the guidelines in Appendix 11 (page 145).

13-cis-RA will be administered according to Appendix 16 (page 169).

Surgical management

If after four courses of chemotherapy IDRFs are not present or there has been no response (and

consequently IDRF still positive) surgical excision should take place. Patients who complete 6

courses of chemotherapy prior to surgery will have a surgical excision following the sixth course.

In cases where IDRFs cause concern discuss with the Surgical Coordinators. If a tumour remains

IDRF positive after chemotherapy this is not an absolute contraindication to surgery. Resection

may still be recommended if evaluation of the primary tumour suggests that the risk to life, or of

major functional loss, is less than the risk from leaving residual disease. The decision-making-

process needs to be documented.



After the first 2 courses of chemotherapy a reassessment is required, with clinical and ultrasound

evaluation. If US reassessment is unclear then CT or MRI is required.

After 4 courses of chemotherapy a CT scan or MRI is required to assess disease response. If the

reassessment after four courses of chemotherapy shows:

disease progression (event), the National Coordinator should be contacted for a Phase 2

treatment.

any response and IDRF negative: surgery, followed by chemotherapy (courses 5 and 6).

any response and IDRF positive: further chemotherapy (courses 5 and 6).

no response (and consequently IDRF still positive): surgery, followed by chemotherapy

(courses 5 and 6). In cases where IDRF cause concern discuss with the Surgical

Coordinators. The decision-making-process needs to be documented.

After the 6 courses of chemotherapy, if surgery has not been performed, IDRF are evaluated

again:

If IDRF negative, surgery.



If IDRF positive, surgery. In cases where IDRF cause concern discuss with the Surgical

Coordinators. If a tumour remains IDRF positive after chemotherapy this is not an

absolute contraindication to surgery. Resection may still be recommended if evaluation of

the primary tumour suggests that the risk to life, or of major functional loss, is less

than the risk from leaving residual disease. The decision-making-process needs to be

documented.

Reassessments to evaluate disease response should also be carried out two months after the

completion of radiotherapy and then once more 12 weeks later.




2. Metastatic progression is defined by the appearance of distant disease (any localisation).

Clear documentation of progression and or relapse will be required in the clinical trial forms.


In the case of local progression after the initial 2 courses of VP/Carbo, the child should receive the

scheduled chemotherapy (i.e. 2 courses of CADO). In the case of metastatic progression after the

initial 2 courses of VP/Carbo, the child should be considered for treatment in the high risk study.

If the progression appears later, the Study Coordinator should be contacted for a phase 2

treatment.





(L1) INSS Stage 1,

MYCN amplified

Any age

Reassessment

2 x VP/Carbo

2 x CADO

Reassessment

1 x CADO

Reassessment

Radiotherapy

6 courses of

13-cis-RA

1 x VP/Carbo




INSS stage 1, MYCN amplified


WEEK Dx 1 4 7 10 13 17 21

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

MYCN status

CONFIRMED

*

CHEMOTHERAPY

Cycle 1 2 3 4 5 6

VP/Carbo • • •

CADO • • •

SURGICAL Excision *

LOCAL

RADIOTHERAPY

•

13-CIS-RA x6 •





Eligible patients

INSS Stage 1 neuroblastoma, MYCN amplified.

NOTE: If no enlarged lymph nodes are noticed during surgery and thus lymph nodes are not

examined this is still considered INSS Stage 1.

Treatment

All patients will receive 6 courses of chemotherapy:

Courses 1 and 2: VP/Carbo

Courses 3 and 4: CADO

Course 5: VP/Carbo

Course 6: CADO


modification.

After completing 6 courses of chemotherapy, patients within this group will receive radiotherapy

followed by six courses of 13-cis-RA.

Radiotherapy will be given after 6 courses of chemotherapy, radiotherapy to the pre-

surgery tumour volume should be administered according to the guidelines in Appendix 11

(page 145).

13-cis-RA will be administered according to Appendix 16 (page 169).

Surgical management

Surgical resection is undertaken at diagnosis in this group of patients.


Following every two courses, clinical assessment and ultrasound in abdominal cases should be

carried out. At completion of therapy a CT or MRI scan and MIBG are required.

Reassessments to evaluate disease status should also be carried out two months after the

completion of radiotherapy and then once more 12 weeks later.



Progression is defined as any new localisation of disease either local or metastatic. Clear

documentation of progression will be required in the clinical trial forms.


In the case of local progression after the initial 2 courses of VP/Carbo, the child should receive the

scheduled chemotherapy (i.e. 2 courses of CADO). In the case of metastatic progression after the

initial 2 courses of VP/Carbo, the child should be considered for treatment in the high risk study.

If the progression appears later the Study Coordinator should be contacted for a phase 2 treatment.





Stage M, MYCN non-amplified

≤ 12 months

any histology

2 x VP/Carbo

Reassessment

Metastatic CR

(liver excluded)

Response

Surgical

resection

No response and/or

progression

2 x CADO

Reassessment

Discuss with National and/or

International Coordinators.

2 x VP/Carbo

Reassessment Persistent

metastases

Persistent

metastases

2 x CADO

Reassessment

Persistent

metastases




INRG stage M, ≤12 months


WEEK Dx 1 4 7 10 13 17 21

Date

PRE-TREATMENT

EVALUATION

*

IMAGING * * * * *

BIOPSY *

TOXICITY

MONITORING

*

*

*

*

*

PATHOLOGY

CONFIRMED

*

CHEMOTHERAPY

Cycle 1 2 3 4 5 6 7 8


CADO • • • • • •

SURGERY† * * *


Week 7, following 2 cycles VP/Carbo:

Responding disease: give another 2 cycles VP/Carbo

Progression or/and No-response: give 2 cycles CADO

†Following the 4th, 6th or 8th cycle of chemotherapy, if metastatic CR (liver excluded):

Surgical resection of the primary should be undertaken; there is no indication for further chemotherapy.

If following:

6 cycles of chemotherapy (2 VP/Carbo and 4 CADO --

patients with no response on week 7, following 2 VP/Carbo) or

8 cycles of chemotherapy (4 VP/Carbo and 4 CADO--

patients with response on week 7, following 2 VP/Carbo)

metastases persist, contact trial coordinator for further treatment options. Consider HR-group therapy

(including MAT).

OR OR




Eligible patients

Infants (≤12 months) with stage M neuroblastoma, without MYCN amplification

Treatment

These infants will start treatment with two courses of VP/Carbo.

Providing there is evidence of response they should then receive a further 2 courses of

VP/Carbo.

If metastatic CR has not been achieved after 4 courses of VP/Carbo then up to 4 courses of

CADO should be given.

Infants with disease progression or no evidence of response after 2 courses of VP/Carbo

should switch to CADO.

If there is residual metastatic disease after 2 courses of CADO a further 2 courses should be

given, with a further reassessment after these.


modification.

There will be no additional treatment given following surgery, unless there is disease progression.

In particular it is NOT recommended that radiotherapy is given to residual macroscopic primary

tumour.

Surgical management

If after four courses of chemotherapy the treatment has resulted in metastatic CR then surgery to

the primary should be carried out. The definition of metastatic CR considers a response of all

metastatic sites involved at diagnosis except for the liver.

For patients requiring extra courses of CADO, surgery should only be carried out once the patient

is in metastatic CR. If after a total of 4 courses of CADO the infant still has evidence of metastatic

disease, it is recommended that further treatment options (including any Phase II studies) be

discussed with the National coordinator or the Trial coordinator.

If a tumour remains IDRF positive after chemotherapy this is not an absolute contraindication to

surgery. Resection may still be recommended if evaluation of the primary tumour suggests

that the risk to life, or of major functional loss, is less than the risk from leaving residual disease.

If IDRF persist after metastatic CR, resection should still be carried out; discuss with National or

International Surgical coordinators if in doubt.



After 2 courses an assessment of disease response should be undertaken. If this shows

response then a further two courses of VP/Carbo are given. If no response is seen then two

courses of CADO are given.

After 4 courses a full assessment of all involved sites should be undertaken. If this shows the

patient to be in metastatic CR then surgery to the primary tumour should be carried out at this

time. The definition of metastatic CR considers a response of all metastatic sites involved at

diagnosis except for the liver.

Assessment of response to CADO should be carried out after 2 courses, and if metastatic CR

is achieved surgery to the primary can be undertaken at that time.






2. Metastatic progression is defined by the appearance or increase in size of any bone, pleura-

lung, and/or CNS metastasis.

Clear documentation of progression and or relapse will be required in the clinical trial forms


In the case of progression (local or metastatic) after the initial 2 courses of VP/Carbo, the child

should receive the scheduled chemotherapy (i.e. 2 courses of CADO). If the progression appears

later, the Study Coordinator should be contacted for a phase 2 treatment.

10.4.5 Treatment recommendations for patients with stage L2, age >18 months, without a defined

histotype (i.e. neuroblastoma NOS)

Certain patients with neuroblastic tumours are not able to be classified with a clear definition of

the histotype and are considered as neuroblastoma NOS (not otherwise specified). In these cases it

is recommended that another biopsy be taken in order to obtain a more accurate diagnosis and

consequently stratify the patient to the appropriate study group.

If a second biopsy is not available or gives the same result (NOS) the patient cannot receive

treatment stratified by histotype and should receive standard “historic” treatment: 6 courses of

chemotherapy and surgical resection as in study group 8, with no radiotherapy and no 13-cis-RA.

These patients should still be registered and will be “study patients” (rather than “trial

patients”), patient status data (according to section A18.8, page 176) will be collected on

them regularly but they will be analysed separately.

These patients must be consented using the “without histotype” consent form (page 206).

10.4.6 Chemotherapy and/or surgical procedures outside the recommendations of this protocol

In order for a complete treatment picture to be drawn for these patients, it is important that all

chemotherapy and/or surgical procedures are entered into the LINES database using the

appropriate chemotherapy and/or surgery CRFs via the SIOPEN-R-NET. This should be done

even if the chemotherapy and/or surgical procedure is not within the recommendations of this

protocol.



11. INVESTIGATIONS DURING AND AFTER TREATMENT

11.1 Minimal Mandatory Investigations during and at the end of treatment

After Course Number

At

Dx

1

2

3

4

5

6

End Of

therapy

Complete Blood Count

Biochemistry

Serum LDH,

Renal function evaluation Including the Schwartz's formula

Urine catecholamines

Biological studies

MYCN

DNA index

Array-CGH / MLPA

Bone marrow aspirates

& trephine biopsy

Stage

M

Stage M

Ultrasound, CT or MRI

*

+

Scintigraphy MIBG (Tc MDP)

Stage

M

X-ray of any bone uptake of

MIBG

Audiogram (for patients receiving

carboplatin)

Echocardiogram (for patients

receiving anthracycline)

* Preferable to use US to evaluate response. + Evaluate with CT and /or MRI, comparable with imaging at diagnosis



11.2 Investigations after completion of treatment

11.2.1 Tumour Assessment/Detection

As a minimum, imaging of the primary tumour site should be evaluated every 12 weeks in the

first year after completion of treatment and then annually for 5 years.

11.2.2 Toxicity Assessment

Renal Follow-Up: GFR should be assessed at 2 years and 5 years from diagnosis in patients

considered to be at risk of renal toxicity.

Auditory Follow-Up: Audiometry should be assessed at 2 years and 5 years from diagnosis in

those treated with carboplatin.

Cardiac Follow-Up: Echocardiography should be carried out 2 years, 5 years and 10 years

from diagnosis in those patients that received an anthracycline.



12. LONG TERM FOLLOW UP

Long term follow-up in children with neuroblastoma should follow the different national policies

for LONG TERM FOLLOW-UP in children cured of a paediatric cancer. Minimal mandatory

recommendations are the following:

12.1 Tumour assessment

In an asymptomatic patient, the following are recommended:

1. Full Clinical examination (including blood pressure)

2. Imaging of the primary site (x-ray, ultrasound, or CT or MRI scan as appropriate), at least

once a year.

3. Metastatic assessment: In case of residual skeletal MIBG positive, repeat MIBG scan

every 3 months until negative or progression. If stable over 1 year, repeat it on a yearly

basis for up to 5 years.

12.2 Toxicity assessment

The toxicity assessment needs to be related to the treatment received by the patient, with regards

to renal, audiological and cardiological follow-up. Those who have had extensive abdominal or

pelvic radiotherapy may have prolonged thrombocytopenia.

12.2.1 Renal Follow-Up

Annual blood pressure measure at out-patient clinic.

GFR should be assessed at 2 years and 5 years from diagnosis. In children who can give a

reliable 24 hour urine collection, endogenous creatinine clearance is acceptable. Where this is

not possible, then GFR estimation by DTPA or CrEDTA is preferred. Children who had an

end of treatment GFR of less than 80ml/min/1.73m² should have a repeat GFR and serum

magnesium 5 years off treatment.

DMSA scan at 5 years in patients having received abdominal irradiation.

12.2.2 Auditory Follow-Up

Ototoxicity is usually permanent or irreversible. An adequate assessment at the end of treatment is

strongly advised, and audiometry should be performed at 2 years and 5 years from diagnosis for

those children treated with carboplatin.

If the child has sudden severe hearing loss which is not only a high-frequency loss then serious

otitis media should be excluded and the audiometry repeated after 6 months.

12.2.3 Cardiac Follow-Up

In this protocol, the cumulative dose of anthracycline does not exceed 240 mg/m2.

Echocardiography at 2 and 5 year from diagnosis for children treated with anthracycline is

recommended.

12.2.4 Etoposide: second malignancy follow up

It is essential that any second malignancy occurring amongst the children treated with

chemotherapy is registered urgently with the data centre.



13. STATISTICAL CONSIDERATIONS

The majority of studies comprise a single arm strategy the success or otherwise of which will be

judged by reference to historical information that is summarised for each study below. It is also

recognised, even with wide multinational collaborations, that the numbers of patients within the

various subgroups will not be sufficient to base sample size calculations on conventional methods

of test size and power. Hence the general objective is to obtain as many patients as possible within

a reasonable time frame (initially set at 5 years and thereby to estimate the endpoint statistic, with

a two-sided 95% confidence interval, for the respective endpoint measures and use these as the

basis for conclusions in the single arm parts of the protocol. In addition, adopting a Bayesian

approach, the probability that the endpoint of concern is worse than the specified target will be

calculated. Also, a general requirement is to monitor the individual studies with respect to

unacceptably high event rates so guidelines for this are provided for each study within the

protocol for the data monitoring committee (DMC) who will review the data on an annual basis. It

is anticipated, should any of the guidelines indicate unacceptably high event rates, that the DMC

will recommend an appropriate course of action to the trial steering committee. In the majority of

studies, event free survival (EFS) is the endpoint of concern and will be estimated by the Kaplan-

Meier technique. This is defined as the time from the date of diagnosis (or the date of

randomisation for study group 1) to the date of the first event, that is, progressive disease,

recurrence or death. A definition of progressive disease can be found in the treatment strategy

sections (LR: section 9, page 43 and IR: section 10, page 73) under the appropriate study group.

13.1 Low Risk Study

13.1.1 Stage L2

Total anticipated recruitment number: 240

13.1.1.1 Group 1

NCA genomic profile, No LTS

Randomisation:

(1) Chemotherapy (CO 2-4 courses ± VP/Carbo 2 courses) – If the tumour becomes IDRF

negative after 2, 4, or 6 courses then surgery is performed and no further chemotherapy is

given.

(2) Observation: observed with regular assessments for 1 year, with surgery indicated only if

IDRFs are negative at 1-year after diagnosis. If progression with LTS occurs then

chemotherapy (VP/Carbo 2 courses ± CADO 2 courses) is initiated.

Randomisation will be stratified according to country.

Number of patients anticipated in 5-years: 150.

Objectives: To maintain an OS of 100% with less treatment than has been given historically. The

treatment burden will be less in some patients who will only be given chemotherapy if

progression occurs and chemotherapy may be avoided completely in others. The primary endpoint

is the time of initiation of VP/Carbo administration during the first year following diagnosis.

Trial size: It is anticipated that the 1-year rate of initiation of VP/Carbo will be approximately

55% in the chemotherapy group and possibly less than 30% in the observation group. With a two-

sided test size of 5% and power 85% a trial of 150 patients will be sufficient to detect this

reduction of 25%.



Monitoring: In order to monitor the possibility of an increased number of events this trial will be

formally reviewed by an independent Data Monitoring Committee should the 1-year death rate

exceed 1% at any time during the course of the trial in either arm of the trial.

In this randomised trial, it will not be possible to determine reliably whether an OS of 100% is

maintained in the experimental arm given the very low anticipated event rate. A hazard ratio will,

if possible, be calculated but the CI is likely to be very wide and hence uninformative. Adopting a

Bayesian approach, the probability that outcome is more than X% worse in the experimental arm

than in the standard arm will be given for a range of value of X. Clinical judgement can then be

used to decide whether OS in the experimental arm is acceptable.

13.1.1.2 Group 2

NCA genomic profile 1, with LTS

Single arm: Chemotherapy (VP/Carbo) to eliminate Life Threatening Symptoms (LTS) – surgery

if /when IDRF negative.

Number of patients anticipated in 5-years: 60

Objective: To maintain a 2-year EFS of at least 90% and an OS of at least 95%

Monitoring: In order to monitor the possibility of an unacceptable increased number of events the

trial will be formally reviewed by an independent Data Monitoring Committee should the

cumulative 1-year event rate exceed 10% at any time during the course of the study.

13.1.1.3 Group 3

SCA genomic profile, irrespective of LTS

Single arm: Chemotherapy (VP/Carbo) – surgery if/when IDRF negative


Objective: To improve the 2-year EFS to at least 90% and maintain the OS of close to 100%

Monitoring: In order to monitor the possibility of an unacceptable increased number of events the

trial will be formally reviewed by an independent Data Monitoring Committee should the

cumulative 1-year event rate exceed 10% at any time during the course of the study.

13.1.2 Stage Ms


13.1.2.1 Group 4

NCA genomic profile, No LTS

Single arm: Observation only as no LTS present. Chemotherapy is given if an event occurs

(progression or the development of symptoms). A surgical resection of the primary mass is not

recommended. If symptoms develop chemotherapy with VP/Carbo is given as in group 5.


Objectives: Maintain 2-year EFS over 85 % and OS of at least 98%.



Monitoring: In order to monitor the possibility of an increased number of events the trial will be

formally reviewed by an independent Data Monitoring Committee should the cumulative 1-year

event rate exceed 10% or the 1-year death rate exceed 1% at any time during the course of the

study.

13.1.2.2 Group 5

NCA genomic profile, with LTS

Single arm: chemotherapy with no change as compared to previous practice. Treatment with

minimal number of courses of chemotherapy VP/Carbo 2-4 or VP/Carbo 2 and CADO 2 if the

symptoms do not respond to VP/Carbo, to resolve to resolve LTS. A surgical resection of the

primary mass is not recommended.


Objective: Maintain a 2-year EFS over 85% and OS of at least 98%




study.

13.1.2.3 Group 6

SCA genomic profile, irrespective of LTS

Single arm: Chemotherapy: Patients with no LTS receive VP/Carbo x 4 courses. Patients with

LTS will receive VP/Carbo 2 courses followed by CADO 2 courses if LTS do not resolve or a

further 2 courses of VP/Carbo. Surgery is recommended if primary mass is IDRF negative.


Objectives: Aim to improve 2-year EFS to over 70%. Retrospective data show EFS of 60% in

this group. It would be clinically worthwhile to improve this 2 year EFS to 70% with the addition

of upfront VP/Carbo chemotherapy.



event rate exceed 30% at any time during the course of the study.

13.2 Intermediate Risk Study

13.2.1 Stage L2


13.2.1.1 Group 7

Age >18 months, differentiating neuroblastoma or differentiating ganglioneuroblastoma nodular

Single arm: Chemotherapy (4 courses of VP-Carbo or 2 courses of VP-Carbo and 2 of CADO)


Objectives: Maintain 3-year EFS of 90% with 3-year OS at 100%




specifically reviewed by the independent DMC should the cumulative 1-year event rate exceed

5% or the 1-year death rate exceed 1% at any time during the course of the study.

13.2.1.2 Group 8

Age >18 months, undifferentiated and poorly differentiated neuroblastoma or undifferentiated and

poorly differentiated ganglioneuroblastoma nodular

Single arm: Immediate Chemotherapy (2 courses of VP-Carbo and 2 of CADO). Surgery if IDRF

negative. Delayed chemotherapy (1 course of VP/Carbo and 1 of CADO, or 2 courses of CADO).

Surgery if not obtained before. Radiotherapy. 13-cis-RA: 6 courses.


Objectives: To improve 3-year EFS to 70% with 3-year OS at 90%




13.2.2 INSS stage 1


13.2.2.1 Group 9

MYCN amplified neuroblastoma

Single arm: Adjuvant chemotherapy (6 courses: 3 of VP-Carbo and 3 of CADO). Radiotherapy.

13-cis-RA: 6 courses.


Objectives: To improve 3-year EFS at least to 50% with 3-year OS at 80%




13.2.3 Stage M


13.2.3.1 Group 10

Age < 12 months

Single arm: Immediate chemotherapy (4-8 courses of VP-Carbo and CADO) with the aim to

obtain metastatic remission and IDRF negative. Surgery after at least 4 courses, if IDRF negative

and metastatic remission achieved.


Objectives: Maintain 3-year EFS of 85% with 3-year OS at 95%




study.



14. NEONATAL SUPRARENAL MASSES: AN OBSERVATIONAL

APPROACH

14.1 Background

The incidence of suprarenal tumours/masses has increased in the last decade due to the expanded

use of prenatal ultrasonography in routine obstetric care and in the neonatal and early infancy

care. The differential diagnosis of these masses ranges from benign (adrenal haemorrhage) to

malignant processes (neuroblastoma, adrenal carcinoma). Knowledge on perinatal suprarenal

masses, although based on a relatively large literature, is scattered amongst studies on very few

cases with no methodical approach and often short follow up [48-50]. Therefore, the optimal

management of these masses has not been clearly defined. Neuroblastoma at this age is an

intriguing entity with a very good prognosis in most cases. The SIOPEN Group, based on their

results in the first multicenter European Trial for infants with neuroblastoma (INES) and the

world-wide experience provided in the literature, is launching this European surveillance study for

these masses.

14.1.1 Review of anatomy and embryology [51]

The adrenal glands may be visible by ultrasound as early as at 20 week gestation and usually are

distinguishable by 30 weeks in the majority of the foetuses. The glands appear lenticular in the

transverse plane and pyramidal in the longitudinal one, with a hypoechoic rim around and an

echogenic central stripe. The foetal gland, larger than the one in the adult, constitutes 0.2% of the

total body weight in the foetus. At birth, the glands are located anteromedially to the superior pole

of their corresponding kidney, covered with perirenal fat and encased in Gerota´s fascia. The

blood supply is derived primarily from 3 sources (the phrenic artery superiorly, the aorta medially

and the renal artery inferiorly) and the venous drainage goes to the inferior cava (right adrenal)

and left renal vein (left adrenal). The lymphatic drainage also differs for the 2 glands (periaortic

lymph nodes for the right and accompanying the venous drainage for the left one).

The adrenal parenchyma is divided into the inner medulla and the outer cortex. The medulla

originates from the ectodermal chromaffin cells of the neural crests (neuroblasts) that eventually

differentiate and organize into many small ganglia that may secrete epinephrine and

norepinephrine in response to neural stimuli. Other chromaffin cells differentiate from the adrenal

precursors and migrate ventrally to form the aortic paraganglia. Cortical cells are derived from the

splachnic mesoderm that arises in the adrenogenital ridge early in gestation. Medullary or cortical

cells may fail to migrate completely and may develop into extra-adrenal rests, commonly in the

kidney but also along the route of descent of the gonads. Approximately half of newborns have

adrenocortical rests that usually involute within a few weeks. The dual embryonic origin of the

adrenal gland explains the broad spectrum of tumour histologies that may present as suprarenal or

para-adrenal masses in this period of life.

14.1.2 Perinatal tumours and foetal diagnostic imaging

The reported prevalence of perinatal tumours ranges from 1.7-13.5 per 100,000 live births.

However, those in foetuses that are stillborn are probably underreported [52]. Congenital

neuroblastoma is one of those tumours with a tendency to regress and a benign clinical course

despite a clear malignant histological picture. Foetal diagnostic imaging continues to develop and

improve allowing more accurate diagnosis of foetal tumours [53], appropriate surveillance and

timing of intervention and delivery [54]. The vast majority of foetal tumours are detected by

obstetric US during the second and third trimesters. The technique is helpful in determining

location, content, relationship to surrounding structures and the presence of heart failure, hydrops

or associated malformations. Nevertheless, complementary examinations are necessary. Foetal

MRI has shown the potential to provide additional information in the work-up of tumours detected

by US. Foetal MRI defines the exact extent of the tumour better than US, also complications



within the tumour or caused by the tumour. A multidisciplinary approach with access to a tertiary

referral centre is advisable to optimise foetal outcome.

14.1.3 Differential diagnosis

Adrenal haemorrhage (AH): This may present with physical and radiographic findings that are

indistinguishable from other suprarenal masses. The incidence of adrenal haemorrhage is nearly 2

cases per 1000 live births; however, advances in prenatal ultrasonographic imaging may increase

this figure. Because the majority of these haemorrhages spontaneously resolve it is important to

clearly distinguish them from other suprarenal masses, in particular neuroblastoma, which can

spontaneously resolve in the same time frame. When the adrenal haemorrhage is diagnosed in

utero, the lesion is characterized by an echogenic mass, and it is more frequent on the right side.

On follow-up ultrasound the mass appears increasingly hypo-echoic and usually involutes. Many

lesions completely resolve, leaving only residual minor calcifications. However, the cyst may not

regress and then other explorations might be necessary. If the diagnosis is not made antenatally,

the infant rarely may present signs/symptoms of adrenal insufficiency. Most of the times, there

will be hyperbilirubinemia, anaemia, scrotal hematoma. Boys are affected more frequently and the

right side also more commonly. The aetiology is unknown, being related to birth trauma, foetal

hypoxia, maternal hypotension and sepsis; but these factors might not be present at all. It has also

been related to vascular factors (venous congestion, arterial insufficiency). Adrenal haemorrhage

can be sequentially followed-up by ultrasound, without harming the baby [55].

Enteric duplication cysts may present in the suprarenal location. However, they appear as early

as the 16th week, allowing some means of discrimination from cystic NB [56].

Subdiaphragmatic extralobar pulmonary sequestration (SEPS): They are important to

distinguish from neuroblastomas because they may not require surgery. These rare congenital

lung abnormalities consist of non-functioning pulmonary tissue that lack tracheobronchial

communication and receive their blood supply from anomalous systemic arteries. They may be

intra or extralobar. These latter ones occur between the diaphragm and the lower lobe of the lung.

The majority of them are diagnosed before the age of 10 and frequently associated with congenital

abnormalities such as diaphragmatic hernias. The typical sonographic appearance suggests a solid

mass, which persists with stable size postnatally most of the times and occasionally regresses or

grows.

Adrenal cytomegaly [57]: This phenomenon is not well known yet. It is characterized by the

presence of large polyhedral cells with eosinophilic granular cytoplasm and enlarged nuclei in the

adrenal cortex. It is thought to be a degenerative process, but not a malignancy. It rarely forms

cysts and, can be detected during prenatal ultrasonographic examinations.

Adrenocortical tumours: The incidence of adrenocortical tumours is approximately 3 per

million and these tumours comprise less than 1% of all paediatric neoplasms. Most of them are

functional and may be detected easily if there is sufficient clinical suspicion (hypertension,

precocious puberty) though patients often present with a large abdominal mass. These tumours are

more common in girls and might be associated with other syndromes like Beckwith-Wiedeman,

hemihypertrophy. The family history may include adrenal carcinoma or other cancers (Li-

Fraumeni, Wilms, hepatocellular carcinoma) or not, since most likely they occur sporadically. The

diagnosis of a functioning adrenocortical tumour can be confirmed by a myriad of biochemical

tests. Free cortisol levels in 24 hour urine are the most common screening test.

Other rare diseases: congenital adrenal hyperplasia, can present with an increase in adrenal size

and other signs/symptoms; pheochromocytoma and paraganglioma have rarely described in

neonates. Hydronephrosis, multicystic dysplasia of the upper pole of a duplex collecting system.

Neonatal adrenal abscesses should also be taken into account in the differential diagnosis.

Neuroblastoma: see below.



14.1.4 Neuroblastoma biology

14.1.4.1 Benign behaviour

Neuroblastoma is the most common solid abdominal tumour in children, with a very intriguing

biology. Prognosis is related to age, stage and biological factors such as MYCN amplification. It is

well-known its favourable course in early infancy (neuroblastoma in situ, screening programmes).

The European experience with the first multicentre study has shown its excellent good prognosis

for all stages, including metastatic ones, with none or little treatment (surgery + mild

chemotherapy), except for the very rare and scarce group of infants suffering from non localised

neuroblastoma with MYCN amplification. Experiences conveyed by American, Japanese and

German groups are similar [11, 58, 59].

14.1.4.2 Neuroblastoma in the pre and neonatal period [60-65]

Obstetric ultrasonography has been used over the last few decades with increasing frequency and

precision. As technology and protocolised routine examinations have improved, the finding of

suprarenal masses in this population has steadily increased. Although there are not exact figures

on the general incidence of suprarenal masses in this population, we know that the detection of

neuroblastoma at this age has risen also steadily. More than 55 cases of antenatal neuroblastoma

have been reported since the first report by Fenart in 1983, and these figures underestimate the

real incidence of neuroblastoma at this age. These “foetal” neuroblastomas represent a unique

group of tumours with a prognosis and management that is different from that in older infants and

children.

More than 90% of the “so-called” foetal neuroblastomas are located in the adrenal glands and

almost half of them have a cystic component. They are usually discovered during a routine

obstetric ultrasound examination or during one in the first 3 months of age (usually performed for

the diagnosis of another suspected paediatric problem, i.e., hydronephrosis or the asymptomatic

finding of an abdominal mass). These tumours are usually located in the adrenal gland and their

sonographic image can vary from solid to cystic to hyperechoic to mixed foci of calcification.

Foetal adrenal NBs are usually found with US during the third trimester of gestation; two thirds of

them are located on the right side. 50% of tumours are solid isoechoic masses (diameter 2 to 4

cm). The remaining cases are either purely cystic hypo-anechoic masses or complex structures

containing both fluid and echogenic solid material. The latter group is typically larger in size,

measuring between 3 to 10 cm in their widest diameter. During foetal life it is common for the

solid masses to increase in size on serial antenatal ultrasounds exams, whereas the cystic and

complex masses may grow-off or shrink during foetal life [56].

The stage at presentation is usually stage 1 or 4s, and most of the isolated reports have shown a

very benign course, usually after resection for stage 1 and maybe some mild chemotherapy for

stage 4s. Nevertheless, neuroblastoma can metastasise in utero and placenta (maternal symptoms

of catecholamine excess and foetal hydrops) and there is a very small group of neonates with

unfavourable features [48, 49].

In the INES 99.4 population of infants with MYCN amplified neuroblastomas [10], there were 4

patients diagnosed in the first month of life, all of them were clearly metastatic at diagnosis (liver,

cutaneous, skull, and one with a general dissemination of the tumour shown in the autopsy).

MIBG was performed in 3 cases (Primary tumours: 1 weak +, 1 strong +, 1 negative; liver uptake

in one, no bone uptake) and bone marrow studies in 2 (evaluable and + in one). Survival: 1 alive

without evidence of disease after 99.4 strategy including megatherapy, 1 died in the first hours of

life (autopsy: neuroblastoma in all organs, including brain). For the great majority of patients

included in the INES study and trials 99.1, 99.2 for non-amplified stage 1, 2, 3 and 4s the survival

and disease free survival have been excellent as reported in different scientific forums [3, 10].



However, surgery performed at this early age and even mild chemotherapy may have secondary

effects, either then or later on. Although not studied in depth, the available literature suggests that

this type of surgery may have more risks in this young population: severe haemorrhage (up to

3%), respiratory failure, major vascular injury (4%), intestinal complications (infarction,

intussusception), and splenic injury. Renal and intestinal long-term complications may also

appear. Of course, anaesthetic procedures also have an overall risk of 0.43%, and even risk of

cardiac arrest. The overall mortality for young infants undergoing this surgery is greater than the

older counterpart [66-68].

14.1.4.3 Utility of imaging studies and biological markers in the neonatal period

Ultrasonography (US)

A few parameters may be considered: structure, size, vascularisation, change in size and structure

during observation.

Structure: It may be cystic, solid, mixed.

Cystic structure is associated with proved NB in 47% of cases, whereas it is associated with

presumed AH in 21 % of cases, and with presumed other conditions in the remaining 32% of

cases.

Solid structure is associated with proved NB in 74.5% of cases, with proved AH in 6% of cases

and with proved other conditions (mostly SEPS) in 19.5% of cases.

A mixed structure is associated with NB in 54% of cases, with the remaining cases associated

with presumed other conditions mostly including AH[69].

Size: Size does not appear to be a parameter suitable to differentiate AHs from NBs.

Change in size: Just a single study[69] deals with the “Change in Size” parameter. According to

this study, in a relatively large number of cases observed (24 newborns) enlargement and size

stability were associated with verified NB in the 86% and 91% of cases, respectively. Regression

of the size was always considered associated with presumed AHs. However, this study does not

report the length of the observation period in which size changes were observed.

Vascularisation: It has been proposed [55] that colour and power Doppler sonography may be

useful in differentiating between AH and perinatal NB. In a very limited series of cases it has been

observed that in most cases AHs do not show vascularisation on Doppler studies, whereas solid

NBs may show vascularisation. However, it also known that Doppler studies do not help in

differentiating an AH from an haemorrhage occurring within a cystic NB [60].

Changes in echogenicity: A neonatal solid suprarenal mass which changes its echogenicity while

it shrinks (usually: solid -> mixed -> cystic -> disappearing with possible residual calcifications)

has been traditionally associated with a presumed AH.

CT scan CT may be contributory in differential diagnosis when it shows a systemic vessel in a pulmonary

sequestration, in case US failed to show it [61]: however some pitfalls are well known. It may be

also helpful to the diagnosis of NB when identifying subtle calcifications within the mass or in the

adjacent retroperitoneum [69], and it is useful to evaluate local NB extension before surgical

excision. In case of cystic lesions, rim enhancement may suggest the possibility of cystic NB

[63]. However, it cannot be considered a routine imaging modality in a newborn with a suprarenal

mass because of its high radiation exposure.

MRI

In case of cystic lesions, MRI may be useful in determining the nature of the cyst content. Two

different intensities may be demonstrated within the mass, with debris-fluid levels suggesting

intracystic haemorrhage: however, MRI is unable to differentiate between adrenal haemorrhage

and haemorrhage occurring within a cystic NB [65]. Not differently form CT, MRI may be useful

when it demonstrates a systemic vessel within a solid suprarenal mass (possible Subdiaphragmatic

Extralobar Pulmonary Sequestration (SEPS)), although this finding is not constant and

pathognomonic of SEPS.



MIBG

MIBG is useful for both identification of a neuroblastic tumour as well as for detection of

metastatic disease. It has a very high (> 90%) positive predictive value for the presence of a NB

primary tumour, but its utility is limited by the fact that only less than 70% of perinatal NB are

MIBG avid and the negative predictive power is about 55%[69].

Urinary catecholamines

High levels of urinary catecholamines are helpful in confirming the diagnosis of NB. However, in

patients with perinatal NB, the sensitivity is 52% only, with a negative predictive power of 46%.

In patients with prenatally detected NB, urinary catecholamines will be normal in two thirds of

them, with a negative predictive power of about 70% [69].

See summary table below:

Negative Predictive

Power

Sensitivity

41%55%46%(post)

72%(pre)

21%70%52%(post)

36% (pre)

MRI/CTMIBGCatecho.

Differential Diagnosis

14.1.5 Summary

Based on all the above facts, there has been a considerable debate in the scientific community on

the best management of suprarenal masses discovered ante or neonatally. There are currently

several groups conducting their own studies. The European SIOPEN Group is launching this

multicentre observational approach in Europe, with a non-interventional strategy with three main

anticipated problems:

1. Differential diagnosis of suprarenal masses: Inclusion and exclusion criteria should be

clear, since in the majority of the cases we are going to observe masses that have not been

biopsied and we risk the fact of observing non-neuroblastoma cases “labelled” as

neuroblastoma.

2. What kind of diagnostic examinations are necessary in this population? Should a complete

staging be performed according to INSS in all cases in spite of invasive or long-sedation

examinations? Balance safety with efficiency/efficacy.

3. What kind of research tests can be performed to be “relatively” confident that patients with

MYCN amplified tumours are going to be excluded from this observational approach

without having biological tumour material to analyze?

14.1.6 Definitions

The following definitions will be used for the analysis:

Suprarenal Neuroblastoma: a suprarenal mass which shows MIBG uptake and/or elevated urinary

catecholamines

Supposed Adrenal Haemorrhage: a suprarenal mass which does not show MIBG uptake and/or

elevated urinary catecholamines, which subsequent to diagnosis has a typical evolution of a

haematoma on serial US examinations.



Other adrenal masses: any other mass which does not show MIBG uptake and/or elevated urinary

catecholamines, which may shrink, stay stable or enlarge during the observation period. No

definite diagnosis is feasible without pathology examination.

14.2 Hypothesis

Infants with suprarenal masses discovered antenatally or neonatally (suspected neuroblastoma)

can be managed non-operatively (without initial surgery) with close monitoring without

jeopardizing their outcome.

14.3 Type of study

A multi-centre, non-blinded, one armed prospective trial.

14.4 Aims of the study

14.4.1 Primary aim

To maintain a 3-year event free survival over 80% with a non-operative therapeutic approach

(serial monitoring, surgery if warranted) in infants with a localised suprarenal mass discovered

ante or neonatally.


To manage infants with suprarenal masses discovered ante or neonatally with a uniform

approach in Europe in a multicentre setting.

To maintain an excellent overall survival with a non-operative therapeutic approach (serial

monitoring, surgery if warranted) in infants with a local suprarenal mass discovered ante

or neonatally.

To determine the 3-year surgery-free survival in infants with suprarenal masses discovered

ante or neonatally and managed conservatively (non initial surgery).

To find out the natural history of perinatal suprarenal masses, according to the definitions

set up for the study.

To study the kinetics of regression in those suspected suprarenal neuroblastomas in infants

with suprarenal masses discovered ante or neonatally and managed conservatively (non

initial surgery).

1. To collect tissue from those suprarenal masses excised in order to perform standard and

investigational pathological and biological studies (INPC, MYCN, 1p, 11).

To collect frozen plasma from all patients included in the study in order to perform

research.

14.5 Eligibility

14.5.1 Inclusion criteria

Age less than or equal to 90 days when the suprarenal mass is discovered.

Suprarenal mass detected by ultrasound and/or MRI. The suprarenal mass may be cystic

and/or solid, but IT CANNOT REACH THE MIDLINE AND should MEASURE ≤ 5 CM

AT THE LARGEST DIAMETER.

No regional involvement: MRI scan does not show evidence of positive ipsi/contralateral

lymph nodes or other spread outside the suprarenal gland.

No metastatic involvement.

Frozen plasma available.

Informed consent.

Availability to do the adequate follow-up



14.5.2 Exclusion criteria

Age older than 90 days.

Suprarenal mass bigger than 5 cm.

Regional involvement.

Metastatic involvement.

Inability to undertake mandatory diagnostic studies (biological markers, US, MRI, MIBG).

Follow-up not guaranteed by parents/guardians.

14.6 Diagnostic procedures

A neonate in whom a suprarenal mass is visualized by ultrasound, either during pregnancy or

neonatally (≤ 90 days after birth) should be referred to an experienced centre in Paediatric

Oncology.

Date of diagnosis is defined as the date of the first ultrasound where the suprarenal mass

was visualised. If this is antenatally, take the date of birth. The first day of the

observational period would be the date of diagnosis.

Once the mass is discovered, the following studies must be carried out:

Imaging: The initial ultrasound has to be repeated at the referred centre and completed with

abdominal MRI (within the first 9 weeks) to rule out regional or metastatic involvement. Data

on size and characteristics of the mass will be collected. This data will be mandatory to

register the patient in the study.

Blood sampling: Blood counts, biochemistry including LDH will be performed

Urine sampling: Catecholamines [70-73] and free cortisol in urine will be performed.

MIBG: to be done within the first 9 weeks of age, in order to confirm the diagnosis of

neuroblastoma and rule out metastasis. It is mandatory in all those suprarenal masses with

increased catecholamines. In those lesions smaller than 1 cm with negative catecholamines it

will not be mandatory. If the time period differs to your National Guideline, then your

National Guideline should be followed.

Bone marrow aspirations and biopsies are not required, as they are invasive and difficult

techniques in neonates.

14.7 Therapeutic strategies

Observational approach:

Children, who fulfil the eligibility criteria and have completed the mandatory diagnostic

procedures and whose parents/legal guardians consent to participation in this study, will be

observed at the prescribed intervals during the course of the study.

- This observation will consist of physical examination, follow-up ultrasound and

urinary catecholamine tests every 3 weeks until week 12. (NOTE: if the week 0

catecholamines where negative, then repeat catecholamines are only mandatory if

an increase in the size of the mass is seen. For patients with positive week 0

catecholamines repeat tests every 3 weeks, although recommended, are not

mandatory. However, it is mandatory that at least 2 repeat samples are taken for

these patients within the first 12 weeks (week 6 and 12 are the preferred time

points).

- At this point, two features could be observed:

Initiation of regression: US and markers will be performed at week 18, 30 and 48.

If regression has not been observed, US and markers can be performed monthly until week

48.



NOTE: For regression of the tumour, no specific decrease in the size of the mass is

required. However, in the opinion of the treating physician the mass has to be

undoubtedly smaller.

- If there is evidence of an increase of tumour volume or an increase in catecholamine

metabolite excretion the children will go off the observational approach follow-up and will

switch to follow the off-observation recommendations.

- If at month 12 (48 weeks of observation) the suprarenal mass still persists, pathological

analysis is recommended.

- In the case where complete regression is observed (at or prior to week 48) the patient

should be followed annually for 3 years.

WEEK 48 = 1 YEAR OF OBSERVATION is the final point of the study. Then, MRI is

mandatory in both regressing and non-regressing masses.

follow-up

(weeks)

0 3 6 9 12 18 30 48

US + + + + + + + +

Catecholamines* + + + + + + + +

MRI** + +

MIBG **

* If the week 0 catecholamines where negative, then repeat catecholamines are only mandatory if

an increase in the size of the mass is seen. For patients with positive week 0 catecholamines repeat

tests every 3 weeks during the first 12 weeks, although recommended, are not mandatory.

However, it is mandatory that at least 2 repeat samples are taken for these patients within the first

12 weeks (week 6 and 12 are the preferred time points). After week 12 for patients with positive

week 0 catecholamines:

repeat tests at weeks 18, 30 and 48 are mandatory for patients where regression is

observed at week 12

repeat tests every 4 weeks, although recommended, are not mandatory for patients where

regression is not observed at week 12. However, it is mandatory to repeat tests every 8

weeks.

** It is recommended that MRI and MIBG will be done at a time-point between week 0 and 9

according to national policies. Reasons to perform both at the same time point are not only for

diagnosis but to rule out metastatic disease in neuroblastoma. Ensure that thyroid blockade is

undertaken with MIBG scan. NOTE: If the time period differs to your National Guideline, then

your National Guideline should be followed.

If regression has not been observed, US and catecholamines* can be performed monthly until week 48.

If regression has been observed, US and catecholamines* can be performed at week 18, 30 and 48.


Version 3.0; 23 May 2011 Page 107 of 237

US, LDH, Catecholamines and

free cortisone in urine and every 3

weeks until week 12

MRI within the first 9 weeks. If antenatally,

discuss with radiologist

MIBG (see text, page 105)

Diagnosis of neuroblastoma

based on MIBG +ve and/or high

catecholamines.

Regressing mass: US and markers

weeks 18, 30, 48.

Increasing size and/or increasing

metabolites: CONSIDER OFF STUDY. See text.

Non regressing mass: monthly US and markers until

week 48.

No diagnosis of NB; Low likehood of NB

due to MIBG -ve and/or negative catecholamines

Regressing mass: US and markers

weeks 18, 30, 42.

Non regressing mass: consider MRI and discuss surgery

past week 18.

Regressed mass No disappearance at week 48: MRI and

surgery

Regressing mass: at that moment, US

and markers every 2-3 months until

week 48.

Non regressing: MRI and surgery. Week

48

No NB: follow-up needed according to

etiology.

NB: staging according to

INRGSS. Treatment and follow-up accordingly.

Increasing size: OFF study: MRI and

surgery.

Follow-up week 48. Consider surgery if no disappearance.

MRI pre-op.


Version 5.0; 1 October 2015 Page 108 of 237

Children whose parents/legal guardians do not consent to inclusion into the observational

approach will be treated according to LNESG-2 algorithm.

Off -observation criteria and strategy:

There are clear and certain criteria to consider a surgical approach. Obviously, the first step is

a full reassessment to determine the appropriate surgical approach (INRGSS L1=surgical

excision, INRGSS L2= biopsy).

- A 40% increase of the original tumour volume associated or not with an increase

in catecholamines. NOTE: An increase in catecholamines alone is not criteria for

removal from study.

- If at the end of the observational period (1 year), a mass is persisting this will be

excised without regarding catecholamine metabolites.

- If the suprarenal mass disappears, but the catecholamine metabolites persist

elevated (evidence of neuroendocrine tumoural function), a diagnostic and staging

procedure should be performed according to LINES to rule out the presence of

occult disease.

- Suspicion of progressive disease (clinical symptoms/signs of progression or

metastases) will prompt the cessation of the observational approach and the

patient’s complete reassessment according to LINES.

In all other cases, closer monitoring by ultrasound and urine catecholamines is recommended,

based upon the individual criteria of the responsible centre, until stabilization is reached or

surgery is decided. At the point surgery is decided, the central database will be informed.

Surgery:

Adrenalectomy should be performed following standard procedures when the off-criteria

study is applied. Management of the surgical material must follow the E-SIOP Biology

(ENQUA) Guidelines in case a neuroblastoma is pathologically confirmed, biological studies

must be carried out.

14.8 Registration and study monitoring

All patients with a suprarenal mass will be registered in the database. Those that fulfil the

inclusion criteria will be the core of the study (observational approach patients) and the mandatory

data to summit to the central data base will include the ultrasound measures at inclusion and

catecholamines determination performed. Otherwise the patient will not be considered for final

analysis. The final analysis will divide patients in three categories:

Adrenal NB: a suprarenal mass which shows MIBG uptake and/or elevated urinary

catecholamines

Supposed Adrenal Haemorrhage: a suprarenal mass which does not show MIBG uptake and/or

elevated urinary catecholamines, with typical evolution by US.

Other adrenal masses: any other mass which does not show MIBG uptake and/or elevated

urinary catecholamines, and may shrink, stay stable or enlarge during the observation period.

No definite diagnosis is feasible without pathology examination.

The group of patients who are excluded from the study due to rejection of informed consent by

their parents or legal guardians and followed the standard LNESG-2 strategy (immediately

surgery) is of special interest for the Study Committee. It will be asked to those parents and the

LNESG-2 Committees to forward a copy of the pathological report and biology data on that

surgery for research purposes.



14.9 Definition of an event

For the subset of patients with suprarenal neuroblastoma (MIBG positivity and/or elevated

catecholamines) an event would be defined as having progressive disease (higher than L1) at the

time of resection; where progression is defined as ≥ 40% increase in tumour volume.

Progression/recurrence after resection will be defined as:

Any new mass lesion, in a patient under observation for a suprarenal mass, which is

subsequently biopsy proven neuroblastoma.

Any new mass lesion, either local or distant, in a patient who has undergone resection of a

suprarenal neuroblastoma, which is subsequently biopsy proven neuroblastoma.

MIBG positive for metastatic tumour.

Bone marrow positivity.

These events should be managed according to the current SIOPEN guidelines by including the

patients in the right study (LINES or HR-NBL-1), following staging procedures and biological

studies.

14.10 Statistical considerations

This study, as the majority of studies within this protocol, comprise a single arm strategy the

success or otherwise of which will be judged by reference to historical information (summarised

in the background). It is also expected that the numbers of patients within this subgroup will not

be sufficient to base sample size calculations on conventional methods of test size and power.

Hence the general objective is to obtain as many patients as possible within a reasonable time

frame (initially set at 5 years) and thereby to obtain the 95% confidence interval for the endpoint

measure and use this as the basis for conclusions.

The characteristics of the study are the following:

Single arm: Interventional. Serial monitoring based on imaging and laboratory studies in order to

avoid initial surgery in small suprarenal masses.

Number of patients anticipated in 5 years: 100. The number of patients anticipated to be recruited

in this observational approach over this period has been calculated using data from the INES

Study database.

Objectives: Maintain 3-year EFS of 80% with 3-year OS over 95%.




study.

For study aim 14.4.2 (b); the overall survival rate will be calculated.

For study aim 14.4.2 (c); the percentage of patients who are spared resection will be estimated.

The 3-year resection-free survival rate will be calculated.

For study aims 14.4.2 (c) and (d); descriptive statistics on demographic and biological data will be

calculated.

14.11 Collection of samples to study MYCN amplification in plasma [74-76]

Peripheral blood samples from neuroblastoma patients are collected on EDTA at diagnosis.

Plasma should be prepared by centrifugation of blood at 2000 rpm (700g) for 10 min.

The supernatant (plasma) is aliquoted, (Warning: When you perform aliquots, please avoid

touching the pellet).



Frozen quickly in liquid nitrogen and stored at -80°C. (This sending should be done in dry

ice by Express transporter).

To analyse the circulating MYCN DNA 200 to 300 µl of plasma are necessary.

Address of delivery

The samples should be sent to:

Valérie COMBARET

Laboratoire de Recherche Translationnelle

Batiment Cheney D

Centre Léon BERARD

28 rue Laennec

69373 Lyon cedex 08

FRANCE



APPENDIX 1. THE INTERNATIONAL NEUROBLASTOMA RISK GROUP

(INRG) STAGING SYSTEM

A1.1 Modified INRGSS as used in LINES

Stage Description

L1 Localised tumour not involving vital structures as defined by the

list of image-defined risk factors*

L2 Locoregional tumour with presence of one or more image

defined risk factors

M Distant metastatic disease (except stage Ms)

Ms Metastatic disease in children younger than 12 months** with

metastases confined to skin, liver, and/or bone marrow

* In this study “confined to one body compartment” is not considered. The reason for this is

detailed in section 4.3.2.1, page 27.

** In this study, we use the age cut off of ≤ 12 months. The reason for this is detailed in

section 4.3.2.2, page 27.

A1.2 Original INRGSS

Stage Description

L1 Localised tumour not involving vital structures as defined by the

list of image-defined risk factors and confined to one body

compartment

L2 Locoregional tumour with presence of one or more image

defined risk factors

M Distant metastatic disease (except stage Ms)

Ms Metastatic disease in children younger than 18 months with

metastases confined to skin, liver, and/or bone marrow

NOTE. See original paper for detailed criteria [7]. Patients with multifocal primary tumours

should be staged according to the greatest extent of disease as defined in the table.



APPENDIX 2. SUMMARY OF SIOPEN NEUROBLASTOMA TREATMENT

STRATEGIES

Disease stage

Age at

diagnosis

MYCN status Life

threatening

symptoms

Segmental

chromosome

alterations *

Study

Group

(LINES)

Study eligibility

Neonatal suprarenal

masses

<90 days

-

NSM**

L1 (excised)

Any

non Amp

LNESG 2

L2

≤18 months

non Amp

No

no 1

Low Risk yes 3

Yes

no 2

yes 3

Ms

≤12 months

non Amp

No

no 4

Low Risk yes 6

Yes

no 5

yes 6

L2

Differentiating

histology***

> 18 months

non Amp

7

Intermediate risk

L2

Poorly/undifferentiated

histology***

> 18 months

non Amp

8

Intermediate risk

L1 - INSS stage 1

Any age

Amp

9

Intermediate Risk

M

≤12 months

non Amp

10

Intermediate Risk

M

> 12 months

non Amp

High Risk

Ms

> 12 months

non Amp

High Risk

L1 INSS stage 2

L2,

M,

Ms

Any

Amp

High Risk

* If no genomic profile result is available, see Low Risk Study treatment strategy section 9.4.7 (page 72) for treatment

guidelines.

** See section 14.5 (page 104) for a detailed list of eligibility criteria.

*** If a clear definition of the histotype is not able to be classified (i.e. neuroblastoma NOS), see Intermediate Risk

Study treatment strategy section 10.4.5 (page 91) for treatment guidelines.



APPENDIX 3. LIFE THREATENING SYMPTOMS

The presence of any of these symptoms is an indication for chemotherapy.

Intraspinal neuroblastoma (See Appendix 13, page 155)

Patients who either have symptoms of spinal cord compression or have a spinal tumour

component that occupies more than one third of the spinal canal on the axial plane and/or the

perimedullary leptomeningeal spaces are not visible and/or the spinal cord signal is abnormal.

Systemic upset

Pain requiring opiate treatment

Gastrointestinal

- Vomiting needing nasogastric/IV support

- Weight loss >10% body weight

NOTE: diarrhoea with VIP does not respond to chemotherapy and is a definite indication

for surgery

Respiratory

- Respiratory distress without evidence of infection

Tachypnoea >60

Oxygen need

Ventilatory support

Cardiovascular System

- Hypertension

- IVC compression +/- leg oedema

Renal

- Impaired renal function, creatinine increased x 2 ULN1

- Poor urine output, less than 2mls/kg/day

- Hydroureter/hydronephrosis

Hepatic

- Abnormal liver function >2 ULN

- Evidence of DIC

- Platelets <50 x 109/l

Bladder/Bowel dysfunction secondary to a mass effect.

A very larger tumour volume causing concern of possible tumour rupture and/or the

possible rapid development of systemic upset.

NOTE: Some of these symptoms will require immediate treatment with chemotherapy. In these

cases the definitive biopsy to obtain material for the genomic profile should be delayed until the

patient is fit enough to have a biopsy which is likely to be either within 7 days of the

chemotherapy administration or just before the second course of chemotherapy is given.

If tumour material for the genomic profile is obtained after the initiation of chemotherapy, the

genomic profile will be considered non informative (no genomic profile result) and the patient

will not be eligible for a genomic profile result treatment stratification.

1 ULN = Upper Limit Normal



APPENDIX 4. LIST OF IMAGE DEFINED SURGICAL RISK FACTORS [7]

The Image Defined Risk Factors (IDRF) have been designed to guide surgical management of

neuroblastoma at diagnosis, in particular to indicate whether biopsy or attempted resection is

recommended as the first surgical procedure.

The same system of risk factors can be applied later during the course of treatment and follow-up,

although it is not designed for this purpose. If a tumour remains IDRF positive after

chemotherapy in specific clinical situations as in Study Groups 3, 6, 7, and 8 this may not be an

absolute contraindication to surgery. Resection may still be recommended if evaluation of the

primary tumour suggests that the risk to life, or of major functional loss from surgery, is less

than the risk from leaving residual disease. In these situations discuss with the National Surgical

Coordinator.

Ipsilateral tumour extension within two body compartments:

Neck-chest, chest-abdomen, abdomen-pelvis

Neck

Tumour encasing carotid and/or vertebral artery and/or internal jugular vein

Tumour extending to base of skull

Tumour compressing the trachea

Cervico-thoracic junction

Tumour encasing brachial plexus roots

Tumour encasing subclavian vessels and/or vertebral and /or carotid artery

Tumour compressing the trachea

Thorax

Tumour encasing the aorta and/or major branches

Tumour compressing the trachea and/or principal bronchi

Lower mediastinal tumour, infiltrating the costo-vertebral junction between T9 and T12

Thoraco-abdominal

Tumour encasing the aorta and /or vena cava

Abdomen/pelvis

Tumour infiltrating the porta hepatis and/or the hepatoduodenal ligament

Tumour encasing branches of the superior mesenteric artery at the mesenteric root

Tumour encasing the origin of the celiac axis, and/or of the superior mesenteric artery

Tumour invading one or both renal pedicles

Tumour encasing the aorta and/or vena cava

Tumour encasing the iliac vessels

Pelvic tumour crossing the sciatic notch

Intraspinal tumour extension whatever the location provided that: More than 1/3 of the spinal canal in the axial plane is invaded

and/or the perimedullary leptomeningeal spaces are not visible

and/or the spinal cord signal is abnormal

Infiltration of adjacent organs, structures

Pericardium, diaphragm, kidney, liver, duodenopancreatic block and mesentery.

Conditions recommended to be recorded, but NOT considered IDRFs:

Multifocal primary tumours

Pleural effusion (with or without malignant cells)

Ascites (with or without malignant cells)



APPENDIX 5. RESPONSE EVALUATION

This study will use volume measurements for the tumour response assessment. Tumours do not

necessarily grow or shrink in a rounded fashion and 3D evaluation is likely to be more accurate

than uni or bidimensional measurement. The size of the tumour should be calculated using

volume calculation. This maybe obtained from the commonly used elliptical approximation (D1 x

D2 x D3 x 0.52) or by common software algorithms which are implemented in most radiological

workstations. For more information on tumour measurement see section A8.5, page 128.

In this study the following definitions will be used for response evaluation criteria:

Complete Response (CR): Complete disappearance of all measureable disease.

Stable Disease (SD): No tumour shrinkage to demonstrate any objective reduction in

tumour volume and not a sufficient increase in tumour volume to

qualify for PD.

Progressive Disease (PD): An increase in tumour volume of >40% or the appearance of a

new site of disease. If 3D measurements are not obtainable then

PD is defined as an increase of 25% in the product of 2

dimensions [77].

The definition of partial response (PR) is not used in this protocol for determining treatment.

However, PR will be used to assess the disease status of patients for data collection.

Partial response is defined as any shrinkage of the primary tumour and/or a decrease in size or

number of metastatic sites.



APPENDIX 6. PATHOLOGY GUIDELINES

The “local pathologists” should be informed by the “local” paediatric oncologists of the possible

inclusion of a patient in the LINES and that they have to send slides and pathological reports

within 2 weeks of finalisation of their histopathological report to the national coordinating

pathologist (see A6.3.3).

The material obtained, in the majority of cases by needle biopsies or surgical open biopsies, and in

the minority of cases by surgical resection (complete, with minimal residual disease or

incomplete) must be sent fresh without any preservatives, fixatives or formalin and as quickly as

possible to the pathologist. The division of tumour material is performed by the pathologist with

adequate frozen tissue banking for MYCN FISH and molecular studies (see A7.3, page 121) on

one hand, and formalin fixation for morphological assessment on the other.

A6.1 Needle biopsies

When a needle biopsy is indicated instead of surgical procedure, the “general procedure for

handling the material” is the same as in the surgical procedures (see A6.2).

To date, there is no data to indicate the minimal volume of tumour material required to permit

application of the INPC classification. Empiric data consider that only 2cmx2cm or greater

surgical biopsies allow INPC characterisation. For these reasons, sufficient tissue must be

obtained without taking any risk of adverse effect or complication for the patient, ideally from two

different areas of the tumour, and/or with several needle core biopsies to provide sufficient tissue

for diagnostic studies. Note that the amount of tissue for biological studies will be depend on

overall amount sent to pathologist who will have to consider the pathological features more

critically than just confirming NB.

This implies that in case of needle biopsies the INPC histoprognostication

(Favourable/Unfavourable), and INPC morphological characteristics independent of the age (i.e.

differentiation and MKI) developed in the INRG risk grouping scheme will be determined on a

prospective basis, with mention of the size of the formalin fixed tumour material submitted for

histological evaluation. The differentiation and MKI will be determined in the stroma poor

component of the tumour. These criteria should be assessed only if a minimum of 5000 tumour

cells are present in the studied material.

In the “Pathology Form”, it will be specified that in the case of a stroma rich tumour, it is not

possible to exclude a nodular component in another area of the same tumour, requiring a

correlation with clinical and radiological data.

In case of multiple core biopsies, if one or more cores are occupied by a stroma rich tumour and

other core(s) by a stroma poor tumour, a diagnosis of “ganglioneuroblastoma stroma mixed

nodular” could be proposed.

The accurate volume of frozen material for biological studies (MYCN determination, molecular

evaluation) is determined with the biologist. The non frozen material must be formalin fixed.

A6.2 Surgical open biopsies and surgical resections

Including: complete, with minimal residual disease or incomplete.

In all cases where surgical resection is performed before chemotherapy, the guidelines for

pathology process are the same as those in the LNESG-2 protocol, which is available on the

SIOPEN-R-NET website.



In these samples the tissue volume is sufficient to apply the morphological criteria of the INPC

[46]. It will therefore be possible to assign the tumour to one of the four PNTs categories [47],

i.e.: neuroblastoma stroma poor, ganglioneuroblastoma stroma mixed nodular,

ganglioneuroblastoma stroma rich intermixed, ganglioneuroma, and to determine histoprognostic

categories: Favourable or Unfavourable. Moreover, the differentiation (i.e.: undifferentiated,

poorly differentiated and differentiating) and the MKI (low <2%, intermediate between 2 and 4%,

and high >4%) will be determined in the neuroblastoma stroma poor component of the tumour for

each individual tumour as developed in the INRG pre-treatment risk grouping scheme.

These data will be quoted in the “Pathology Form”.

In case of fine needle aspiration, or in case of core biopsy with less than 5000 tumour cells in the

studied material, the INPC is not applicable.

A6.3 Review process

Please also refer to the Pathology Form (see A6.4, page 119) and the Local Pathologists

Agreement Form (see A6.5, page 120).

A6.3.1 Material to be sent

One HE-stained section from each paraffin block of the tumour must be sent to one of the national

coordinating pathologist from the ESIOP pathologist panel (Gabriele Amann, Klaus Beiske,

Catherine Cullinane, Emanuele S.G. d’Amore, Claudio Gambini, Samuel Navarro, Michel

Peuchmaur).

Only in the case of an undifferentiated tumour: 1 HE per block and either 1 paraffin block or 5 or

10 unstained sections (for immunohistochemical studies).

In order to ease their review task, clinical information on catecholamines, site of tumour, MIBG

data, or other relevant data should be sent to the national coordinating pathologist together with

the slide(s).

The tumour cell content of the frozen specimen dedicated to biological studies will be analysed by

the local pathologist.

A6.3.2 Timeline

The “local pathologists” must send slides and pathological reports within 2 weeks of finalisation

of their histopathological report to the national coordinating pathologist. The national

coordinating pathologist sends his/her report within 4 weeks to: local pathologist, local paediatric

oncologist, and national clinical coordinator.

Finally, review in real time means that it takes 3 to 6 weeks from biopsy to completion of the

national review.

A6.3.3 Pathology reviewers

Names, address and function of local and national clinical coordinators of each participating

country should be clearly established and accessible to the national coordinating pathologist of the

ESIOP pathologist panel.

We, the pathologists of the ESIOP pathologist panel, agree to review cases from our own

countries and from those countries which are not represented in the review panel. The pathologists

of the latter countries should be informed by the national clinical coordinator and the LINES

clinical coordinator.



ESIOP pathologist panel (see A22.7, page 219 for contact details):

K Beiske: Scandinavian countries + Belgium + Greece

G Amann: Austria + Czech Republic + Hungary

ES d'Amore and C Gambini: Italy + Serbia + Slovakia

C Cullinane: UK + Ireland + Baltic countries + Australia

S Navarro: Spain + Poland + Portugal

M Peuchmaur: France + Israel + Switzerland

A6.3.4 Logistical issues

To overcome comprehension difficulties with different languages, the international form -

including the International Neuroblastoma Pathology Classification (INPC) and the grading of

differentiation of the tumours should be used by the local pathologists (see A6.4, page 119).

If after national review the grading (characterisation of the differentiation of the tumour) cannot

be established, the national coordinating pathologist will send the case to one of the other

pathologists of the review panel in order to try to obtain a consensus on the grading of such

tumours. This final consensus could be either:

grading not done. Peripheral Neuroblastic tumours NOS, or

after discussion the grading could be proposed

The delay from biopsy to the consensus and sending the answer must be less than 12 weeks.

We, the pathologists of the ESIOP pathologist panel will organise 2 international meetings per

year for the review and discussion of every case over a period of 2 working days per meeting.

Such international review will be performed as a quality control system.



A6.4 Pathology form

PATHOLOGICAL ASSESSMENT

Biopsy or Surgery date: birth date: D D M M Y Y D D M M Y Y

Date of the pathological diagnosis D D M M Y Y 1/ Specimen •1 needle biopsy (<5000 tumour cells) •2 needle biopsy (>5000 tumour cells) •3 open surgical biopsy •4 surgical resection

2/ Diagnosis according to INPC:

In case of core biopsy and stroma rich tumour, it is not possible to eliminate a nodular component in another area of the same tumour, requiring a correlation with clinical and radiological data.

In case of multiple core biopsies, if some cores are occupied by a stroma rich tumour and other core(s) by a stroma poor tumour, a diagnosis of “ganglioneuroblastoma stroma mixed nodular” will be proposed.

In case of Fine needle aspiration, or in case of core biopsy with less than 5000 tumour cells in the studied material, INPC is not applicable.

Neuroblastoma Ganglioneuroblastoma Ganglioneuroma Other •1 NOS •5 NOS •10 Maturing GN •12 NB tumour not classifiable •2 Undifferentiated •6 Intermixed •11 Mature GN •13 Small round cell tumours, NOS •3 poorly differentiated •7 Nodular, undifferentiated* •14 Only necrotic material •4 differentiating •8 Nodular, poorly differentiated* •15 Non NB tumour

•9 Nodular, differentiating* MKI •1 Low •2 Intermediate •3 High •4 Not evaluable Calcifications •1 Absent •2 Present •3 Not evaluable

(*: for ganglioneuroblastoma nodular, the degree of differentiation, the MKI and the calcification are assessed in the nodule, or in the nodule with the worst prognostic criteria if several nodules) 3/ INPC Prognostic category: (1) favourable, (2) unfavourable, (3) not evaluable •

4/ Miscellaneous: Nucleoli in undifferentiated or poorly differentiated neuroblastoma tumour cells: (1) small, (2) large • Nuclear morphology: (1) conventional, (2) polymorph •

5/ Tissue specimen designated for biology:

% of tumour cells1) % of Schwannian cells1)

% of other cells (e.g. lymphocytes, fibroblasts)

Necrosis •1 No •2 Yes If yes, % of necrosis2) •3 NE 1) estimated % of all nucleated cells 2) estimated % of total area

Please forward 1 HE-stained section from each paraffin block, within 2 weeks of your diagnostic

report, to one of the members of the European Neuroblastoma Reference Panel (without sending

your report!)

Pathologist’s Signature:

Date:



A6.5 Local pathologist agreement form

AGREEMENT FORM FOR THE PRIMARY (LOCAL) PATHOLOGIST: LINES Protocol

As the pathologist who is responsible for the primary handling of neuroblastic tumours in my

department, I am aware that the final conclusion about histoprognosis in neuroblastic tumours:

must be based on the assessment of at least two pathologists from different institutions

and should be reached within 4 weeks of finalisation of local histopathological report.

Therefore, for all patients who, according to the information provided by the paediatric

oncologist(s) at my institution, will be enrolled into the LINES protocol, I agree to:

send 1 HE-stained section from each paraffin block of the tumour to one of the reference

pathologists in the SIOP European Neuroblastoma Pathology Review Panel (Gabriele

Amann, Klaus Beiske, Catherine Cullinane, Emanuele S.G. d’Amore, Claudio Gambini,

Sam Navarro, Michel Peuchmaur) within 2 weeks of finalisation of my histopathological

report,

discuss by phone/mail/other the diagnosis and prognostic classification according to INPC

with the reference pathologist in order to obtain a final conclusion which is agreed upon

by both.

Name of the responsible pathologist: ……………………………………………

Name and address of institution: ……………………………………………

……………………………………………

……………………………………………

………………….. ……………………………………………..

Date Signature



APPENDIX 7. BIOLOGY GUIDELINES

A7.1 Mandatory investigations

Mandatory investigations within this protocol are:

Tumour cell content of the tumour sample

DNA index

MYCN copy number

Genomic profile of cases without MYCN amplification using a technique for pangenomic

analysis (MLPA or array-CGH).

In addition, it is highly recommended to store RNA for expression studies.

For patients included in the Neonatal Suprarenal Mass study:

Serum for the determination of MYCN status based on circulating DNA should be sent to

Valérie Combaret.

Handling of biological material according to the pathology guidelines and adequate material for

biological studies is mandatory! All mandatory investigations have to be carried out in the

National Reference Laboratories or other Reference Centres, i.e. laboratories participating in the

European Neuroblastoma Quality Control Assessment (ENQUA) study. Results from other

laboratories will not be taken into account.

If the DNA index can not be done at a local or national level, a representative paraffin block

should be sent to Klaus Beiske or Rosa Noguera for determination of ploidy (static method).

A7.2 Schedule for results

MYCN results have to reach the data centre within 10 days.

DNA index results and tumour cell content information have to be available within 3 weeks and

sent to the laboratory in order to help the MLPA/array-CGH/SNP interpretation.

For patients for whom these results are decisive for treatment stratification, i.e. patients included

in the Low risk section of the protocol, the genomic profile results will have to reach the data

centre and be returned to the clinician within 4 weeks, including central quality review.

For all other patients (i.e. patients for whom the pangenomic results will not be decisional) these

results have to reach the data centre within 8 weeks.

A7.3 Tumour material for biological studies of neuroblastoma

Surgical biopsies or fragments

In this case, provide, if possible, a fresh tumour sample in culture medium and a frozen tumour

sample in a cryotube. In case the tumour fragment is small, it should be frozen only.

Needle-core biopsies

A minimum of 2 biopsies (ideally 3) snap-frozen in separate microtubes is necessary for

biological analyses (plus one for Pathology).

Fine-needle aspirates frozen in a cryotube.



A7.4 Tumour handling

Tumour handling should be done according to the guidelines developed by the SIOPEN Biology

(ENQUA) Group [44, 78].

Fresh tumour sample to be used for:

o Pathology

o DNA index (flow cytometry or static photometry)

o MYCN status and other FISH studies

Frozen tumour tissue to be used for:

o Frozen sections: tumour cell content

MYCN status (FISH)

Other FISH studies

o DNA extraction: Pangenomic/mulitlocus analysis (MLPA/array-CGH/SNP)

Plasma to be used for:

o MYCN status (for patients of Neonatal protocol, without availability of tumour tissue)

Protocol to collect plasma samples:

Blood samples from neuroblastoma patients are collected at diagnosis in EDTA tubes. They are

centrifuged at 2000 rpm (700g) for 10 minutes. The supernatant (plasma) is aliquoted, frozen

quickly in liquid nitrogen and stored at -80°C.

To analyse the circulating MYCN DNA 200 to 300 µl of plasma are necessary.

Remark: Don’t touch the pellet during the collect of plasma to prevent the contamination by the

blood cells.

A7.5 Recommendations for determination of MYCN status

Interphase FISH (I-FISH) as technique to determine MYCN status is preferred because it has a

number of advantages compared to other techniques, most important of which is direct quality

control of the specimen by the microscopist. Use of a reference probe on the same chromosome is

required. For recording the MYCN-FISH results the terms summarised in Ambros et al. [43]

should be used. Heterogeneous MYCN amplification requires meticulous analysis of the tumour

specimen. At the present time, the prognostic significance of heterogeneously MYCN amplified

neuroblastomas is not known and the prognostic impact of 2p gain is still under investigation.

A7.6 Recommendations for MLPA

Extraction of tumour DNA from a sample (frozen tumour sample or tumour cells) containing at

least 60% of tumour cells, according to standard procedures.

Currently, the INRG Biology Subcommittee suggests using multilocus or pangenomic techniques,

enabling an analysis of all relevant genomic loci in a single experiment. Besides the pangenomic

array-CGH technique, multigenomic techniques like multiplex ligation-dependent probe

amplification (MLPA) will likely be the major technique for detecting structural and segmental

chromosomal alterations in the future. MLPA is a recent PCR based technique that detects a large

variety of segmental alterations and gene amplifications in a robust manner. At least two probes

are used for every target sequence. The probes are hybridized to the tumour sample DNA, ligated,

and then amplified. The amplification of the probes is dependent on the presence of the target

sequence in the tumour DNA, and quantification of the losses, gains and amplifications in small

amounts of DNA can be determined by analyzing the ratio of the reaction product to the number

of target sequences/alleles. Thus, simultaneous investigation of a large number of loci, covering



all currently known important aberrant regions found in neuroblastomas, can be performed in a

single assay. The robust nature of the results and the relatively low cost of the MLPA kits make

this technique attractive for routine NB analysis. The technique is validated, and a consensus for

platform(s), procedures, terminology, categorization and analysis has been reached (Ambros et al.

CCR in submission).

The MLPA technique enables the simultaneous investigation of a large number of loci of interest

(i.e. 1p, 1q, 2p: MYCN, DDX1, NAG, ALK, 3p, 4p, 7p, 9p, 11q, 12p, 14q, 17q). Both, structural

alterations, such as amplifications, segmental chromosome alterations (gains and losses) as well as

numerical chromosome changes are detectable.

A7.7 Recommendations for array-CGH

Extraction of tumour DNA from a sample (frozen tumour sample or tumour cells) containing at

least 60% of tumour cells, according to standard procedures.

Slides: Commercialised slides or slides prepared by a National Reference Laboratory can be used.

They should contain DNA sequences (BAC/PAC clones, oligonucleotide sequences, SNP)

distributed evenly along the whole genome. Chromosomal regions of special interest in

neuroblastoma, such as chromosomes 1p, 11q or 17q, may be covered more densely. Duplicate or

triplicate presence of DNA sequences used on the slides allow for calculation of a mean for a

given locus, rendering results more robust.

Tumour DNA and an equal amount of normal reference DNA quality and quantity checked,

digested, purified and labelled with distinct fluorochromes. Following precipitation and

denaturing, the samples are hybridised to the slides. The slides are then washed and ready for

acquisition of images. Images will be acquired using an appropriate software [4].

A7.8 Recommendations for SNP-array

SNP (single nucleotide polymorphism) arrays can be used as an alternative technique for whole-

genome screening of aberrations in neuroblastoma tumours. When using conventional CGH

arrays the comparison/normalisation is performed directly on the array glass with tumour and

normal DNA labelled with different fluorochromes. In contrast, with SNP arrays the comparison

tumour vs. normal is performed in the computer with profiles from normal controls analysed

earlier. The SNP arrays have the advantage that allelic/haplotype information can be included in

the analysis in that for instance imbalances and copy-neutral LOH can be readily analysed.

Several commercially available SNP arrays are available. They are generally very robust and

uniform and give excellent genome profiling results. They also come in different densities; Arrays

with some 250, 000 SNPs evenly spaced in the genome have been available for some years, and

more recently arrays with combinations of some 1 million SNPs and 1 million copy number

probes have been released.

Molecular biology-based methods, including MLPA, array-CGH and SNP array, are sensitive to

contamination by DNA of normal cells. To use these techniques, a sample with a tumour cell

percentage exceeding approximately 60% is required to obtain good results. In tumour samples

containing less than 60% of neuroblasts (samples from primary tumours with extensive necrotic,

haemorrhagic and/or calcification areas or metastatic sites), I-FISH might potentially provide

useful information about the presence of a segmental chromosomal alteration corresponding to the

studied chromosome locus if such an alteration is clearly present in at least 60% of the observed

tumour cells. In these cases at least 3 images showing nuclei with decisional segmental

chromosome alteration results will be uploaded into the SIOPEN-R-NET database immediately

for central review.



If results obtained by one pangenomic/multilocus technique are uninterpretable, or if no

alterations are detected, the sample is to be analysed by another pangenomic/multilocus technique

and/or DNA will be sent to another SIOPEN reference laboratory for reanalysis. Alternatively, I-

FISH could be performed, but only clear-cut results will be accepted. Also consider new DNA

extraction when sufficient material with adequate tumour cell content is available.

If necessary, for instance in case of difficulties for interpretation of the result, DNA should be sent

immediately to another laboratory, as well as a frozen section/H+E slide to check the indicated

tumour cell content.

Genomic profiles without any significant genomic changes (neither Numerical Chromosome

Alterations nor Segmetnal Chromosome Alterations) will not be taken into account for treatment

decisions.

In case of tumours with different components (for instance a tumour with different sections, only

one of which shows segmental chromosome aberrations): if one sample gives interpretable CGH-

or SNP-array or MLPA results, this will be sufficient for interpretation. That is, if one component

of the tumour, containing >60% tumour cells, shows segmental chromosomal aberrations these

should be taken into account for making the clinical decision (even if the other components do not

have these alterations).

A7.9 Central review

All decisional genetic results including multilocus or pangenomic profiles will be uploaded into

the SIOPEN-R-NET database immediately for central review in real time.

Clinical decision making based on genomic results means that the result should be validated by a

second independent person. This is easier for MLPA/array-CGH-/SNP results as uploaded

images will show the whole profile, whereas for FISH only a limited number of images can be

reviewed. A central review system (on a rotational basis) will be organised within the SIOPEN

Biology Group.

The final clinically relevant conclusion of the genomic profile, confirmed following central

review by the SIOPEN Biology Group, will be returned to the clinician within 4 weeks of the

biopsy.

A7.10 Genomic classification

Based on data obtained by MLPA/array-CGH/SNP tumours will be classified into genomic

groups. Within this protocol the following terms are used:

Definition of genomic profiles: Chromosomal alterations detected by MLPA/array-CGH /SNP

NCA GENOMIC PROFILE - presence of numerical chromosomal alterations (NCA) only

SCA GENOMIC PROFILE - presence of any segmental chromosomal alteration (SCA)

observed recurrently in neuroblastoma (deletion of chromosome 1p, 3p, 4p, or 11q; gain of 1q,

2p, or 17q) without or with numerical chromosomal alterations

NO RESULT

No genomic result: pangenomic/multilocus analysis not done because of: no adequate

tumour cell content, no DNA, no tumour, etc

Genomic result not evaluable: pangenomic/multilocus analysis done but not informative because

of: a flat curve, too much fluctuation, too high standard deviations, or too many outliers.



Genomic result not prognostically informative: pangenomic/multilocus analysis done and

interpretable, but the detected segmental alterations concern only chromosomes from which

prognostic information is not available so far.

If the tumour sample is obtained after the beginning of chemotherapy, the genomic profile

result is not to be taken into account and should be considered as “no result”.

The genomic profile, obtained by MLPA/array-CGH/SNP, as well as the single genetic alterations

will be recorded and entered in the SIOPEN-R-NET site.



APPENDIX 8. IMAGING GUIDELINES

A8.1 Introduction

The aim of these guidelines is to define standard procedures that will enable accurate evaluation

of the resectability of a localised NB by detection of possible Imaging Defined Risk Factors

(IDRFs) and to assess response to treatment.

A8.2 Methods of investigation

For the most frequent abdominal and pelvic locations, ultrasonography (US) is usually the first

modality used at diagnosis in children because of wide availability, non-invasiveness and absence

of ionizing radiation. US allows accurate analysis of tumour relationships with adjacent organs

and vessels with the use of Doppler imaging. The liver is also well assessed with sonography in

children. In newborns, US may also be used to depict intraspinal involvement. However, US is

associated with important limitations: a low inter-observer reproducibility, a limitation in

assessing highly calcified tumours because of acoustic shadowing and a high limitation in

retrospective review of the data which is mandatory for clinical trials. Therefore, CT-scan or MRI

should always be performed at diagnosis for accurate staging.

CT was initially recognised as a useful technique for the diagnosis and staging of NBs [79]. CT-

scan is widely available in imaging departments and multi-detector row CT (MDCT) machines

now allow fast acquisitions without motion artefacts, reducing the need for sedation in most

situations. Intraspinal extensions are also well depicted and post-treatment software allows

multiplanar reconstructions (MPR). However, CT scan requires intravenous iodine contrast agent

injection for both increasing the soft tissue contrast and for assessing relationships between the

tumour and adjacent vessels. Moreover, CT-scan is associated with significant radiation exposure

and children are known to have a higher inherent sensitivity to ionizing radiations [80].

MRI was secondarily recognised to be the most useful imaging method. MRI should be preferred

because of a higher contrast resolution compared to CT and the lack of ionizing radiation [81, 82].

There is no consensus in the radiologic literature regarding the need for gadolinium contrast

injection to assess NB extensions. T1- and T2 weighted images (WI) actually provide excellent

tissue contrasts and adequate display of vessels can be achieved without the use of contrast [83].

The main limitations of MRI are the local availability in some countries and, compared to CT, the

need for sedation in young children because of longer acquisition time of sequences. MRI is the

recommended modality for investigating intraspinal extension of primary paraspinal tumours [84]

because of the excellent visualisation of the spinal cord, nerve roots and subarachnoid spaces.

A8.3 CT-scan technical recommendations

A8.3.1 Preparation

Performing CT in children requires adapted procedures for immobilisation and warming for

newborns and infants, and analgesic procedures for intravenous line placement.

The use of MDCT currently limits the need for sedation in most situations, provided adequate

immobilisation of young children. There is no consensus about the use of oral contrast agent or

water to delineate the digestive tract. Enema is usually not required, even for pelvic locations.

A8.3.2 CT protocol

Postero-anterior ± lateral projection radiographs are mandatory to adjust the study volume.

Unenhanced series allow depicting intratumoural calcifications but should be used only in

doubtful cases and limited to the tumour volume.

One contrast-enhanced series is usually sufficient to assess the tumour extent.

Well tolerated contrast agents (low osmolarity, non-ionic) should be used with 300 to 350 mg/L

iodine concentration with a total volume of 1.5 to 2 ml/kg. The perfusion rate should be adapted



to patient age and to the IV line diameter, usually between 0.8 and 2 ml/sec to ensure appropriate

enhancement. The scan delay should be adjusted in order to get both arterial and venous

enhancement, and depends on the age and the anatomic region assessed and to the acquisition

time. CT acquisition is usually begun just after the end of contrast injection for neck and chest,

and about 35-40 seconds after the beginning of injection for abdomen and pelvis to get venous

and portal phase. Injection may be performed manually or with an automatic injection device. An

automatic bolus detection option can be used to adjust the scan delay.

The beam collimation should be adjusted to the CT machine (number of detector rows). The

nominal slice thickness should be 1 to 2.5 mm and the reconstruction thickness between 2.5 and 5

mm. The use of a minimal tube rotation time is recommended to reduce the acquisition time.

Recommended pitch values are between 1 and 1.5 providing a reasonable compromise between a

short acquisition time and sufficient z-resolution.

Tube voltage and mAs should be adapted to patient age, weight and to the anatomic region

studied. The tube voltage is usually set between 80 and 100 kVp for the trunk in newborns and

infants, and between 100 or 120 kVp in older children.

The mAs should be adjusted to obtain a final mean absorbed dose (i.e., CTDI volume) in

agreement with the current paediatric recommendations or dose reference levels [85, 86].

A lower mediastinal tumour, infiltrating the costo-vertebral junction between T9 and T12 is a

theoretical risk of medullar ischaemia because of Adamkiewicz anterior spinal artery injury

during tumour resection [87]. MDCT angiography has been described as a possible alternate to

selective angiography [88], and could be used in this indication. Actually, angiography is

currently not mandatory because of the intrinsic risk of angiography in young children and since

this risk is considered very low in children.

A8.4 MRI

A8.4.1 Preparation, sedation and monitoring

The image quality in young children is highly sensitive to motion artefacts. MRI in children

younger than 4 or 5 years should therefore be associated to either sedation or general anaesthesia

with adapted monitoring. Various sedation protocols may be used depending on local strategies.

Auditory protection is mandatory, especially in infants and newborns.

MRI settings should be adjusted to keep the Specific Absorption Rate (SAR) below 1.5 W/kg.

A8.4.2 MRI protocol

MRI scanning always involves customising scan settings to the individual patient and to the local

MRI unit.

There is currently no consensus about the optimal MRI protocol to be used in NB patients.

Appropriate compromise should be obtained between field of view, matrix, slice thickness, signal

to noise ratio and acquisition time, the latest being optimised by phased array coils and parallel

acquisition. Each protocol should be adapted to the available options, notably motion-reducing

sequences and respiratory or cardiac gating.

Field of view and matrix should be adjusted to patient size and anatomic regions. Pixel size

should range between 0.5 to 1 mm and slice thickness between 3 to 6 mm for 2D imaging (1 to 2

mm for 3D gradient echo) in order to optimise the spatial resolution.

Acquisition planes should always include the axial transverse plane and at least one longitudinal

plane (coronal and/or sagittal). Both sagittal and coronal planes are mandatory for paraspinal NBs

to assess foraminal and intraspinal extensions. The orientation of the coronal views should be

adjusted on the sagittal view to provide images demonstrating all foraminal and intraspinal

tumour extent on the same coronal image.

Appropriate sequences are T1-WI (spin echo, fast spin echo or spoiled-gradient echo) and T2-WI

(fast spin echo) or STIR (short-tau inversion recovery). The tissue contrast on T2-WI can be

increased with the use of fat saturation options.



The role of whole-body MRI (WBMRI) in bone marrow assessment for neuroblastoma staging is

also still not defined. Small series provided encouraging results, but further studies should be

obtained. MRI is actually known to be a highly sensitive technique to depict bone marrow

location, however its specificity is low compared to MIBG. However, MRI cannot be used to

assess tumour response as signal abnormalities may persist for months despite good response [89].

Diffusion-weighted MRI could also be helpful for tumour delineation [90] but further studies are

required to assess its contribution. The use of gadolinium-enhanced T1-WI sequences is optional

but, if used, should be performed with fat saturation to increase the contrast against fatty tissues.

Gadolinium should be avoided in children with renal function impairment because of the risk of

secondary nephrogenic systemic fibrosis [91].

A8.5 Tumour measurements

Three-dimensional measurement of the primary tumour should always be provided [7] at the time

of diagnosis, during therapy or observation period, and - in case of residual tumour – during

follow-up.

Measurements are easily obtained for well circumscribed tumours: the largest diameter and the

largest orthogonal diameter are obtained from the transverse plane, and the height of the tumour

from one longitudinal (coronal or sagittal) plane obtained either directly with MRI or with MPR

with CT-scan. Volume calculation may be obtained from the commonly used elliptical

approximation, i.e., D1 x D2 x D3 x 0.52. Current European recommendations are that 3D

measurements are used for the definition of response. The implications of which are that there are

changed cut-off values [77, 92]. NOTE: If 3D measurements are not obtainable then the product

of 2 dimensions should be calculated.

The table below shows the relationship between change in diameter, product and volume* [77].

Diameter, 2r Product, (2r)2 Volume, 4/3пr3

Response Decrease Decrease Decrease

30% 50% 65%

50% 75% 87%

Disease Progression Increase Increase Increase

12% 25% 40%

20% 44% 73%

25% 56% 95%

30% 69% 120%

* Shaded areas represent the response evaluation criteria in solid tumours (diameter) and World

Health Organization (product) criteria for change in tumour size to meet response and disease

progression definitions.

The response evaluation definitions used in this study can be found in Appendix 5 (page 115).

Measurements are more difficult and less reproducible for complex-shaped masses or for tumours

associated to numerous adjacent lymph nodes. In such situations, it is recommended to measure

the whole tumour mass (i.e., primary and adjacent lymph nodes in a single volume). Common

software algorithms which are implemented in most radiological workstations can be also used.

It may be difficult to accurately assess the volume of an intraspinal tumour extension due to the

half moon configuration it commonly takes in that particular site. It is suggested to consider the

ratio between the maximum axial diameter of the tumour within the spinal canal and the diameter

of the spinal canal. In the case where more than 1/3 (33%) of the spinal canal is invaded by the

tumour, an IDRF is present. The craniocaudal extent of the intraspinal component is measured in

relation to the vertebrae involved. The leptomeningeal spaces must be assessed to determine if

they are visible and the spinal cord MRI signal needs to be evaluated.



A8.6 Relationship between the primary tumour and neighbouring vital structures

It is recommended that these relationships between the primary tumour and neighbouring vital

structures should be systematically discussed in a multidisciplinary team including (at least)

radiologists, nuclear physicians, surgeons and paediatric oncologists.

To ensure uniform staging the following terms should be used by radiologists for description of

the relationship between a tumour and the neighbouring vital structures, i.e. structures that can not

be sacrificed without impairment of normal function such as major arteries, veins, and lymphatic

vessels or major nerves. Other vital structures are the heart, diaphragm, lungs and airways, liver

and biliary system, spleen, pancreas, kidney and the urinary tract, bowel and mesentery.

A8.6.1 “Separation”

“Separation” means that a visible layer, usually fat, is present between the tumour and the

neighbouring structure. When a tumour is separated from a vital structure, according to previous

definitions, an IDRF is not present.

A8.6.2 “Contact” and related terms

“In contact” means that no visible layer is present between the tumour and the neighbouring

structure. For vessels, a “contact” means that less than 50% of its circumference is in contact with

the tumour (more than 50% being named “encasement”). The term “compression” is additionally

used for veins with reduced diameter but still partially visible lumen. The term “compression” is

also used for spinal cord and airways when a tumour is in contact and causes the short axis of the

structure to be reduced. For other vital structures (neighbouring organs), a “contact” may be

associated with “displacement” (abnormal anatomic location) or “distortion” (abnormal anatomic

shape) of the structure, but neither with “total encasement” nor “infiltration”. When a tumour is

“in contact” with a vital structure, according to previous definitions, an IDRF is not present.

However, “compression” of the trachea or the spinal cord is considered as an IDRF.

A8.6.3 “Encasement” and related terms

“Encasement” means that a neighbouring structure is surrounded by the tumour. A “total

encasement” means that a vital structure (organ, vessel) is completely surrounded by the tumour.

“Encasement” of a vessel means that more than 50% of its circumference is in contact with the

tumour. The additional term “stretched” means that a partially or totally encased or displaced

vessel is elongated by an expanding tumour, i.e. an artery having a reduced diameter or a

compressed vein with no visible lumen. When a tumour is encasing a vital structure, according to

previous definitions, an IDRF is present.

A8.6.4 “Infiltration”

“Infiltration” means involvement of vital structures other than vessels (infiltration of the vessel

wall can currently not be demonstrated by imaging): an infiltrating tumour has extension into a

neighbouring organ causing the margins between the tumour and the infiltrated structure to be

lacking or ill-defined.

When a tumour infiltrates a vital structure, according to previous definitions, an IDRF is present.

A8.6.5 Compartments, uni- and multifocality

The majority of tumours occur in a single anatomic compartment. However, some tumours may

extend into an adjacent compartment and increase the risk of injury of vital structures during

surgery; these tumours should be classified L2 according to the INRGSS. This usually occurs with

cervico-thoracic locations arising from the stellate sympathetic ganglion, or with lower

mediastinal locations extending into the retroperitoneum along the aorta, or in the pelvic presacral

locations extending upward in the abdomen around the aorta and/or IVC.



An upper abdominal tumour with enlarged lower mediastinal nodes or a pelvic tumour with

inguinal lymph node involvement is considered locoregional disease. However, nonregional

(distant) lymph node involvement, such as retroperitoneal primary with supraclavicular lymph

node, is metastatic disease (stage M).

Multifocal primary neuroblastoma is a rare situation. These tumours should be staged according to

the greatest extent of disease. This special feature should be recorded but will not be considered,

by itself, as an IDRF.

A8.6.6 Effusions

Isolated pleural effusion and ascites, even with malignant cells, are not considered IDRFs in the

INRGSS. Although pleural disease is associated with reduced survival rates in metastatic patients,

isolated pleural effusion or ascites is rare in patients with locoregional disease, and its impact on

outcome is still unclear.

A8.7 IDRF assessment according to anatomic location

A8.7.1 Neck

Cervical NBs arise from the superior cervical sympathetic ganglion located behind the internal

carotid artery or, more rarely, from the stellate ganglion. These tumours usually extend anteriorly

and laterally displacing the carotid artery and internal jugular vein, medially compressing the

airways and upward to the skull base along the carotid artery.

According to the INRGSS, surgical risk factors should be considered when the tumour encases the

carotid and/or vertebral artery and/or internal jugular vein, extends to the base of skull, and/or

compresses the trachea.

A rarer NB involving the cervico-thoracic junction and arising from the stellate ganglion may

encase the brachial plexus roots and/or subclavian vessels and/or vertebral artery. Intraspinal

extension may be also observed. All these conditions are considered IDRFs.

A8.7.2 Thorax

Thoracic NBs mostly arise from the paraspinal sympathetic trunks in the posterior mediastinum.

Foraminal and intraspinal extensions (dumbbell tumours) are typically observed at this anatomical

level. Spinal cord compression may be diagnosed on either neurological signs and/or on the basis

of imaging findings. According to the INRGSS, an IDRF is present when more than one third of

the spinal canal in the axial plane is invaded and/or the perimedullary leptomeningeal spaces are

not visible and/or the spinal cord MRI signal is abnormal, i.e. imaging features usually observed

in symptomatic patients.

A lower mediastinal tumour, infiltrating the costo-vertebral junction between T9 and T12 is still

considered as an IDRF as there is a theoretical risk of medullar ischaemia by Adamkiewicz

anterior spinal artery injury [87]. This artery originates in 85% of the cases between T9 and T12

on the left side, whereas in 15% of the cases it originates on the right side and is usually

associated with a supplementary circulatory circuit. Tumour growth within this area induces the

formation of collaterals. Therefore, arteriography of the medullary artery is not mandatory, since

the preoperative risk is minimal, although it should be mentioned to the parents. On the other

hand, an invasive procedure such as arteriography in very young patients is risky.

Tumours encasing the aorta and/or major branches and tumours compressing the trachea and/or

principal bronchi usually cause major surgical difficulties: therefore these conditions are

considered IDRFs.



A8.7.3 Abdomen

Abdominal NBs arise from either the adrenal gland and/or from sympathetic ganglions (celiac,

superior and inferior mesenteric ganglions) and/or sympathetic fibres and plexuses located along

the aorta and its main branches.

Relationships between the primary tumour and abdominal vessels usually represent the main

limitation for surgery. Therefore, accurate analysis of all arteries and veins is a major issue in

imaging reports. Main abdominal vessels (aorta, celiac axis, superior and inferior mesenteric

arteries, renal arteries and veins, inferior vena cava, iliac arteries and veins, porta hepatis) should

be clearly assessed and classified according to the previously defined terms. According to the

INRGSS, tumours encasing the superior mesenteric artery, the origin of the celiac axis, one or

both renal pedicles, the aorta, the inferior vena cava and/or the iliac vessels are considered IDRFs.

Adjacent organs and structures should also be precisely assessed and classified according to the

previously defined terms.

According to the INRGSS, the following are considered IDRFs: tumours infiltrating the porta

hepatis or the hepatoduodenal ligament, diaphragm, kidney, liver, duodeno-pancreatic block,

and/or the mesentery.

Paraspinal lumbar NBs rarely may be associated with foraminal and intraspinal extensions

(dumbbell tumours) sharing the same pattern and neurological issues as mediastinal primaries.

A8.7.4 Pelvis

Pelvic NBs mainly arise from either the upper hypogastric sympathetic plexus and/or presacral

sympathetic ganglions.

Pelvic NBs are frequently associated with foraminal/intraspinal extensions and associated with

sacral and/or inferior lumbar radicular neurological signs, but not with spinal cord compression.

According to the INRGSS, an IDRF is present when more than one third of the spinal canal in the

axial plane is invaded. However, intraspinal tumour extension below the level of L2 does not lead

to spinal cord compression but only to radicular involvement. This condition is usually not a

contraindication for primary surgery of the extraspinal component and usually does not lead to

emergency neurosurgery.

Relationships between mass and majors pelvic vessels (aorta, inferior vena cava, primitive,

internal and external iliac arteries and veins) should be clearly assessed and classified according to

the previously defined terms. According to the INRGSS, tumour encasement of the aorta and/or

inferior vena cava and/or iliac vessels is associated with IDRF.

The involvement of the sciatic notch is considered an IDRF. The “sciatic notch” in the IDRF

definition is a sagittal-oblique plane at the anatomic level of the greater sciatic notch, joining the

spine of the ischium and the lateral border of the sacrum.



APPENDIX 9. GUIDELINE FOR RADIOIODINATED MIBG

SCINTIGRAPHY IN CHILDREN

A9.1 Purpose

The purpose of this guideline is to offer to the Nuclear Medicine team a useful framework in daily

practice.

The present document is influenced by the conclusions of the reunion on "Consensus Guidelines

for MIBG Scintigraphy" (Paris, November 6, 1997) of the European Neuroblastoma Group and by

those of the Oncologic Committee of the French Society of Nuclear Medicine [93]. This guideline

summarises the views of the Paediatric Committee of the European Association of Nuclear

Medicine. It should be taken in the context of “good practice” of nuclear medicine and local

regulation.

A9.2 Background information and definitions

Meta-iodobenzylguanidine (MIBG) is an aralkylguanidine noradrenalin analogue which, once

iodinated with 131I or 123I, enables successful imaging of neuroectodermally derived tumours,

including neuroblastoma and phaeochromocytomas [94-98] although other tumours, such as

paragangliomas, medullary thyroid carcinomas, carcinoid tumours, Merkel cell tumours of the

skin, and metastases of these tumours, have been shown to take up MIBG [99, 100].

Neuroblastoma is one of the most common solid malignant tumours of childhood. As the stage of

the disease has an impact on both prognosis and treatment, and as 80% of neuroblastoma are

metastatic (stage IV) at the time of presentation, the accurate identification of all lesions is crucial

to evaluate the extent of disease [101, 102]. The specificity of MIBG for detecting primary and

secondary neuroblastoma approaches 100%. The sensitivity in detecting sites on a lesion-by-

lesion basis is about 80% and the sensitivity in terms of staging is 90-95% [103-108].

The high tumour affinity also allows the therapeutic use of the 131I labelled compound [109-111].

MIBG therapy shares with MIBG imaging several points but this guideline only covers its use in

diagnosis and follow-up. This guideline contains information related to the indications for,

acquisition, processing, and interpretation of radioiodinated MIBG scintigraphy in

neuroectodermally tumours in order to obtain scans with high and constant quality.

There was controversy regarding the respective value of MIBG and bone scan in neuroblastoma

patients. It is 2 now clear that neither MIBG scintigraphy nor MDP bone scans alone detects all

the lesions. In some cases bone scan detects lesions that are MIBG negative and in others MIBG

uptake is positive while the bone scan is normal [112]. Thus, initial assessment should include

both MDP bone scan and MIBG scintigraphy. MIBG assesses response to treatment of primary

tumour and metastatic sites. Follow-up evaluation using MDP is limited since one infrequently

discovers new abnormal MDP uptake without corresponding MIBG uptake. Moreover, these

kinds of abnormalities are highly unspecific and may correspond to any lesion not related to

neuroblastoma.

A9.3 Common indications

A9.3.1 Indications

A. Confirmation in suspected neuroectodermally derived tumours including neuroblastoma,

phaeochromocytoma and ganglioneuroma.

B. Staging of the disease.

C. Follow-up of neuroblastoma under chemotherapy, particularly in stage IV and IVs

patients.

D. Before and after surgery of the primary tumour.



E. Follow-up after treatment to exclude a sub-clinical relapse, especially in the bone marrow

and also in the case of any clinical abnormality during follow-up, particularly bone pain.

F. Before planning MIBG therapy

A9.3.2 Contra indications

There are no contra indications.

A9.4 Procedure

Referring physicians should be informed that radioiodinated MIBG is a cyclotron product and

therefore not continuously available at all times. Several days are required to obtain the MIBG

also there is a period (days) between injection and scan depending on which iodine nuclide is

used.

A9.4.1 Information about previous examinations relevant to this procedure

The clinical information related to the patient, the reason for performing the examination, the

biochemical data, the previous treatment (i.e. surgery, chemo- and/or radiotherapy) and diagnostic

procedures should be available.

A9.4.2 Patient preparation

A9.4.2.1 Information with appointment letter:

Child and parents should receive detailed information about the entire procedure. The thyroid

should be blocked (see A9.4.2.2). Ensure that the actions (see A9.4.3) prior to the injection of

tracer have been carefully explained to the family or to ward staff for in-patients. This will require

a special information sheet. Good hydration is advocated to lower the radiation burden.

Prior to Injection: On arrival in the department a local anaesthetic cream can be used; it should

be applied at least 60 minutes before the injection.3

A9.4.2.2 Thyroid blockade

Thyroid blockade is important to protect the thyroid from unnecessary irradiation, an organ that is

more radiosensitive in children than in adults.

Beginning on the day before tracer injections until the day after injection, children from one

month to three years should receive 32 mg potassium iodide daily, from three to thirteen years 65

mg, and over this age 130 mg daily. New-borns receive 16 mg potassium iodide only on the day

before tracer injection.

Rapid blockade by perchlorate (Irenat) is an alternative option.

A9.4.2.3 Drugs interactions

Many classes of medicines interfere with MIBG uptake and storage [113]. Treatment prescribed

before and at the time of MIBG injection and imaging procedure should always be checked with

the referring physician.

Unlike in adult patients, the list of usually concerned substances is actually limited.

The most common are bronchodilators containing: Fenoterol (Berotec ®), Salbutamol

(Ventolin®), Terbutaline (Bricanyl®) and nasal drops and sprays containing Xylometazoline

(Otrivine drops®).

The possibility of over the counter medicines should also be kept in mind.

Many cardiac drugs interfere with MIBG and although such drugs are rarely found in a paediatric

population, special attention should be paid to cases of suspected pheochromocytoma. Those

usually prescribed in children are:

Calcium channel blockers: Nifedipine (Adalat®), Nicardipine (Cardene®), Amlodipine

(Norvasc®),



Angiotensin-Converting Enzyme inhibitors: Captopril (Capoten®), Enalapril (Vasotec®),

Adrenergic Receptor Blockers: Labetalol (Trandate®), Amiodarone (Cordarone®),

Inhibition of sodium pump: Digoxin (Lanoxin®).

A9.4.3 Precautions

Check if thyroid blockade has been given.

Check on going treatment for possible drugs interactions.

The tracer must be injected slowly, preferably over a period up to 5 minutes.

A9.4.4 Radiopharmaceutical

A9.4.4.1 Radionuclide

Iodo-123 (123I)

Iodo-131 (131I)

A9.4.4.2 Pharmaceutical

MIBG (Meta-iodobenzylguanidine)

123I-MIBG is the radiopharmaceutical of choice in children giving high quality images. Gamma

emission energy of 159 keV for 123I is more suitable for imaging than 360 keV for 131I and the

differences in terms of radiation burden permit to inject higher activities with 123I compared to

131I. Result of 123I-MIBG scintigraphy is also more rapidly available for the clinician.

131I-MIBG that is more widely available is an acceptable second choice and may be preferred

when 4 estimation of the tumour's MIBG-retention is required for therapy planning.

A9.4.4.3 Dose Schedule (see A9.5)

Minimum doses: 123I-MIBG = 80 MBq

131I -MIBG = 35 MBq

Recommended maximum doses: 123I-MIBG = 400 MBq

131I -MIBG = 80 MBq

Administered doses should be scaled down to body weight [114].

A9.4.4.4 Injection technique

Slow injection (at least 5 min) in peripheral vein, flushed with saline. Potential adverse effects of

MIBG (vomiting, tachycardia, pallor, abdominal pain) that are not related to allergy are very rare

in case of slow injection. Rapid injection is contraindicated as it causes the above adverse effects.

Injection via central venous catheters should be avoided if possible and in that case injection

should be even slower.

A9.4.4.5 Radiation Burden

Depending on administered activity and on the child's age. i.e.: 3,7mSv for a 5 year old child

receiving

5.18MBq/kg 123I-MIBG [115] and 5.5mSv for a for a 5 year old child receiving 0.74 MBq/kg

131I-MIBG [115].

A9.4.5 Image Acquisition

An adapted environment, an adequate attitude toward the child, a well-trained technologist for

paediatric procedures and involved parents before and during the procedure all help in obtaining a

co-operative child.

Sedation is usually not required for a technical satisfactory examination. The most difficult age is

between 1 and 3 years. In this category, sedation might be necessary [116].



A9.4.5.1 Timing for imaging

Using 123I -MIBG images are acquired 20-24 hours after the injection.

Selected delayed images - never later than 48h - may be useful in the odd cases with equivocal

findings.

Using 131I-MIBG, scanning is generally performed 48h after the injection and can be repeated at

day 3 or later.

A9.4.5.2 Collimator

Use the collimator suitable respectively for 123I and 131I energy depending on manufacturer’s

recommendations.

A9.4.5.3 Positioning of the child

The highest quality images are obtained by having the child as close as possible to the camera

face, if possible on the camera face. A special table - if available - with an aperture for the

collimator will allow imaging of the patient lying directly on the collimator.

A9.4.5.4 Views

Whole body scan imaging with additional spot images including lateral views of the skull can be

considered especially in children able to lie still for the entire acquisition time. However spot

images of the entire body are the reference method.

Skull anterior, posterior

Skull right, left (including arms)

Chest anterior, posterior

Abdomen anterior, posterior

Pelvis (empty bladder) anterior, posterior (lateral if bladder is not empty)

Lower limbs anterior, posterior

These views should be similar to those obtained with bone scans i.e. the toes turned inwards and

the knees together. The ankles should be included.

A9.4.5.5 Computer acquisition set up

Static images: 250.000 counts or 10 min counting per image are necessary (compromise between

best image quality and limitation of scanning time). A pixel size of approximately 2 mm requires

a 256x256 matrix or 128x128 matrix with zoom.

Whole body scanning: a scan speed of 5 cm/min or a total acquisition time of 30 minutes is

appropriate.

A9.4.5.6 Optional images

Landmarking: In certain circumstances, it can be of help to use other tracers in conjunction with

MIBG. In case of difficulties to differentiate between MIBG tumour uptake and retention of

activity in the renal pelvis one may either use furosemide to washout the pelvic activity or use

MAG3/DTPA to identify the kidney. A bladder catheter is not necessary except in particular

cases. However, even in pelvic neuroblastoma, the bladder activity is rarely a problem. The child

(if co-operative) should be encouraged to void prior to the imaging.

SPECT imaging: In cases where uncertainty exists as to the exact site of MIBG activity SPECT

may be useful. The abdomen is the area where this is most likely to occur: lesions in or close to

the liver, as well as close to the bladder or any other area of intense physiological uptake are

particularly good indications to add SPECT.

The feasibility of SPECT will depend on the child and on the equipment available (multiple head

camera) [117-119].



A9.4.6 Intervention

Not relevant.

A9.4.7 Processing

Not relevant.

A9.4.8 Hard copy output

Grey scale of all images that have been acquired.

A9.4.9 Interpretation, reporting and pitfalls

A9.4.9.1 Normal biodistribution

Knowledge of normal biodistribution is necessary to avoid misinterpretations [120-122]. MIBG is

normally taken 6 up by liver, spleen, myocardium, salivary glands and normal adrenals. The

myocardial uptake may be particularly high, especially in children under 1 year. In the other

cases, there is a balance between liver and myocardial uptake. Skeletal muscles, nasal mucosa,

lungs, urinary tract, colon, gallbladder and thyroid may also demonstrate accumulation of tracer of

variable intensity. Uptake of MIBG by the various organ systems reflects either rich adrenergic

innervation or catecholamine excretion (or both) [96, 122].

Free iodine causes thyroid uptake (which can be blocked with stable iodine or perchlorate (see

A9.4.2.2) and some digestive artefacts [120].

No skeletal uptake should be seen. Extremities show only slight muscular activity and in these

cases the bone may appear as a photon deficient area (as observed at the level of the knees).

A9.4.9.2 Pathology

MIBG soft tissue uptake is observed in the primary tumour and in metastatic sites including

lymph nodes, liver, bone and bone marrow.

Intensity of MIBG uptake can be similar in benign and malignant tumours. Similarly, maturation

of neuroblastoma may occur with no change in intensity of uptake.

MIBG skeletal uptake can be observed either as focal areas of increased uptake or as a diffuse

skeletal uptake. The increase of tracer uptake is related either to bone metastases or to bone

marrow infiltration or both.

A9.4.9.3 False negative scans

One or several lesions could be missed due to a variety of reasons:

physical reasons related to limited spatial resolution which will prevent detection of small

lesions,

anatomical reasons, with lesions located close to a voluminous primary or metastatic mass, or

close to

soft tissue with a high physiological uptake (myocardium, thyroid and salivary glands, liver,

kidney,

bladder and colon) [106, 120],

no tumour uptake or low tumour uptake related to physiopathological reasons arising from

tumour

heterogeneity, ischaemic necrosis in the tumour mass, lack of granules, loss of tumour

capacity to take up the tracer, pharmaceutical inhibition or unknown reason [103].

A9.4.10 Quality control

Due to the long acquisition time, it is important to check for movement in each image recorded.

This has to be done before the child leaves the department.



A9.5 Issues requiring further clarification

The amount of activity to be injected, reported in D.3 is higher than that proposed by EANM

paediatrics task group [114]. There is actually no scientific basis to suggest that there is a

“correct” amount of radiotracer to inject.

Accurate staging is a priority and experience gained from MIBG therapy has taught us that

sensitivity of detection with MIBG increases with increased activity injected.

Acknowledgement:

The authors thank Dr CA Hoefnagel, Dr J Lumbroso and Dr S Meller for their valuable advice.

Guidelines issued date: December 29, 2002



APPENDIX 10. SURGICAL GUIDELINES

A10.1 Introduction

These guidelines are intended for the surgical management of all patients with neuroblastoma who

are not classified as “High Risk” patients using the International Neuroblastoma Risk Group

(INRG).

The role of the surgeon in the management of the primary tumour in Very Low, Low and

Intermediate Risk neuroblastoma has been re-evaluated following previous studies [6, 123]. These

have highlighted the increased risk of complications associated with attempted resection in the

presence of Surgical Risk Factors (SRF), in contrast to excellent results following complete

resection of localised tumours with no SRF.

Since many of these patients have an excellent overall prognosis even when complete resection of

their tumour is not possible, it is important to avoid unnecessary surgical morbidity.

SRF are now termed Image Defined Risk Factors (IDRF) and are based on cross sectional

imaging before any surgery. IDRF are intended to ensure consistent interpretation of the pre-

treatment extent of disease, but they also guide surgeons in the pre operative identification of

children of all ages in whom neuroblastoma resection is likely to be hazardous and or incomplete.

The Image Defined Risk Factors (IDRF) are listed in Appendix 4, page 114. Data from previous

studies have shown a discrepancy between the interpretation of imaging by surgeons, and by

radiologists. It is therefore important that the imaging be assessed against the list of IDRF by a

radiologist with appropriate paediatric expertise. The INRG stage can then be determined by a

multidisciplinary team (MDT), which should include the surgeon.

A10.2 Aims of surgery

To achieve complete excision of localised neuroblastoma with minimal morbidity.

To provide sufficient tissue for histological and biological studies by biopsy in those children

where resection would be hazardous.

To remove residual tumour after chemotherapy or natural involution only in those situations

where the benefit is likely to outweigh the risk

A10.3 Surgical procedures at presentation

At presentation this group of tumours fall into one of three broad categories, which guide the

initial surgical approach.

Tumours suitable for resection at presentation:

- L1 by INRG definition (localised tumour: IDRF negative)

Tumours suitable for biopsy only at presentation:

- L2 by INRG definition (localised tumour: IDRF positive)

- M and MS tumours by INRG definition (metastatic disease)*

Tumours suitable for observation only (no resection or biopsy)

- Adrenal mass in selected infants under 90 days of age at presentation

*Excision of the primary tumour may be an alternative diagnostic procedure to biopsy in

metastatic tumours, provided the primary tumour is IDRF negative.



Since pathological diagnosis, MYCN status and other biological parameters remain central to the

management of neuroblastoma, biopsy or resection at presentation is mandatory in all children,

except those less than 90 days at presentation with an adrenal mass less than 5 cm in diameter.

A10.4 Subsequent/secondary surgical procedures

Following chemotherapy or natural involution the criteria for attempting delayed resection are

influenced by the age of the child, and also sometimes by the IDRF.

The IDRF have been designed to guide surgical management of neuroblastoma at diagnosis, in

particular to indicate whether biopsy or attempted resection is recommended as the first surgical

procedure.

The same system of risk factors can be applied later during the course of treatment and follow-up,

although it is not designed for this purpose. If a tumour remains IDRF positive after

chemotherapy this is not an absolute contraindication to surgery. Resection may still be

recommended if evaluation of the primary tumour suggests that the risk to life, or of major


A10.4.1 Excision of primary mass should be attempted for:

Persistent suprarenal mass in infants under 90 days at presentation, following period of

observation, provided the tumour is IDRF negative (Section 14.7, page 105).

Enlarging suprarenal mass in an infant under 90 days of age at presentation, according to the

study guidelines, provided the tumour is IDRF negative (Section 14.7, page 105).

L2 tumours in children less than 18 months at presentation, where the IDRF have become

negative following chemotherapy or natural involution. Study groups: 1 (page 47), 2 (page 54)

and 3 (page 58).

Ms tumours in children less than 12 months at presentation with segmental chromosomal

alteration, provided the tumour is IDRF negative. Study group 6 (page 69).

Metastatic (M) tumour in children under 12 months at presentation (bone, lung, and/or CNS

metastases), with no evidence of residual metastatic disease (liver excluded), irrespective of

IDRF status. Study group 10 (page 88).

L2 tumours in children over 18 months of age at presentation with a differentiating tumour

which has become IDRF negative following chemotherapy. Study group 7 (page 78).

L2 tumours in children over 18 months of age at presentation with undifferentiated or poorly

differentiated tumours, irrespective of IDRF status. Study group: 8 (page 81).

A10.4.2 Excision of primary mass should be discussed with the surgical coordinators for:

Persistent or enlarging suprarenal mass in infants under 90 days at presentation, which is

IDRF positive at the time when the guidelines suggest that resection should be carried out.

L2 tumours in children less than 18 months at time of presentation, with segmental alterations,

where the IDRF remain positive after chemotherapy. Study group 3 (page 58).

Ms tumours in children less than 12 months at time of presentation, with segmental

alterations, where the IDRF remain positive after chemotherapy. Study group 6 (page 69).

L2 tumours in children over 18 months at presentation with a differentiating tumour which

remain IDRF positive. Study group 7 (page 78).

Tumours which remain IDRF positive, and are causing persistent symptoms at the end of the

chemotherapy schedule.

Discussion is essential in these cases, because the advantage of excision depends upon careful

assessment of risks vs. benefits.



A10.4.3 Excision of primary mass is not recommended for:

L2 tumours in children less than 18 months at the time of presentation, where the IDRF

remain positive following chemotherapy or natural involution, unless there are segmental

alterations. Study groups 1 (page 47) and 2 (page 54).

Metastatic (Ms) tumours in children under 12 months at presentation, with no segmental

chromosomal alterations irrespective of IDRF status. Study groups 4 (page 62) and 5 (page

65).

M tumours in children less than 12 months at presentation, who still have metastatic disease at

sites other than the liver at the end of the chemotherapy schedule. Study group 10 (page 88).

A10.5 Timing of subsequent procedures (where applicable)

A10.5.1 Neonatal suprarenal masses (NSM): an observational approach

Enlarging tumour (see guidelines, page 108) – excise (or biopsy) immediately.

Persisting tumour – excise at 1 year of age, providing IDRF negative.

A10.5.2 Low risk study

Children receiving chemotherapy:

Excision of localised tumour is recommended as soon as the tumour becomes IDRF

negative, in order to keep the chemotherapy load to a minimum.

Children who have received no treatment (“observation only”):

Excision is recommended 1 year from diagnosis, provided that IDRF are negative.

A10.5.3 Intermediate risk study

The timing of surgery is influenced by the INRG risk grouping, so refer to the individual

treatment schedule.

A10.6 Definitions

A10.6.1 Complete Excision

- Complete excision is defined as the removal of all visible tumour, including the removal

of abnormal lymph nodes and the sampling of normal lymph nodes.

- It is important to assess the likelihood of microscopic residual tumour even if macroscopic

complete tumour resection has been achieved. This can be aided by pathological

examination of biopsies taken from the tumour bed as well as examination of the tumour

margins.

- However, microscopic residual tumour only demonstrated by pathological examination

should not influence the surgeon’s assessment of completeness of excision.

A10.6.2 Excision with minimal residual disease (<5% of original or <5ml volume)

- Macroscopic residual disease remains after operation.

- The amount should be estimated by the surgeon in millilitres and as a percentage of the

original tumour volume.

A10.6.3 Incomplete gross excision

- More than 5% or 5ml of tumour remain after attempted excision.

- The residual should be evaluated by postoperative imaging.

A10.6.4 Biopsy

- Any procedure which leaves more than 50% of the tumour is defined as a biopsy.

- Biopsy should be the first procedure in all tumours that have Image Defined Risk Factors.



A10.7 Surgical Procedures

A10.7.1 Biopsy (please refer also to the pathology guidelines)

- Sufficient tissue must be obtained, ideally from two different areas of the tumour, to allow

histological diagnosis and biological studies. In particular, it is essential that sufficient

material be obtained for the accurate determination of MYCN status and other biological data.

- Open biopsies are the most satisfactory but many may feel this is an unacceptable additional

procedure given that the diagnosis can be made by less invasive means. Multiple needle core

biopsies can provide sufficient tissue for diagnostic studies. Optimum treatment is critically

dependent on correct tissue handling. It is mandatory that the pathologist is prepared to

receive the samples. Fresh tissue should be delivered to the pathologist immediately under

sterile conditions. Formalin or other preservatives must NOT be used.

- Fine needle aspiration is acceptable if there is no other alternative and is an effective source

for additional tumour cells for biological studies.

A10.7.2 Surgical approach for tumour resection

A10.7.2.1 Abdominal tumours

For abdominal lesions a transverse laparotomy incision is recommended.

If necessary this can be extended to a thoracoabdominal approach

A10.7.2.2 Pelvic tumours

For a pelvic lesions a midline incision may be preferable to Pfannensteil

A combined laparotomy and posterior sagittal approach may be required for some low lesions.

A10.7.2.3 Thoracic/cervical tumours

Large thoracic lesions may require a double thoracotomy at a distance of 2-3 intercostal

spaces, using the same soft tissue incision or a thoracoabdominal approach if just above the

diaphragm.

For lesions in the dome of the thorax, and thoracic inlet, alternative approaches may be

considered:

- A ’trap-door’ incision including neck, clavicle and sternum [124].

- A trans-manubrial approach provides similar surgical access to the ‘trap-door’ incision,

with less disruption of bones and muscle [125].

In some instances, a combined surgical approach is required which may need more than one

operating session. Examples include:

approach through cervicotomy and thoracotomy in cervico-mediastinal or upper-mediastinal

NB

bilateral thoracotomy in some cases of mediastinal NB extending beyond the midline (bilateral

thoracotomies can be carried out synchronously or with an interval between the procedures)

laparotomy combined with a sacrococcygeal/posterior sagittal approach

laminotomy associated with thoracic or abdominal operation.

Resection of large paravertebral tumours may result in vertebral instability. In these situations

consider advice from spine/orthopaedic surgeons.

A laparoscopic approach is acceptable provided oncological principles are strictly adhered to.



A10.7.3 Lymph Node Evaluation

To accurately INSS stage a localised tumour it is necessary to ascertain involvement of non-

adherent lymph nodes. Therefore, if possible, non-adherent lymph nodes should be sampled

during complete resections, even if the lymph nodes appear macroscopically normal.

Depending on the site of the primary tumour, lymph nodes from the following regions should

be examined and removed if they appear abnormal:

Lateral cervical region: jugular chain and supraclavicular area;

Chest: mediastinal lymph nodes above and below the tumour;

Abdomen: coeliac nodes (infra-diaphragmatic), mid-aortic (at renal level) and iliac region

(bilaterally).

A10.7.4 Intraspinal Extension

In patients with no IDRF the extraspinal mass should be removed even though intraspinal disease

remains.

Macroscopic disease may be left in the intervertebral foramina, especially when there is a risk of

leakage of spinal fluid and/or jeopardising the blood supply of the spinal cord.

A10.7.5 Intraspinal disease with neurological symptoms.

The urgent need in this situation is relief of pressure on the spinal cord rather than excision of the

primary. Operation is not usually the best strategy. For more information see A14.3, page 157.

A10.7.6 Nephrectomy

Nephrectomy is not acceptable as part of immediate excision.

A10.7.7 Tumour Relation with Great Vessels.

In order to gain further information on the accuracy of the pre-operative imaging, the intra-

operative findings should be described in detail. Particular attention should be given to the

technical difficulties encountered when the tumour is in contact with the vessels.

A10.7.8 Risk factors encountered at operation

If risk factors are encountered unexpectedly during operation, serious consideration should be

given to abandoning the procedure.

A10.7.9 Clips

Titanium or absorbable clips should be used if necessary to avoid interference with subsequent

imaging.

A10.8 Definition of Major Surgical Complications

Death within 30 days of operation, or obviously related to the operation at any time.

Serious haemorrhage >30% blood volume.

Serious vascular injury leading to loss of tissue

Any spinal cord injury.

Serious peripheral nerve injury leading to loss of function.

Any organ failure.



APPENDIX 11. CHEMOTHERAPY GUIDELINES

If chemotherapy is assigned to be given to eligible patients, three different regimens might be

applicable: CO, VP/Carbo, or CADO. The chemotherapy regimens applicable in this protocol are

described below.

It is very important to note that according to patient weight, the final drug dose might be

calculated based on:

Body surface area (BSA) for patient weighing > 10 kg or

Weight for patients weighing ≤ 10 kg

For infants weighing below 5 kg, chemotherapy drug doses should be reduced by a further 33%.

Chemotherapy courses should be given at the indicated intervals (see below) provided the

absolute neutrophil count (ANC) is >1.0 x109/l and the platelet count (PLT) is >100 x 109/l.

If the count has not recovered from the previous course of chemotherapy, treatment should be

delayed for a week, and the count checked again.

A11.1 CO (Cyclophosphamide and vincristine)

Courses of CO are given at 14 day intervals

DAY 1 2 3 4 5

Cyclophosphamide • • • • •

Vincristine •

DRUG Dose (mg/kg) (mg/m2) Administration Cyclophosphamide 5.0 mg/kg 150 mg/m2 Slow bolus injection daily x 5 (or PO)

Vincristine 0.05 mg/kg 1.5 mg/m2 Slow bolus injection day 1 only

Cyclophosphamide is given on days 1-5 of each course and vincristine is given on day 1 only.

A central line is not mandatory for chemotherapy administration. In those infants where

treatment is given via a peripheral line, great care should be taken to avoid extravasation.

Additional hydration is not required in the absence of vomiting, as long as oral intake is

satisfactory. If this is not the case, then intravenous fluids at "maintenance" rates (depending on

the age and weight of the child) should be given for the duration of chemotherapy.

A11.2 VP/Carbo (Etoposide {VP16} and carboplatin)

Courses of VP/Carbo are given at 21 day intervals

DAY 1 2 3

Carboplatin • • •

Etoposide • • •

DRUG Dose (mg/kg) Dose (mg/m2) Administration Carboplatin 6.6 mg/kg 200 mg/m2 5% dextrose (5 ml/kg) over 1 hr

daily x 3

Etoposide (VP16) 5.0 mg/kg 150 mg/m2 0.9% saline (12.5 ml/kg) over 2 hrs

daily x 3



Both these drugs are given on days 1-3 of each course.

It is advisable that a central line is inserted for chemotherapy.

In those patients where treatment is given via a peripheral line, great care should be taken to avoid

extravasation.

Additional hydration is not required in the absence of vomiting, as long as oral intake is

satisfactory. If this is not the case, then intravenous fluids at "maintenance" rates (depending on

the age and weight of the child) should be given for the duration of chemotherapy.

A11.3 CADO (Cyclophosphamide, doxorubicin and vincristine)

Courses of CADO are given at 21 day intervals

DAY 1 2 3 4 5

Cyclophosphamide • • • • •

Doxorubicin • •

Vincristine • •

DRUG Dose (mg/kg) Dose (mg/m2) Administration Cyclophosphamide 10 mg/kg 300 mg/m2 5% dextrose (5 ml/kg) over 1 hr, daily x 5

days

Doxorubicin 1 mg/kg 30 mg/m2 0.9% saline over 6 hours on days 4 and 5

Vincristine 0.05 mg/kg 1.5 mg/m2 (max 2 mg) Bolus injection on days 1 and 5

A central line is essential and mandatory for the administration of doxorubicin over 6 hours.

Hydration at the level of x2 maintenance or 3 litres/m2 is required.

A11.4 Dose modification

If significant infective problems occur (CTCAE Grade 4), consider reducing the doses of

myelosuppressive therapy by 20% for subsequent courses.

If an allergic reaction occurs during the administration of Etoposide, appropriate measures should

be taken. However, the drug should be tried again with the next course at a slower rate and with

steroid premedication.

In the case of marked ptosis or other neurological deficit (other than loss of tendon reflexes),

consider reducing or omitting the next vincristine dose. In the case of CADO chemotherapy, the

second vincristine of CADO should be postponed by one week.

If there is CTCAE Grade 2 renal toxicity (see page 159), repeat GFR and modify the dose of

Carboplatin.



APPENDIX 12. RADIOTHERAPY GUIDELINES

A12.1 Indications

A12.1.1 Indication

Study group 8: Stage L2, age > 18 months, MYCN non-amplified, poorly differentiated and

undifferentiated neuroblastoma or poorly differentiated and undifferentiated

ganglioneuroblastoma nodular.

Study group 9: INSS Stage 1 neuroblastoma, MYCN amplified.

A12.1.2 No indication

Intermediate risk stage M, and Low risk neuroblastoma.

A12.2 Treatment centres

Children should only receive radiotherapy in centres suitably equipped, staffed and experienced in

paediatric radiotherapy, supported by the necessary paediatric backup facilities. This includes

radiotherapy doctors with training and experience in paediatric radiotherapy fully integrated into

the core paediatric oncology multidisciplinary team, the availability of play specialists and

paediatric anaesthetic facilities, and sufficient on-site paediatric support to deal with emergencies.

This may mean that some children cannot receive treatment in the radiotherapy centre closest to

their homes. It will need to be explained to families that this inconvenience is offset by improved

safety and quality.

Centres should be able to demonstrate compliance with external quality standards.

A12.3 Timing of radiotherapy

Radiotherapy is undertaken after completion of chemotherapy and after surgery if that has been

performed. Sufficient time needs to be allowed for the patient to recover, but radiotherapy will

normally commence between 21 and 42 days after the previous treatment.

A12.4 Assessment

The doctor responsible for radiotherapy ideally should meet with the patient and family members

as soon as it is clear that radiotherapy will form part of treatment. This initial consultation will

help to make it clear that radiotherapy is an important part of treatment, and confirm that the

radiotherapy team is part of the overall team caring for the child. An introduction to the

radiotherapy team at this stage is important as is may prevent the family from feeling that

radiotherapy is an afterthought, or has been suggested because things have not gone according to

plan, and a frank discussion should help to allay any myths or misunderstandings about

radiotherapy and its late effects.

It is helpful if at this early stage the patient and family meet not just the doctor, but also other

members of the radiotherapy team including radiographers and play specialists. Early familiarity

with the team will reduce subsequent anxieties, and will help to prevent the problems which can

arise after delayed referral.

An important part of the initial assessment is to decide whether it is likely that, with appropriate

preparation, the child will be able to cooperate with the radiotherapy team and be able to tolerate

the requirements of immobilisation awake, and feel comfortable being left alone in the treatment

room. If not, arrangements will have to be made for radiotherapy planning and treatment under

general anaesthesia. The requirement for anaesthesia is not absolute, and sometimes children need



to start the process with general anaesthesia, but with continuing input from play specialists may

be able to transition to treatment awake as they gain understanding and confidence.

Assessment of normal tissue function in organs at risk is important. For example in the case of

adrenal primary tumours it is important to know not only overall renal function but differential

function. It is advisable to have an up-to-date renal assessment including glomerular filtration rate

(GFR) measurement and dimercaptosuccinic acid (DMSA) scan.

A12.5 Informed consent

It is important that families receive accurate and complete information about the indications for,

practicalities of, and likely early and late side effects of, the radiotherapy required. This should be

given verbally by members of the radiotherapy team, and be supplemented by written

information. Information leaflets can be tailored to local circumstances by containing contact

details of important team members and links to other sources of reputable information, should the

family wish to explore these.

Children should be given relevant information in a sensitive and age appropriate manner.

Families from ethnic minority backgrounds, or speaking other languages, should have the services

of an interpreter available at every interaction with the radiotherapy team.

Children and their families should have an opportunity for their questions to be answered in full

and in an unhurried manner. Often, if there are anxieties about radiotherapy, it is recommended

that families have more than one opportunity to discuss the information and ask questions.

Finally, the receipt of informed consent for radiotherapy treatment should be documented

according to local practice.

A12.6 Immobilisation and positioning

It is essential for the accurate and safe delivery of radiotherapy that the patient does not move

during planning or treatment. As indicated above, this may require the use of daily general

anaesthesia. For young children who are to be treated awake, the input of an experienced play

specialist in preparation can be indispensable.

For abdominal tumours, and also in the more unusual circumstances of a thoracic or pelvic

tumour, the position chosen will normally be supine, with the arms resting by the sides. If field

arrangements other than anterior-posterior/ posterior-anterior (AP/PA) are envisaged for any

specific reason, the arms may need to be in a different position.

The use of a vacuum-bag, surf-board, shell, or alternative immobilisation device is recommended

according to usual departmental practice, to ensure accurate reproducibility of set-up on a daily

basis.

Departments should regularly audit the accuracy of set-up in different circumstances in order to

define appropriate margins for treatment.

A12.7 Imaging, planning and target volume definition

The target volume is defined on the basis of the post-chemotherapy extent of disease, prior to

surgery being undertaken. If surgery has taken place, the volume to be irradiated is not influenced

by the completeness or otherwise of the surgery.



It can be helpful for the child to have a planning CT scan performed in the radiotherapy

department in the treatment position after chemotherapy and before surgery. Alternatively, or in

addition, cross sectional imaging with MRI or CT can be used to determine the extent of the

disease at this point in time.

A planning CT scan will be undertaken with the patient immobilised in the treatment position

after surgery and prior to radiotherapy. This scan will be used for target volume definition and

dosimetry.

Cross sectional imaging, CT or MRI, obtained previously may be fused with the planning scan to

aid target volume definition, provided that the positioning of the patient is compatible.

Alternatively, boundaries identified on the earlier imaging may be manually transferred to the

planning scan.

The Gross Tumour Volume, GTV, defined as the post-chemotherapy, pre-surgical, extent of

disease, should be delineated on the planning scan. This will include the primary tumour and any

adjacent involved lymph nodes. Patients with remote lymph nodes involved by tumour at the time

of diagnosis have metastatic disease and are not treated in this protocol.

The Clinical Target Volume, CTV, is defined as the GTV, expanded by a margin normally 10mm

in all directions. However the CTV should be expanded from the GTV anisotropically so that it

does not extend into adjacent structures, e.g. kidney or liver, which were definitely not infiltrated

by tumour. Similarly it should exclude structures such as liver or small bowel which were not

infiltrated by disease but which have fallen back into the place occupied by tumour prior to

surgery. However, in view of the importance of uniform irradiation of complete vertebral bodies

to avoid the development of scoliosis, the CTV may need to be expanded by more than 10mm in

some directions to achieve this.

The Planning Target Volume, PTV, is defined as the CTV expanded by a margin which allows

for internal organ motion and daily set-up error. The CTV to PTV distance should be determined

by data from departmental audits, but is likely to be at least 5mm.

Planning and beam arrangements should take into account the need to give homogenous

irradiation to the PTV while minimising exposure to organs at risk.

Usually the most satisfactory beam arrangement will be a parallel opposed pair of AP/PA fields.

Sometimes it is appropriate to angle the fields slightly away from 0° and 180° to allow the medial

field borders to be non-divergent.

Occasionally more complex arrangements of beams will result in a better dose distribution in the

PTV and greater avoidance of organs at risk.

Multi-leaf collimation (MLC) or customised blocks should be used to shape fields to spare normal

tissue.

Wedges or field-in-field boosts may be used to improve dose homogeneity, but full inverse

planned intensity modulated radiotherapy with multiple non-coplanar beams is not generally

recommended as the larger volume of tissue outside the target receiving low dose radiation may

result in an increased risk of second malignancies.



Dose volume histograms showing the dose to the target volume and relevant organs at risk should

be produced to inform approval of the plan, and to document the doses received as a guide to

follow-up.

A12.8 Prospective Quality Control

It is important that radiotherapy is delivered to the highest possible standards. To achieve this all

radiotherapy plans will be reviewed and approved prospectively by the radiotherapy committee

prior to treatment. Following a radiotherapy planning scan, target volume definition and

planning/dosimetry at the treatment centre, all planning images, plans, dose volume histograms

and radiology on which radiotherapy plans are based must be sent (preferably in electronic form

on CD or alternatively in hard copy) to the Sponsor where they will be uploaded onto the

SIOPEN-R-NET database. All information sent to Sponsor should be clearly labelled with the

patient’s study number and date of birth. In case any questions arise regarding the sent

information a contact person, email address and phone number must also be supplied.

NOTE: Images must be sent to the Sponsor by priority post, ideally 10 working days prior to the

planned start date of radiotherapy, or failing that definitely to have arrived 5 working days prior to

the intended treatment start date. Treatment centres must notify the Sponsor that the information

has been sent.

The Sponsor’s address for sending the above information to is:

LINES RT PROSPECTIVE REVIEW

FUNDACION PARA LA INVESTIGACION HOSPITAL UNIVERSITARIO LA FE.


AVDA DE CAMPANAR 21

46009 VALENCIA

SPAIN

Notification that the information has been sent should be emailed to Adela Canete

([email protected]) and the Clinical Trial Unit ([email protected]).

The Radiotherapy Coordinator will be automatically notified by email of the request for a review

following uploading of the data into the SIOPEN-R-NET. Following review, a response will be

automatically generated and sent to the treatment centre which will be either to go ahead with

radiotherapy as planned or to make some modifications.

A12.9 Energy, dose and fractionation

The use of megavoltage photons from a linear accelerator is necessary. Usually 6MV photons

provide optimal treatment. Occasionally other energies, or mixed beam energies may be better.

The use of cobalt or orthovoltage equipment is contra-indicated. Electron beams will not give an

appropriate dose distribution. The use of other techniques including intraoperative treatment,

hadrons (protons or light ions) or brachytherapy is not recommended within this clinical trial.

The prescribed dose should be 21 Gy in 14 fractions of 1.5 Gy. This dose does not depend on the

response to chemotherapy or the extent of surgery. The dose is not affected by the presence or

absence of residual disease.

Sometimes it is not possible to deliver a dose of 21Gy to the PTV as defined in section above,

without exceeding normal tissue tolerance. In these cases an intelligent compromise is necessary.



It is possible to compromise on volume, or dose, or both. In most cases it is recommended that a

two-phase technique is used to spare organs at risk. A Phase 1 dose of 15 Gy in 10 fractions to the

PTV as defined above should not exceed tolerance, and then a Phase 2 dose of an additional 6 Gy

in 4 fractions may be given to a smaller volume, ideally encompassing all macroscopic residual

disease but sparing normal organs, can be used.

It is important to document clearly the rationale behind the use of any compromise technique or

dose.

Treatment should normally be one fraction per day, for five days a week. This means that the 14

fractions are delivered over 18 to 20 days. If there interruptions to treatment for medical reasons,

machine breakdown, public holidays etc. compensation should be made for the gap so that the

overall treatment time, if at all possible does not exceed 20 days. The simplest and best way of

doing this is to treat on days which are normally days off (for example treat on Saturdays around

public holidays) or give two treatments per day, with a minimum inter-fraction interval of six

hours. This approach is radiobiologically preferable to giving additional fractions, or a larger

fraction size, to compensate.

A12.10 Normal tissue tolerance

The dose to normal, uninvolved tissues should be kept As Low As Reasonably Achievable,

consistent with delivering the required radiation dose to the target volume. This is known as the

ALARA principle.

A12.10.1 Kidneys

Because of the usual location of neuroblastoma in the adrenal region, some irradiation of the

kidneys to the full dose of 21 Gy is normally inevitable. Care should be taken to ensure that

overall and differential renal function (GFR and DMSA scan) has been assessed and documented

after chemotherapy and surgery. Pre-existing renal impairment may impose additional constraints

on radiotherapy. However, assuming normal renal function, it may not be possible to have any of

the ipsilateral kidney outside the PTV, in which case the dose to at least 80% of the contralateral

renal volume should be kept below 12 Gy.

A12.10.2 Liver

The dose to the whole liver should not exceed 15 Gy. The volume of liver receiving 21Gy should

not exceed 50%. Care needs to be taken to ensure that hepatic function is normal before

radiotherapy. If there has been hepatic veno-occlusive disease as a result of chemotherapy, for

example, tolerance limits may be lower and advice should be sought from the radiotherapy

coordinator.

A12.10.3 Lungs

The whole of the lungs may safely be irradiated to a dose not exceeding 15Gy, unless lung

tolerance may have been affected by chemotherapy, in which case advice should be sought from

the radiotherapy coordinator. The volume of lung tissue receiving a greater dose than 15 Gy

should not exceed 20%.

A12.10.4 Bone

Growth of vertebral bodies and other bones is affected by radiotherapy. Care should be taken to

ensure that whole vertebrae are irradiated uniformly to prevent the development of scoliosis.

A12.10.5 Spinal cord

A dose of 21 Gy should be acceptable for any length of spinal cord.



A12.10.6 Bowel

A dose of 21 Gy should be acceptable for any volume of bowel.

A12.10.7 Bladder

A dose of 21 Gy should be acceptable for the bladder.

A12.10.8 Heart

A dose of 21 Gy should be acceptable for the heart, assuming cardiac function is normal.

A12.10.9 Gonads

Both the ovaries and testes are very sensitive to radiation. It is not likely that they will receive a

significant dose, but in the event of a pelvic tumour requiring radiotherapy, an individual

discussion will be needed, and advice should be sought from a fertility expert as to whether any

measures are available which may help to preserve fertility.

A12.11 Supportive care

The blood count should be checked before treatment. Irradiation of anaemic children should be

avoided, as anaemia is known to compromise the outcome of radiotherapy. If necessary, a blood

transfusion should be given to bring the Haemoglobin up to 12 g dl-1. Unless larger than average

volumes are to be irradiated there should be no adverse effect of radiotherapy on the blood count.

Routine blood counts during treatment are therefore unnecessary unless they are abnormal to

begin with.

Radiotherapy to the abdomen may cause nausea and vomiting, and this can usually be prevented

by the use of granisetron 20 micrograms per kilogram twice daily, or equivalent anti-emetics.

Radiotherapy may cause loss of appetite, and nutrition is important, especially in children

receiving daily general anaesthesia. The advice of a paediatric dietician and nutritional

supplements may be needed.

Radiotherapy can be stressful for children, their parents and other family measures. This can have

both a psychological and practical basis. Psychological supportive care for the family can be very

beneficial. Travelling for radiotherapy, or staying away from home, can be expensive. Families

should have access to support from social workers, or other appropriate healthcare professionals

who can advise on benefits and other practical issues.

A12.12 Geometric verification

It is important to have quality assurance procedures for radiotherapy delivery, particularly

geometric variation for set-up errors. This is usually done nowadays with electronic portal

imaging, but double exposure portal films are an alternative. Different departments may have their

own policies and procedures, but a recommended practice is shown below.

Fraction 1

¤ Acquire orthogonal image set, minimising dose to critical structures (where possible)

¤ If field edge verification is needed, where possible image all treatment fields

¤ Assess for and correct gross errors immediately

Fractions 2, 3

¤ Image orthogonal set

¤ Assess each image and correct gross errors for each fraction where necessary



Fraction 4

¤ Calculate the overall systematic error (average of the isocentric set-up error) in each

orthogonal direction

¤ Apply the systematic set-up error correction

Fraction 5

¤ If the set-up has been corrected, confirm by repeat imaging (typically two or more

fractions)

¤ If practical, calculate the new overall systematic set-up error and correct

Weekly & first day of each phase of treatment plan

¤ Image orthogonal set each week

¤ Assess each image and correct gross errors for each fraction where necessary

¤ If set-up error is greater than the tolerance value, check by repeat imaging (typically two

or more fractions)

¤ Apply any systematic set-up error correction

Daily verification may be required for treating tumours planned with very small margins or

hypofractionated techniques. Concomitant exposures should be considered. Tolerances and action

levels to use will vary, particularly with the immobilisation and treatment technique used as well

as compliance of the patient and should be chosen accordingly.

Sufficient stable anatomy needs to be visible in the verification images to ensure accurate

matching can be made. For typical neuroblastoma fields, the vertebral bodies will be the only

clearly identifiable structure. The images need to extend sufficiently in both the cranial and caudal

directions to allow the individual vertebrae to be identified. So as with small treatment beams, or

beams covering soft tissue targets only, open fields or double exposures are necessary. While the

verification images should be taken avoiding exposing dose to critical structures where possible,

by reducing the imaging borders, they do need to be large enough to be useful.

A12.13 Documentation

It is important that there is a summary of radiotherapy treatment which is accessible to treating

paediatric oncologists, and is also available in long term follow-up. This should include adequate

patient identifiers and sufficient details of the diagnosis and other principal treatments i.e.

chemotherapy and surgery, to allow it to be read in context.

With specific regard to radiotherapy, it should include as a minimum the following:

Institution where treatment given

Responsible clinician

Anaesthesia yes/no

Patient position

Immobilisation

Date of start of treatment

Date of completion of treatment

Site treated

Beam energy

Beam quality

Field set-up

Beam shaping

Total dose administered



Total Number of fractions

Fraction size

Overall treatment time

If the treatment has been given in more than one phase, dose and volume to each

Reasons for any compromise or deviation from protocol

The summary should also have at least the following images

Planning film, digitally reconstructed radiograph or portal image to show principal field for

each phase and beam shaping

Plan, at least transverse centre slice, but ideally in axial, sagittal and coronal planes

Dose volume histograms for target volume and organs at risk

A12.14 Follow-up

It is important that the patient is followed up in the long term with regard to late effects as well as

for local tumour control and survival. Periodic assessments of relevant normal organ function

should be systematically documented. Appropriate psychosocial support should be available as

the child grows up and reaches maturity to answer questions which might arise and to address

proactively other survivorship issues.



APPENDIX 13. SUPPORTIVE CARE

Treatment according to this protocol should be restricted to institutions with experience in

supporting patients with multisystem derangements and who are familiar with the administration

of combination chemotherapy.

A full range of supportive care and multidisciplinary approach should be available.

The supportive care details that appear below are guidelines only. In cases where local

institutional guidelines differ to these, the local guidelines maybe used; this is at the discretion of

the local investigator.

A13.1 Anti-Emetics

Antiemetic therapy should be administered according to institutional guidelines, e.g. Ondansetron

5 mg/m² or 0.15 mg/kg (maximum single dose 8 mg) p.o./i.v. every 8 hours.

A13.2 Hydration

Sufficient hydration (2 to 3 l/m² or twice maintenance) with appropriate electrolyte

supplementation must be provided p.o. or i.v.. Monitoring of blood pressure, cardiac and

respiratory rates, body weight, and diuresis are mandatory, especially for the CADO treatment.

A13.3 Blood component therapy

Due to the risk of graft versus host reactions in newborn/ infants and in patients on chemotherapy

all blood products (except fresh frozen plasma) should be irradiated with at least 15 Gy prior to

transfusion, according to national policies. The use of leukocyte filters for leukocyte depletion

(CMV negativity) is advised.

A13.4 Red blood cells

Keep the haemoglobin level above 8 g/dl.

A13.5 Platelets

Platelet substitution is advised when the platelets are < 10 x 109/l, and/or if there is clinical

evidence of bleeding.

A13.6 Central lines

The use of central lines is not mandatory for all patients. It is strongly recommended for the

administration of the CADO chemotherapy.

A13.7 Treatment guidelines for febrile neutropenia

All participating institutions must be familiar with managing of febrile neutropenia according to

the accepted general principles of supportive care.

During episodes of fever and neutropenia patients need to be admitted to hospital for adequate

diagnostic measures and appropriate treatment.

If there is fever (>38OC) and the neutrophil count is less than 1.0 X 109/L, then the centre’s

usual combination of broad-spectrum antibiotics should be commenced following clinical and

laboratory evaluation.

If fever persists (>38OC) for 48 hours despite broad- spectrum antibiotics, in the absence of

defined source and site of infection, then antifungal therapy should be started, regardless of

the clinical condition of the patient. Antifungal treatment can follow each treating Institution’s

policy. Liposomal amphotericin can be an effective broad spectrum treatment. Then,

appropriate renal function monitoring and potassium supplementation should be instituted



with its use. Itraconazole and voriconazole should not be considered since it should not be

combined with vincristine. In addition a chest X-ray (CXR) should be carried out.

If fever persists for a further 48 hours (i.e. a total of 96 hours), without another identified

cause complete clinical and laboratory investigation should be repeated. That could include a

chest CT, an abdominal ultrasound and a cardiac ultrasound.

According to Local Institutional policy, antibiotic and antifungal treatment can be modified.

A13.8 Pneumocystis pneumonitis prophylaxis

For patients receiving chemotherapy Pneumocystis carinii pneumonitis (PCP) prophylaxis is

advisable, but may be given according to the recommendations of each national group. PCP

prophylaxis is usually considered mandatory for patents receiving VP16- CARBO and CADO

chemotherapy.

Patients should be considered for prophylactic sulfamethoxazole/trimethoprim (5 mg TMP

/kg/day divided in two equal doses and given orally 3 days a week). For sulpha-intolerant patients

it is recommended that inhaled pentamidine or a preparation of trimethoprim only is used as

prophylaxis. PCP prophylaxis using a pentamidine nebuliser at three-weekly intervals can be

encouraged for children who are able to co-operate with jet inhalation (necessary to be effective)

which is usually only the case for children of school age. Intravenous pentamidine can also be

used.

A13.9 Nutrition

Appropriate nutrition and adequate calorie intake should be aimed for in all patients. This is

especially important for young patients with significant hepatomegaly and stage IVs disease, as

well as patients receiving VP16/ CARBO or CADO chemotherapy.

The early start of supplementary nutrition is highly recommended.

Once a 10% weight loss occurs, the institution of nasogastric alimentation with a high caloric

nutritional formula or parenteral nutrition via a central venous line if enetral feeds are not

tolerated, according to local policies is recommended.

A13.10 Renal function monitoring

Glomerular function – GFR

Serum creatinine should be monitored prior to each chemotherapy course. Glomerular

function is to be assessed according to national / group guidelines, applying either timed urine

collection, isotope clearance, or the calculated creatinine clearance.

According to Schwartz's formula [126], creatinine clearance (Ccrea) can be calculated from single

serum samples :

where F is proportional to body muscle mass, hence depending on age and gender:

- LBW infants 0-12 months F = 0.33

- AGA infants 1-12 months F = 0.45

- Children (boys and girls) F = 0.55

- Adolescent girls F = 0.55

- Adolescent boys F = 0.70

]3m²ml/min/1.7[[mg/dl] serum Crea

[cm]Height x F crea C



APPENDIX 14. SPINAL CORD COMPRESSION

A14.1 Spinal cord compression – background

The connections existing between the sympathetic nervous system and the spinal cord account

for the ability of neuroblastoma to infiltrate the intervertebral foramina with occasional

involvement of the spinal canal. Despite the fact that tumour growth almost invariably remains

extradural, it may still cause spinal cord compression which can progress to irreversible

paraplegia. Early diagnosis and prompt treatment of spinal cord compression in neuroblastoma is

therefore of critical importance.

Dumbbell neuroblastoma tends to present at a younger age, is commonly associated with intra-

thoracic disease and is significantly more frequent amongst children with localised, rather than

metastatic disease.

Evaluation of intraspinal extension has been greatly improved thanks to modern diagnostic

imaging (CT scan and MRI). MRI has proven to be especially effective in demonstrating both the

infiltration of the intervertebral foramina and the invasion of the spinal canal. On the basis of

MRI, 10-15% of children with neuroblastoma have documented foramina or intraspinal

involvement, although only half of them present with neurological signs of spinal cord

compression, and very few develop paraplegia.

Treatment of symptomatic spinal cord compression has evolved over the last few years. Until the

mid-1980s, decompressive laminectomy was usually performed in an attempt to avoid

progression to paraplegia. Surgery was frequently carried out even when paraplegia had already

developed. Radiotherapy was considered less effective, although some reports suggested that it

could be as effective as surgery in metastatic disease.

Both laminectomy and radiotherapy have potential disadvantages. The former requires very

experienced surgeons and may carry the risk of late spinal deformities. Radiation reduces and

alters the growth of vertebrae in the radiotherapy field and increases the risk of late secondary

malignancy.

The use of chemotherapy as an effective alternative to laminectomy or radiotherapy was first

reported in 1984. Since then, several authors have confirmed that chemotherapy can successfully

be used instead of surgery or radiotherapy, without compromising the chance of neurological

recovery. The concern that chemotherapy might not work rapidly enough to prevent permanent

neurological damage has been contradicted by several reported cases.

Some relevant reports concerning intraspinal neuroblastoma have been published in the last

few years.

- In 1999 Hoover et al. reported on the long term outcome of 26 patients with intraspinal

neuroblastoma treated with or without laminectomy between 1985 and 1994 [127].

Twenty four/26 are alive from 2-29 years (median, 10). Fifteen had undergone

laminectomy and 9 of them recovered including 3 with paraplegia

- Katzenstein et al. described 83 children with intraspinal neuroblastoma who were

observed between 1990-1998, of whom 43 (52%) had neurologic symptoms [128]. Two

out of five patients who had moderate deficits (paresis, bowel/blood dysfunction)

completely recovered neurologic function, vs. 17/22 who had mild symptoms, and 6/15

presenting with paralysis. Seven/24 patients who had undergone laminectomy developed

scoliosis vs. 1/49 that had been managed without laminectomy. The authors concluded that

neurologic recovery correlated with the severity of the presenting neurologic deficits.

Recovery was similar for patients treated with chemotherapy or laminectomy. However

orthopaedic sequelae were much more frequent in children who were managed with

laminectomy.

- De Bernardi et al. reported on 76 children presenting with symptomatic spinal cord

compression in the period between 1979-1998 [129]. The incidence in the overall



neuroblastoma population was 5.2%, and decreased from 9.7% in the first quinquennium

to 5.2%, 4.8%, and 2.7% in the three subsequent ones (however recent protocols do not

allow patients who are treated with chemotherapy before diagnosis to be registered). Some

children with progressive neurological symptoms are treated with chemotherapy on an

emergency basis and are thus excluded from the protocols database (an obvious bias)).

Seventy five/76 patients had motor deficit (mild in 43, moderate in 22, severe (i.e.

paraplegia) in 10), 47 had pain, 30 had a deficit of sphincter control, and 11 had sensory

loss. Treatment of spinal cord compression consisted of radiotherapy in 11 (early) patients,

laminectomy in 32 and chemotherapy in 33. Laminectomy was more frequently performed

in cases of low stage and severe motor deficits, whereas chemotherapy seemed to be

preferred in cases of advanced disease. Thirty three patients achieved full neurological

recovery, 14 improved, 22 remained stable, and 8 deteriorated, including 3 who became

paraplegic. None of the 10 patients with paraplegia recovered or improved. In the other 66

patients neurological response was comparable for the three therapeutic modalities, but

almost all the children treated with laminectomy required additional treatment. Fifty

four/76 patients are alive at the time of the analysis from 4-209 months (median 139).

Twenty six/54 survivors (44%) have late sequelae, mainly scoliosis and sphincter deficit.

The authors conclude that radiotherapy, laminectomy, and chemotherapy show a

comparable ability to relieve or improve SCC. However, patients treated with

chemotherapy did not usually require any additional therapy, whereas patients treated

either with radiotherapy or laminectomy commonly did. None of the patients presenting

with (or developing) severe motor deficit recovered or improved.

- In 2002, Plantaz et al. reported the French experience of children with localised dumbbell

neuroblastoma treated with first line chemotherapy according to the NBL 90-94 protocols.

Seventy eight cases were treated between 1990 and 1998 (Oral presentation, Meeting of

the French Society of Paediatric Oncology). Initial neurological impairment was present in

44 cases (56%), including 20 cases of severe deficit. Sixty-four patients (82%) received

first line chemotherapy. Out of the 38 patients presenting with neurological deficit who

were treated with chemotherapy, recovery was complete in 24 cases and partial in 8. Five

patients failed to recover from long-standing paraplegia. Only 1 patient deteriorated during

therapy, and recovered following emergency neurosurgery. Six patients underwent initial

laminectomy (or laminotomy) among whom 3 recovered. Finally, eight patients had

extraspinal surgery alone. Ten patients (13%) suffer from severe neurological sequelae,

and 8 (10%) from severe orthopaedic sequelae. Among the 21 patients with a spinal

residue at the end of treatment, 3 relapsed; (only 1 pure intra-spinal recurrence).

Chemotherapy enabled the size of the intraspinal mass to reduce, to improve most of the

neurological deficits, and allowed a conservative surgical, resection. Long standing

paraplegia is not improved by chemotherapy or surgery. Laminectomy should be avoided

in the treatment of dumbbell NB, both for decompression, or for the removal of residual

tumour after chemotherapy. No laminectomies have been performed in France for

dumbbell neuroblastoma since 1995.

- The European experience within INES 99.01 recorded 40 cases of dumbbell tumours of

which 25 had life threatening symptoms. Out of the 116 infant unresectable

neuroblastomas 11 (27%) underwent a primary neurosurgical approach (unpublished data).



A14.2 Diagnosis and Evaluation of Spinal Cord Compression

Diagnosis: Early detection of spinal cord compression can be difficult especially in younger

children. The most common symptoms are back pain, reduced mobility of the legs and/or arms,

sensory and sphincter dysfunction.

The presence of a motor deficit is particularly important since children who develop complete

motor loss usually experience little or no recovery. Infants with congenital dumbbell tumours

have a particularly poor outcome with regards to neurological recovery.

The extent of motor loss can be graded as follows;

0 Capable of unassisted ambulation may have pain and/or difficulty with micturition.

1 Only can walk with assistance.

2 Antigravity strength alone.

3 Presence of trace movement alone.

4 Complete motor and sensory loss.

Note: most episodes of spinal cord compression in neuroblastoma occur in infants, for whom this

scale is only partially applicable.

Evaluation: MRI is the best method to detect infiltration of the intervertebral foramina and

invasion of the spinal canal by neuroblastoma. Although a CT scan may be adequate in some

cases, it is recommended that MRI be used wherever possible, particularly in the diagnostic

work-up of infants with cervical, thoracic, or pelvic disease.

It may be difficult to accurately assess the volume of the intraspinal tumour due to the half-

moon configuration it commonly takes in that particular site. As indicated in the Imaging

Guidelines (Appendix 8, section A8.5, page 128) intraspinal tumour extension should be

considered as IDRF provided that more than 1/3 of the spinal canal in the axial plane is

invaded, and/or the preimedullary leptomeningeal spaces are not visible, and/or the spinal

canal is abnormal.

Myelography and lumbar puncture are of no diagnostic use, and are absolutely

contraindicated.

A14.3 Treatment of spinal cord compression

Spinal cord compression without symptoms

- The regular use of MRI has increased the number of cases with documented infiltration of

foramina (with or without invasion of the spinal canal). However, in the majority of cases,

especially when the intraspinal component is modest (less than 33% of the diameter), there

are no neurological symptoms.

- There is very little, if any, evidence that an asymptomatic intraspinal tumour will grow any

more after a resection of an extraspinal component. Information related to the few, well

documented cases suggests that the intraspinal neuroblastoma in patients with no

neurological symptoms tends to remain stable or even regress without specific treatment.

- If the spinal cord component occupies greater than 33% of the spinal canal, and/or the

leptomeningeal spaces are not visible and/or the spinal cord signal is abnormal the

recommendation of this protocol is to treat these patients with chemotherapy even in

the absence of signs or symptoms of spinal cord compression.

Spinal cord compression with neurologic signs

- Patients with localised neuroblastoma who present with signs of spinal cord compression

require urgent specific treatment. If neurological deficits are present and if there is clinical

progression rapid therapeutic decisions must be made in a matter of hours or at most

within 1-2 days. The decision regarding whether to administer emergency chemotherapy

to these infants with a neurological deficit should be taken after urgent discussions

between the oncologist and the neurosurgeon. Laminectomy or laminotomy is preferable



only in infants showing a very rapid neurologic deterioration. This however occurs

infrequently.

- Once it has been decided that urgent chemotherapy is needed it should never be delayed in order to obtain a pre-chemotherapy biopsy sample. There is no urgent indication to remove

the extraspinal tumour (which is likely to be unresectable, and surgery runs the risk of

worsening the neurological deficit). The tumour should be biopsied (by Tru-cut, fine needle,

or open biopsy), when the patient is stable within 7 days of starting chemotherapy. Initial

chemotherapy in this situation should never be postponed in order to try and obtain a biopsy.

- Medical treatment of Spinal Cord Compression is as follows:

- Dexamethasone 0.5 mg/kg I.V. bolus followed by 0.2 mg/kg/day I.V. in 3 divided daily

doses. Chemotherapy is given using VP/Carbo. A second course should be given 21 days

after the beginning of the first course. (See Appendix 11, page 143, of the protocol for

further details about chemotherapy doses and administration).

- A further MRI scan should be obtained following the first course of chemotherapy. If

either no improvement occurs or if deterioration of the neurological signs are observed,

and the intraspinal component has shown no response to treatment i.e. has not shrunk in

size, laminotomy and an excision of the intraspinal component should be considered.

- If the symptoms persist and the tumour remains unresectable on reassessment with MRI after

2 courses of VP/Carbo, then alternative chemotherapy should be given according to the

protocol, CADO.

- Some patients can have persistent neurological signs and symptoms from neurological

damage caused at initial presentation of the SCC. If these neurological signs are stable over 2

courses of chemotherapy and the reassessment imaging does NOT show progressive disease

it is generally not appropriate to continue on with extra courses of chemotherapy.

A14.4 Asymptomatic intraspinal residual tumour following chemotherapy

It is not necessary to remove any residual asymptomatic intraspinal tumour following chemotherapy.



APPENDIX 15. TOXICITY

Taken from CTCAE version 4.0, published: 28th May 2009 (v4.03: June 14, 2010)

A semi-colon (;) indicates ‘or’ within the description of the grade.

A single dash (-) indicates a grade is not available.

Activities of Daily Living (ADL):

Instrumental ADL refer to preparing meals, shopping for groceries or clothes, using the telephone,

managing money, etc.

Self care ADL refer to bathing, dressing and undressing, feeding self, using the toilet, taking medications,

and not bedridden.

NOTE: The following list is provided as a supportive tool for clinicians. The treatment given

within this trial has well known side effects. Therefore, for the purposes of this trial only SAEs

will be reported (see Appendix 19, page 178).

Adverse Event Grade 1 Grade 2 Grade 3 Grade 4

Blood and lymphatic system disorders

Febrile neutropenia - - ANC <1.0 x109/L with a single temperature of >38.3 degrees C (101 degrees F) or a sustained temperature of ≥ 38 degrees C (100.4 degrees F) for more than one hour.

Life-threatening consequences; urgent intervention indicated

Definition: A disorder characterized by an ANC <1.0 x109/L and a single temperature of >38.3 degrees C (101 degrees F) or a sustained temperature of ≥ 38 degrees C (100.4 degrees F) for more than one hour.

Neutrophil count decreased (x109/L)

<LLN - 1.5 <1.5 - 1.0 <1.0 - 0.5 <0.5

Definition: A finding based on laboratory test results that indicate a decrease in number of neutrophils in a blood specimen.

Platelet count decreased (x109/L)

<LLN - 0.75 <75.0 - 50.0 <50.0 - 25.0 <25.0

Definition: A finding based on laboratory test results that indicate a decrease in number of platelets in a blood specimen.

Cardiac disorders

Cardiac arrest - - - Life-threatening consequences; urgent intervention indicated

Definition: A disorder characterized by cessation of the pumping function of the heart.

Chest pain - cardiac Mild pain Moderate pain; limiting instrumental ADL

Pain at rest; limiting self care ADL

-

Definition: A disorder characterized by substernal discomfort due to insufficient myocardial oxygenation

Heat failure Asymptomatic with laboratory (e.g., BNP (B-Natriuretic Peptide)) or cardiac imaging abnormalities

Symptoms with mild to moderate activity or exertion

Severe with symptoms at rest or with minimal activity or exertion; intervention indicated

Life-threatening consequences; urgent intervention indicated (e.g., continuous IV therapy or mechanical hemodynamic support)

Definition: A disorder characterized by the inability of the heart to pump blood at an adequate volume to meet tissue metabolic requirements, or, the ability to do so only at an elevation in the filling pressure.




Gastrointestinal disorders

Abdominal pain Mild pain Moderate pain; limiting instrumental ADL

Severe pain; limiting self care ADL

-

Definition: A disorder characterized by a sensation of marked discomfort in the abdominal region.

Constipation Occasional or intermittent symptoms; occasional use of stool softeners, laxatives, dietary modification, or enema

Persistent symptoms with regular use of laxatives or enemas; limiting instrumental ADL

Obstipation with manual evacuation indicated; limiting self care ADL


Definition: A disorder characterized by irregular and infrequent or difficult evacuation of the bowels.

Diarrhea Increase of <4 stools per day over baseline; mild increase in ostomy output compared to baseline

Increase of 4 - 6 stools per day over baseline; moderate increase in ostomy output compared to baseline

Increase of ≥7 stools per day over baseline; incontinence; hospitalization indicated; severe increase in ostomy output compared to baseline; limiting self care ADL


Definition: A disorder characterized by frequent and watery bowel movements.

Dry mouth Symptomatic (e.g., dry or thick saliva) without significant dietary alteration; unstimulated saliva flow >0.2 ml/min

Moderate symptoms; oral intake alterations (e.g., copious later, other lubricants, diet limited to purees and/or soft, moist foods); unstimulated saliva 0.1 to 0.2 ml/min

Inability to adequately aliment orally; tube feeding or TPN indicated; unstimulated saliva <0.1 ml/min

-

Definition: A disorder characterized by reduced salivary flow in the oral cavity.

Mucositis oral Asymptomatic or mild symptoms; intervention not indicated

Moderate pain; not interfering with oral intake; modified diet indicated

Severe pain; interfering with oral intake


Definition: A disorder characterized by inflammation of the oral mucosal.

Vomiting 1 - 2 episodes (separated by 5 minutes) in 24 hrs

3 - 5 episodes (separated by 5 minutes) in 24 hrs

≥ 6 episodes (separated by 5 minutes) in 24 hrs; tube feeding, TPN or hospitalization indicated


Definition: A disorder characterized by the reflexive act of ejecting the contents of the stomach through the mouth.

Hepatic disorders

Hepatic failure - - Asterixis; mild encephalopathy; limiting self care ADL

Moderate to severe encephalopathy; coma; life threatening consequences

Definition: A disorder characterized by the inability of the liver to metabolize chemicals in the body. Laboratory test results reveal abnormal plasma levels of ammonia, bilirubin, lactic dehydrogenase, and alkaline phosphatase.




Injury, poisoning and procedural complications

Dermatitis radiation Faint erythema or dry desquamation

Moderate to brisk erythema; patchy moist desquamation, mostly confined to skin folds and creases; moderate edema

Moist desquamation in areas other than skin folds and creases; bleeding induced by minor trauma or abrasion

Life-threatening consequences; skin necrosis or ulceration of full thickness dermis; spontaneous bleeding from involved site; skin graft indicated

Definition: A finding of cutaneous inflammatory reaction occurring as a result of exposure to biologically effective levels of ionizing radiation.

Laboratory Investigations

Alanine aminotransferase increased

>ULN - 3.0 x ULN >3.0 - 5.0 x ULN >5.0 - 20.0 x ULN >20.0 x ULN

Definition: A finding based on laboratory test results that indicate an increase in the level of alanine aminotransferase (ALT or SGPT) in the blood specimen.

Aspartate aminotransferase increased

>ULN - 3.0 x ULN >3.0 - 5.0 x ULN >5.0 - 20.0 x ULN >20.0 x ULN

Definition: A finding based on laboratory test results that indicate an increase in the level of aspartate aminotransferase (AST or SGOT) in a blood specimen.

Creatinine increased >1 - 1.5 x baseline; >ULN - 1.5 x ULN

>1.5 - 3.0 x baseline; >1.5 - 3.0 x ULN

>3.0 baseline; >3.0 - 6.0 x ULN

>6.0 x ULN

Definition: A finding based on laboratory test results that indicate increased levels of creatinine in a biological specimen.

Nervous system disorders

Seizure Brief partial seizure; no loss of consciousness

Brief generalized seizure

Multiple seizures despite medical intervention

Life-threatening; prolonged repetitive seizures

Definition: A disorder characterized by a sudden, involuntary skeletal muscular contractions of cerebral or brain stem origin.

Vascular disorders

Thromboembolic event

Venous thrombosis (e.g., superficial thrombosis)

Venous thrombosis (e.g., uncomplicated deep vein thrombosis), medical intervention indicated

Thrombosis (e.g., Uncomplicated pulmonary embolism [venous], non-embolic cardiac mural [arterial] thrombus), medical intervention indicated

Life-threatening (e.g., pulmonary embolism, cerebrovascular event, arterial insufficiency); hemodynamic or neurologic instability; urgent intervention indicated

Definition: A disorder characterized by occlusion of a vessel by a thrombus that has migrated from a distal site via the blood stream.

NOTE: Any toxicity that results in death is considered Grade 5.

A full list of CTCAE version 4.0 toxicities is available free on the internet at:

http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm



APPENDIX 16. DRUG INFORMATION

A16.1 Carboplatin

Formulation Vials containing solution of 50 mg/5 mls, 150 mg/15 mls or 450 mg/45 mls.

Storage At room temperature.

Reconstitution Dilute in water for injection BP or 5% dextrose to a minimum strength of

0.5 mg per ml.

Stability When reconstituted with water for injection or glucose 5%, 8 hours at room

temperature, 24 hours in refrigerator. After dilution in glucose 5% infusion

stable for 24 hours in refrigerator.

Administration In this protocol as 1 hour intravenous infusion.

Supplier Commercially available.

Toxicity Frequencies Common

Happens to 21 - 100 out of

every 100 children

Occasional

Happens to 5-20 children out of

every 100

Rare

Happens to <5 children out

of every 100

Immediate: Within 1-2

days of receiving drug

Nausea, vomiting Hypersensitivity reactions2

(anaphylaxis, bronchospasm,

hypotension), constipation,

diarrhoea

Metallic taste, rash,

mucositis

Prompt: Within 2-3 -

weeks, prior to the next

course

Myelosuppression1 (anaemia,

neutropenia, leukopenia,

thrombocytopenia), Electrolyte

abnormalities (↓ Na, K, Ca,

Mg)

↑ LFT’s (Alk Phos, AST),

abdominal pain,

Nephrotoxicity (↓ GFR, ↑ Cr

and BUN), asthenia

↑ bilirubin

Delayed: Any time

later during therapy,

excluding the above

conditions

Ototoxicity (tinnitus, hearing

loss)

Peripheral neuropathy with

mild paresthesias,

diminished sense of

vibration, light touch,

pinprick, and joint position,

alopecia; temporary loss of

vision to light and colours

Late: Any time after

completion of treatment

Secondary leukaemia

Unknown

Frequency and

Timing

Foetal toxicities and teratogenic effects of carboplatin have been noted in animals and may

cause foetal harm when administered to pregnant women. It is unknown whether the drug is

excreted in breast milk. 1 Thrombocytopenia is more severe or dose limiting. 2 Hypersensitivity reactions are seen more frequently with repeated courses of therapy (after six courses in adults).

(L) Toxicity may also occur later.



A16.2 Cyclophosphamide

Formulation 100 mg, 200 mg, 500 mg and 1 G vials for reconstitution.


Stability Unreconstituted vials stable for 5 years at room temperature. A solution of

Cyclophosphamide appears to be chemically stable for at least 28 days

when stored 4 oC.

Reconstituted solution (20 mg/ml) should be used within 3 hours when

stored at room temperature, unless prepared under strict aseptic conditions,

when it may be used within 8 hours.

Administration Mesna is not required with the dose prescribed in this protocol.



Happens to 21-100

children out of every

100

Occasional

Happens to 5-20


100

Rare

Happens to <5 children

out of every 100

Immediate: Within

1-2 days of receiving

drug

Anorexia (L), nausea

(L), vomiting (L)

Abdominal discomfort,

Diarrhoea

Transient blurred vision, nasal

stuffiness with rapid

administration, arrhythmias (rapid

infusion), skin rash, anaphylaxis,

SIADH

Prompt: Within 2-3

weeks, prior to the

next course

Leukopenia, alopecia,

Immune suppression

Thrombocytopenia,

Anaemia, Hemorrhagic

cystitis (L)

Cardiac toxicity with high dose

(acute – CHF hemorrhagic

myocarditis, myocardial necrosis)

(L), hyperpigmentation, nail

changes, impaired wound healing,

infection secondary to immune

suppression

Delayed: Any time


excluding the above

conditions

Gonadal dysfunction:

azoospermia or oligospermia

(prolonged or permanent)1 (L)

Amenorrhea1 Gonadal dysfunction : ovarian

failure1 (L), Interstitial

neumonitis, Pulmonary fibrosis2

(L)


completion of

treatment

Secondary malignancy (ALL,

ANLL,

AML), bladder carcinoma (long

term use > 2 years), bladder

fibrosis

Unknown Frequency and Timing: Foetal toxicities and teratogenic effects of cyclophosphamide (alone or in

combination with other antineoplastic agents) have been noted in humans. Toxicities include: chromosomal abnormalities,

multiple anomalies, pancytopenia, and low birth weight. Cyclophosphamide is excreted into breast milk.

Cyclophosphamide is contraindicated during breast feeding because of reported cases of neutropenia in breast fed infants

and the potential for serious adverse effects. 1 Dependent on dose, age, gender and degree of pubertal development at time of treatment 2 Risk increased with chest radiation and high dose. (L) Toxicity may also occur later.



A16.3 Doxorubicin

Formulation Vials containing 10 mg or 50 mg in solution (2 mg/ml).

Vials containing 10 mg, 50 mg as powder.

Storage Solution at 2-8 oC in refrigerator.

Powder can be stored at room temperature.

Reconstitution Each 10 mg should be reconstituted with 5 mls water for injection.

Solution should be further diluted in 0.9% saline or 5% dextrose.

Stability Powder 4 years at room temperature. Solution 2 years (2-8 oC in

refrigerator) or 1 month at room temperature.

Reconstituted solution (100 g/ml) in 5% dextrose or 0.9% saline is stable

for 28 days when stored in refrigerator.

Solutions should be protected from light during storage and administration

unless concentration is > 500 g/ml and freshly prepared. Substantial

photodegradation may occur at concentrations below 100 g/ml if

exposed to light.

Administration The drug should be mixed with 0.9% saline and given over 6 hours.



Happens to 21-100

children out of every 100

Occasional

Happens to 5-20 children out of

every 100

Rare

Happens to <5 children out of

every 100

Immediate:

Within 1-2 days of

receiving drug

Nausea, vomiting, pink or

red colour to urine, sweat,

tears and saliva

Hyperuricemia, facial flushing,

sclerosis of the vein

Diarrhoea, anorexia,

erythematous streaking of the

vein (flare reaction),

extravasation (rare) but if occurs

= local ulceration, anaphylaxis,

fever, chills, urticaria, acute

arrhythmias

Prompt:

Within 2-3 weeks,

prior to next course

Myelosuppression

(leukopenia,

thrombocytopenia,

anaemia),

alopecia

Mucositis (stomatitis and

esophagitis), hepatotoxicity

Radiation recall reactions,

conjunctivitis and lacrimation

Delayed:

Anytime later during

therapy

Cardiomyopathy1 (CHF occurs

in 5-20% @ cumulative doses

≥450mg/m²)(L)

Cardiomyopathy1 (CHF occurs

in

<5% @ cumulative doses

≤400mg/m²) (L), ulceration and

necrosis of colon,

hyper-pigmentation of nail bed

and dermal crease, onycholysis

Late:

Anytime after

completion of therapy

Subclinical cardiac

dysfunction

CHF (on long term follow up in

paediatric patients)

Secondary malignancy (in

combination regimens)

Unknown Frequency

and Timing:

Foetal and teratogenic toxicities. Carcinogenic and mutagenic effects of doxorubicin have been

noted in animal models. Doxorubicin is excreted into breast milk in humans. 1Risk increases with chest radiation, exposure at a young or advanced age. (L) Toxicity may also occur later.



A16.4 Etoposide (VP16)

Formulation Vials containing 100 mg Etoposide in 5 ml.


Reconstitution Ideally dilute to a concentration of 0.25-0.4 mg/ml in 0.9% sodium

chloride.

Stability Vials are stable for 5 years at room temperature. At concentrations of 0.4

mg/ml in 0.9% saline solutions are stable for 96 hours at room temperature

in normal fluorescent lighting, in PVA containers.

Solution in PVC infusion bags should be used immediately, to avoid

leaching out of potential carcinogenic plasticisers.

Administration By intravenous infusion over 2-4 hours (protected from light).

Caution: Anaphylactic reaction usually manifested as severe hypotension

may occur if infusion given too rapidly. Avoid extravasation.



Happens to 21 - 100


Occasional

Happens to 5-20 children

out of every 100

Rare


out of every 100

Immediate: Within


drug

Nausea, vomiting Anorexia Transient hypotension during

infusion; anaphylaxis (chills, fever,

tachycardia, dyspnoea,

bronchospasm, hypotension)

Prompt: Within 2-3

weeks, prior to next

course

Myelosuppression (anaemia,

leukopenia), alopecia

Thrombocytopenia,

diarrhoea, abdominal pain,

asthenia, malaise, rashes

and urticaria

Peripheral neuropathy,

Mucositis, hepatotoxicity, chest pain,

thrombophlebitis, congestive heart

failure, Stevens-Johnson Syndrome,

exfoliative dermatitis

Delayed: Any time


excluding the above

conditions

Dystonia, ovarian failure,

amenorrhea, anovulatory cycles,

hypomenorrhea, onycholysis of nails


completion of treatment Secondary malignancy (preleukaemic

or leukemic syndromes)

Unknown Frequency and Timing: Foetal toxicities and teratogenic effects of etoposide have been noted in animals at

1/20th of the human dose. It is unknown whether the drug is excreted in breast milk.



A16.5 Vincristine

Formulation 1 mg, 2 mg, 5 mg vials with 10 mls diluent. 1 ml, 2 ml, 5 ml vials of

solution 1 mg/ml.

Also available in pre-filled syringes containing 1 mg in 1 ml, 2 mg in 2 mls

unpreserved.

Storage At 2-8 oC in refrigerator.

Stability Depends on formulation: Lyophilised powder (0.6 oC) 3 years. Solution (0-

6 oC) 2 years. Reconstituted injection is stable for 14 days (2-8 oC).

Diluted infusion (in 0.9% saline, 5% dextrose or Ringer’s lactate) is stable

for 24 hours at 20 g/ml.

Administration By bolus intravenous injection. Ensure that needle is well into the vein to

avoid extravasation.



Happens to 21 -100


100

Occasional


out of every 100

Rare


out of every 100

Immediate: Within


drug

Jaw pain, headache Extravasation (rare) but if occurs =

local ulceration, shortness of breath,

and bronchospasm

Prompt: Within 2-3

weeks, prior to the next

course

Hair loss, constipation Weakness, abdominal pain,

mild brief myelosuppression

(leukopenia, thrombocytopenia,

anaemia)

Paralytic ileus, ptosis, diplopia, night

blindness, hoarseness, vocal cord

paralysis, SIADH, seizure, defective

sweating

Delayed: Any time


excluding the above

conditions

Loss of deep tendon

reflexes

Peripheral paresthesias

(including numbness, tingling

and pain); clumsiness; wrist

drop, foot drop; abnormal gait

Difficulty walking or inability to

walk, sinusoidal obstruction

syndrome (SOS, formerly VOD) (in

combination), blindness, optic

atrophy, urinary tract

disorders (including bladder atony,

dysuria, polyuria, nocturia, urinary

retention), autonomic neuropathy

with

postural hypotension; 8th cranial

nerve damage with dizziness,

nystagmus, vertigo and hearing loss


the completion of

treatment

Unknown Frequency and Timing: Foetal toxicities and teratogenic effects of vincristine (either alone or in combination

with other antineoplastic agents) have been noted in humans. The toxicities include: chromosome abnormalities,

malformation, pancytopenia, and low birth weight. It is unknown whether the drug is excreted in breast milk.



A16.6 13-cis-retinoic acid (Isotretinoin®, Roaccutane®)

Source and Pharmacology: The exact mechanism of RA-induced maturation of tumour cells is

not known. It has been observed that cyclic AMP-inducing agents have a synergistic effect on

RA-induced differentiation of the HL-60 promyelocytic leukemic cell line. The observers have

hypothesised that RA may induce increased levels of a cAMP-dependent protein kinase whose

activity is potentiated by increased intracellular cAMP levels. The role of cAMP-dependent

protein kinases in cellular differentiation has been documented. RA also appears to enhance

normal haematopoietic differentiation by increasing the responsiveness of myeloid and erythroid

progenitor cells to the action of myeloid colony stimulating activity and erythropoietin,

respectively.

Metabolism: RA is 99.9% bound in plasma (almost entirely to albumin) and has a half-life of 10-

20 hours. The major metabolite is 4-oxoisotretinoin, and excretion is in the urine and faeces. A

single oral dose of 100mg/m² 13-cis retinoic acid will produce peak plasma levels of 1-2mM. The

mean peak-time was 3.2 hours after 80mg orally, with a terminal t½ of 10 to 20 hours.

Supplier: Commercially available.

Guidelines for cutting Isotretinoin capsules:

Isotretinoin (13-cis retinoic acid) will be provided in 5mg and 20mg capsules depending on the

total daily dose required. All capsules are blister packed. The following guidelines have been

developed to maximise the amount of drug recovered from the capsule and to minimise the risk of

skin contamination especially to women of childbearing age.

Gloves must be worn for this procedure.

1. Remove capsule from blister pack and transfer required number of capsules for each dose to

the plastic medicine pot.

2. Assemble equipment:

- 1 pair non-sterile gloves

- small pair sharp clean scissors (to be used only for this purpose)

- 1 dessert spoon

- 1 teaspoon

- 1 small tray

- small portion of ice cream/yoghurt

- kitchen roll – kept just for this purpose

- cytotoxic waste bin

3. Put on gloves.

4. Place dessert spoon on clean surface.

5. Take a capsule between finger and thumb and hold upright firmly. With the scissors snip the

tip off the capsule into the tray to avoid any possible harm to the eyes.

6. Carefully squeeze the contents of the capsule onto the dessert spoon.

7. Discard empty capsule in cytotoxic waste bin.

8. Use kitchen roll to wipe any drug from gloves then dispose of kitchen roll immediately in the

cytotoxic waste bin.

9. Repeat steps 5 to 8 for each capsule needed.

10. After all the required capsules have been snipped, use the teaspoon to place some soft ice

cream or yoghurt onto the dessert spoon.

11. Using the teaspoon mix the ice cream/yoghurt and medicine together.

12. Give medicine to child.

13. Clean surface with kitchen roll and wash all equipment, including scissors, in hot soapy water.

14. Dispose of gloves in cytotoxic waste bin.

15. Wash hands thoroughly.

16. Return cytotoxic waste bin to hospital when full.




Happens to 21-100


Occasional


out of every 100

Rare

Happens to <5 children out of every

100

Immediate: Within


drug

Nausea and vomiting

Anaphylaxis, bronchospasm

Prompt: Within 2-3

weeks, prior to the

next course

Dry skin (L), dry mucosa

(L), Epistaxis, cheilitis

(L), Photosensitivity,

Elevated ESR, back pain

(L), Arthralgias (L),

Triglyceride elevation (L),

Hypercalcemia (L)

Rash (L), conjunctivitis (L),

headache (L), decrease in

high density lipoproteins

(L), cholesterol elevations

(L), transaminase elevations

(L), anaemia (L)

Alopecia, appetite disturbances,

Hyperglycemia, Hyper or hypo skin

pigmentation, nail changes, eruptive

xanthomas, seizures, dizziness,

Pseudotumor-cerebri (papilledema,

headache, Nausea, vomiting, visual

disturbances), Psychiatric disorders

(depression, aggressive and/or violent

behaviours, suicidal ideation, suicide,

dream disturbances), insomnia,

lethargy, malaise, nervousness,

paresthesias, Weight loss,

myelosuppression, elevated platelet

counts, agranulocytosis, allergic

vasculitis (L), chest pain, pancreatitis

(including very rarely fatal

hemorrhagic pancreatitis), hearing

impairment, Inflammatory bowel

disease, visual disturbances (decrease

in night vision, corneal opacities

which resolve on d/c, photophobia,

colour vision disturbances, cataracts),

oedema, inflammation of the gums,

Drying of respiratory tract with voice

alteration, respiratory infections

Delayed: Any time


excluding the above

conditions

Skeletal hyperostosis Osteoporosis, bone fractures or

delayed healing, premature

epiphyseal closure, Rabdomyolysis,

abnormal menses, renal disturbances

(WBC in urine, proteinuria,

haematuria, renal calculi),

calcification of tendon and ligaments

Unknown frequency and timing: Major human foetal abnormalities related to isotretinoin administration in females have

been documented. There is an increased risk of spontaneous abortion. In addition, premature births have been reported.

Documented external abnormalities include: skull abnormality; ear abnormalities (including anotia, micropinna, small or

absent external auditory canals); eye abnormalities (including microphthalmia); facial dysmorphia and cleft palate.

Documented internal abnormalities include: CNS abnormalities (including cerebral abnormalities, cerebellar malformation,

hydrocephalus, microcephaly, cranial nerve deficit); cardiovascular abnormalities; thymus gland abnormality; parathyroid

hormone deficiency. In some cases death has occurred with certain of the abnormalities previously noted. Cases of IQ

scores less than 85 with or without obvious CNS abnormalities have also been reported. It is not known whether this drug is

excreted in human milk. Because of the potential for adverse effects, nursing mothers should not receive isotretinoin.

(L) Toxicity may also occur later.



APPENDIX 17. DIFFERENTIATION THERAPY WITH 13-CIS RETINOIC

ACID

A17.1 Indication

Indication

Study group 8: Stage L2, age > 18 months, MYCN non-amplified, poorly differentiated and

undifferentiated neuroblastoma or poorly differentiated and undifferentiated

ganglioneuroblastoma nodular.

Study group 9: INSS Stage 1 neuroblastoma, MYCN amplified.

No indication

Intermediate risk stage M, and Low risk neuroblastoma.

These patients will receive 6 cycles of 13-cis RA. Each full cycle consists of two weeks (14 days)

of 13-cis RA given by mouth at a dose of 160mg/m²/day followed by two weeks of rest. Each

cycle is repeated at 28 day intervals. If irradiation is given, differentiation therapy should be

started two weeks after the end of local irradiation, provided that the ANC > 0.5 x 109/l, liver

function < CTC grade 2 toxicity, and renal function, calcium, uric acid and triglycerides ≤ 2 x

normal values. Common toxicity criteria (CTC) can be found in Appendix 14 on toxicity (page

159).

Patients will receive 13-cis-RA 160 mg/m²/day divided into two equal doses, orally Patients ≤

12 kg will be given 5.33 mg/kg/day divided into two equal doses given orally twice a day. Doses

will need to be rounded to the nearest 10 mg. Capsules come as 10, 20, and 40 mg sizes.

It is preferable to administer each p.o. dose without dissolving or trying to modify the capsules. It

can be taken with food and liquids e.g. water, milk, juice. In case of great difficulties, the capsules

can be emptied into a high fat food such as ice cream or peanut butter to administer (see

instructions in Drug Information Section, 13-cis-retinoic acid, page 167).

A17.2 Treatment schedule for 13-cis-RA

W1 W2 W3 W4 W5 W6 W7 W8 W9 W10 W11 W12

RA RA RA RA RA RA

rest rest rest rest rest rest

/ / / / / / / / / / / /

W13 W14 W15 W16 W17 W18 W19 W20 W21 W22 W23 W24

RA RA RA RA RA RA

rest rest rest rest rest rest

/ / / / / / / / / / / /

W: week

A17.3 Suggested supportive care

Topical Vitamin E should be applied to the lips twice a day during 13-cis-RA therapy if

cheilitis develops.

Patients should avoid direct sun exposure while on 13-cis-RA.

Patients should avoid exposure to vitamin A products during 13-cis-RA therapy.



A17.4 Criteria prior to each cycle of 13-cis-RA

ALT < 5 x normal.

Skin toxicity no greater than grade 1.

Serum triglycerides < 300 mg/dl.

No haematuria and/or proteinuria on urinalysis.

Serum creatinine < 1.5 mg/dl.

A17.5 Dose modifications

A dose reduction of 25% (to 120 mg/m²/day or 4 mg/kg/day if child weighs < 12kg) for

subsequent cycles should be made for the occurrence of any grade 3 or 4 CTC toxicity,

EXCLUDING: grade 3 or 4 haematologic, grade 3 hepatic, grade 3 nausea, grade 3 vomiting,

or grade 3 fever. If the same grade 3 or 4 toxicity recurs after a 25% dose reduction, then

decrease the dose by another 20% (to 100 mg/m²/day or 3.33 mg/kg/day if child weighs ≤ 12

kg). If the same grade 3 or 4 toxicity recurs after two dose reductions, then discuss with study

co-ordinator before continuing further therapy.

It has been reported (rarely) that some patients treated with 13-cis-RA develop new areas of

abnormal uptake on bone scan. This is likely to be due to increased bone resorption. If such

changes occur during the 13-cis-RA phase in the absence of any other evidence of tumour

recurrence, discuss with study co-ordinator before reporting as disease progression.

If the criteria to begin the next cycle are not met by the date the cycle is due to begin, delay

the cycle for one week. If the criteria are still not met, treat at 25% dose reduction (120

mg/m²/day or 4 mg/kg/day if child weighs ≤ 12 kg). An additional dose reduction to 100

mg/m²/day (3.33 mg/kg/day if child weighs ≤ 12 kg) should occur if criteria are not met within

one week after due date for subsequent cycles.

If serum creatinine increases by > 50% in any cycle, measured GFR should be carried out

prior to commencing the next cycle. If GFR is < 50 ml/min/1.73 m², then call the study co-

ordinator for dose adjustment.

If patient develops haematuria, proteinuria, and/or hypertension during any cycle of therapy,

withhold medication and contact study co-ordinator.

For localised cheilitis, apply topical vitamin E to lips for subsequent cycles. If this does not

control symptoms sufficiently to allow sufficient oral intake, then decrease dose by 25% to

120 mg/m²/day or 4 mg/kg/day if child weighs ≤ 12kg.

If serum triglycerides are > 300 mg/dl when next cycle is due, delay starting therapy for two

weeks. If still > 300 mg/dl, then start patient on medical therapy for serum triglyceride

reduction and begin cycle at previous 13-cis-RA dosage. If serum triglycerides are < 300

mg/dl by time subsequent cycle is due, then continue at same dosage 13-cis-RA. If

triglycerides are still > 300 mg/dl after one cycle on medical therapy, then reduce 13-cis-RA

dosage by 25% for subsequent cycles.



APPENDIX 18. ORGANISATIONAL AND ADMINISTRATIVE ISSUES

This section is intended to help participating institutions understand the organisational and

administrative issues involved in this clinical trial. To avoid confusion the different roles are

defined as follows:

Sponsor – is the institution which takes responsibility for the initiation, management

and/or financing of the clinical trial. The full name and contact address of the sponsor for

this clinical trial are detailed in section A18.1.2 below.

National Coordinating Centre – is the institution which takes responsibility for the conduct

of the clinical trial in that particular country.

National Coordinator – is the clinician at the National Coordinating Centre who is

responsible for the conduct of the clinical trial in that particular country.

Local Principal Investigator (PI) – is the clinician at the participating institution who is

responsible for the conduct of the clinical trial at that institution.

Local Co-investigator – is an individual at the participating institution who assists the PI

with the conduct of the clinical trial at that institution.

A18.1 Ethical and legal aspects and agreements

A18.1.1 Good Clinical Practice/Declaration of Helsinki

This study will be conducted according to the principles of the Declaration of Helsinki (Appendix

23, page 226) and all its revisions, pertinent national laws and regulations, as well as the

International Conference on Harmonization (ICH)’s Good Clinical Practice, taking into account

the Directive 2001/20/EC of the European Parliament and of the Council April 2001.

A18.1.2 Sponsorship

The Sponsor of the international clinical trial LINES in the legal sense as defined in the Directive

2001/20/EC of the European Parliament and of the Council April 2001 is:

FUNDACION PARA LA INVESTIGACION HOSPITAL UNIVERSITARIO LA FE.


AVDA DE CAMPANAR 21

46009 VALENCIA

ESPAÑA

The Sponsor transfers its duties for every participating country to an authorised institution by

written agreement. This authorised institution will be the National Coordinating Centre for that

country. The National Coordinating Centre will fulfil the transferred duties for the sponsor and

warrants the compliance with all the statutory provisions relevant for the Sponsor.

The Sponsor reserves the right to audit the National Coordinating Centre to control adherence to

all legal requirements.

A18.1.3 Insurance

The National Coordinating Centre of participating countries are responsible for the provision of

insurance or indemnity to cover the liability of the National Coordinator, Local Principal

Investigators, Local Co-investigators and the Sponsor in the respective country, as required by the

directive 2001/20/EC of the European Parliament and of the Council of 4 April 2001 and the

corresponding national laws.

Insurance must be obtained before the initiation of the study.



A18.1.4 Ethics committee (EC)/Institutional Review Board (IRB) approval

The National Coordinating Centre is responsible, on behalf of the Sponsor, for ensuring that every

participating institution within that country has gained approval for conducting the clinical trial

from an appropriate EC/IRB. Copies of the approval should be sent to the Sponsor.

Approval of the EC/IRB must be obtained before initiation of the study.

A18.1.5 Competent Health Authority approval

The Sponsor is responsible for registering the study in the EUDRACT database.

The National Coordinating Centre is responsible, on behalf of the Sponsor, for ensuring that every

participating institution within that country has gained approval for conducting the clinical trial

from an appropriate competent authority. Copies of the approval should be sent to the Sponsor.

Authorisation from the competent authority must be obtained before study initiation.

A18.1.6 Financing

This is a collaborative clinical trial which will be carried out in kind by the participating

institutions. No payments will be made for participation in this clinical trial. Participating

institutions will provide their own financing.

A18.1.7 Informed consent

The Local Principal Investigator, or Local Co-investigator, is responsible for the collection of

written informed consent from each parent/patient participating in the study, after a clear

explanation of the study objectives, design, methods, potential benefits and hazards, data

protection/confidentiality, as well as explaining alternative therapeutic options. In addition, each

parent/patient must have read and understood the Parent and Patient Information Leaflet (PIL)

written in the language of the patient.

The Local PI or Co-investigator must specify to parents/patients that they are free to refuse to

participate in the study and can withdraw from the study at any time and for whatever reason,

without any prejudice on the quality of care that the patient has the right to expect.

It is recommended that parents/patients should have adequate time between being given the

information on the study and being asked to sign the Informed Consent Form (ICF).

The ICF must be dated and signed off by either the Local Principal Investigator (or Co-

investigator) and the patient (and/or their legal representative(s)) in duplicate. One copy is kept by

the patient and the original by the Local PI or Co-investigator.

Consent must be obtained for low and intermediate risk study group patients prior to study

registration using the appropriate consent form (see page 186). Consent should also be obtained

for study group assignment using the low and intermediate risk patient informed consents

(Appendix 21, page 186). Patients who are not eligible for enrolment, and are registered only

must also have consent obtained.

Consent must be obtained for neonatal patients prior to registration.

If new data becomes available that could significantly modify the potential risks of the study or

the consent of the patients, the PIL will be reviewed and updated accordingly. All patients,

including those already on study treatment, must be provided with these new data and need to

give their consent to pursue the study.



A18.2 Data management and protection/confidentiality

A18.2.1 Data Collection/Submission

The LINES trial will be managed via a web based system. Each of the participating institutions

will enter data into the Remote Data Entry (RDE) system hosted by SIOPEN-R-NET.

It is the responsibility of the Local Principal Investigator to insure that the information submitted

is true and has not been falsified in any way.

An audit trail of all changes concerning the contents of the clinical trial database will be

automatically recorded.

Low risk patients without a genomic profile (section 9.4.7, page 72) and intermediate risk L2

patients without a histotype (section 10.4.5, page 91) will be registered and will be study patients

(rather than trial patients). Low risk patients should be consented and the full data set entered.

Intermediate risk patients should be consented and have regular patient status (according to

A18.8, page 176) information entered.

A18.2.2 Registration of a new patient

New patients should be registered via the RDE system as soon as they are identified as a possible

eligible patient and consent for registration has been obtained. After registration the AMED data

is completed. After the completion of the AMED, an automatic notification is sent to signal that

low risk patients should be fast-tracked for biology and intermediate risk L2 patients should be

fast-tracked for pathology.

The SIOPEN-R-NET system will automatically assign the stage and study group for each patient.

In order for this to be possible, the CRFs containing the diagnostic information must be completed

within six weeks from the date diagnosis (see section 8.1, page 39, for the definition of the date of

diagnosis). A patient will be eligible for LINES only within this six week period.

.A patient must be enrolled in LINES and have the study group assigned (i.e. successfully submit

the eligibility form) prior to starting trial treatment. With the following exceptions:

L2 tumours > 18 months without MYCN amplification (i.e. study groups 7 and 8) who may

receive the first two cycles of VP/Carbo while waiting for the pathology review.

Low risk group patients who have LTS may receive the first two cycles of VP/Carbo while

waiting for the biology review.

INRG stage M neuroblastoma, MYCN non-amplified, age <12 months (i.e study group 10)

For these select groups of patients, enrolment into LINES and the assignment of the study group

must be done after no more than the two cycles of VP/Carbo for the patient to remain eligible for

LINES.

NOTE: All patients must have the eligibility form successfully submitted within six weeks from

diagnosis to remain eligible for LINES.

A18.2.3 Randomisation

Patients in study group 1 will be randomised to either chemotherapy or observation.

Randomisation will be stratified according to country and is completed online via the RDE

system. Randomisation must take place within six weeks from the date of diagnosis (see section

8.1, page 39, for the definition of the date of diagnosis).

If access to the RDE system is not possible, the National Coordinating Centre should be

contacted. It is the responsibility of the National Coordinating Centre to ensure that access



problems with the RDE system are fixed so that the patient maybe randomised in a timely

manner.

A18.2.4 Data protection/confidentiality

In accordance with the directive 2001/20/EC of the European Parliament and of the Council of 4

April 2001 and the corresponding national laws all data will be handled strictly confidentially.

Throughout documentation and analysis, clinical trial participants will be identified only by the

unique patient code. Indentifying information will not be published.

Patient data will not be made available to third parties, expect for official representatives of the

Sponsor and regulatory authorities. Trial staff and third parties who have access to patient data

are sworn to secrecy.

A18.2.5 Data storage

All submitted data will be stored electronically. In accordance with the directive 2001/20/EC of

the European Parliament and of the Council of 4 April 2001, the RDE system will ensure:

Access only to authorised personal,

Appropriate and regular backup on electronic media of all data, to permit restoration in

case of loss or damage of the database,

Operation tracking log for each user,

Electronic data audit trails,

Disaster recovery procedures.

A18.2.6 Data transfer

In accordance with the directive 2001/20/EC of the European Parliament and of the Council of 4

April 2001 and the corresponding national laws the unique patient code will ensure that the

transfer of data will be secure, anonymous and encrypted. Patients enrolled in LINES from

outside the EU will also use the unique patient code.

The RDE system will ensure that data transfer is in accordance with the directive 2001/20/EC of

the European Parliament and of the Council of 4 April 2001. It is the responsibility of the

National Coordinator to ensure that transfer of data from their country is in accordance with the

corresponding national laws.

A18.3 Safety pharmacovigilance instructions

The treatment given within this trial has well known side effects. Therefore, for the purposes of

this trial only SAEs will be reported.

Serious Adverse Events (SAEs) will be reported by the Local Principal Investigators to the

Sponsor within 24 hours via the SIOPEN-R-NET using the SAE report form. The Sponsor will

report all SAEs according to current regulations.

For more details on SAE reporting, see Appendix 19 (page 178).

A18.4 Quality control and quality assurance

A18.4.1 Data monitoring committee

An independent data monitoring committee (DMC) consisting of three international experts will

monitor the progress of the clinical trial on ethical and scientific grounds. Review by the DMC

will take place every 12 months.

The DMC receives confidential information on the clinical trial and has the role to:

Review the patient accrual,

Examine the interim analysis,



Monitor the toxicity of treatment,

Examine other pertinent clinical trials,

Review any major modification of the clinical trial proposed by the clinical trial

committee prior to its implementation.

Once the DMC review is complete, recommendations regarding conduct, modification and early

termination are given to the study chair.

A18.4.2 Monitoring

The National Coordinating Centre is responsible for the organisation of an adequate monitoring

process in the respective country. It is the responsibility of the National Coordinating Centre to

insure the quality and accuracy of all data submitted for their country.

A18.4.3 Audits

To guarantee that the conduct of the study is in accordance with ICH-GCP and the national laws,

the Sponsor or his legal representatives reserves the right to audit selected participating

institutions. The auditor will be independent from the staff involved in the proceedings of this

clinical trial.

A18.4.4 Inspections

Inspections of participating institutions maybe performed by the competent authorities at any time

during or after the completion of the clinical trial.

A18.5 Administrative issues

A18.5.1 Protocol compliance

After a patient has been enrolled, it is the responsibility of the Local Principal Investigator to

make every effort that compliance with the protocol is kept.

Protocol violations are any deviations from the procedures outlined in the clinical trial protocol.

That is:

Missed evaluations/incorrect timing of evaluations,

Non-compliance with study medications/intake of prohibited medications, this includes the

inappropriate initiation of chemotherapy for patients under observation.

Major protocol violations need to be reported immediately to the National Coordinating Centre

and the Sponsor. All protocol violations will be listed and discussed with the Sponsor and the

study statistician prior to statistical analysis.

Under practical working conditions, some minor deviations may occur due to circumstances

beyond the control of the Local Principal Investigator. All such deviations will be documented in

the patient’s records, together with the reason for the occurrence, and where appropriate, detailed

in the study report.

A18.5.2 Protocol amendments

Any significant modification of the clinical trial must be approved by the EC/IRB and the

competent authorities, before being undertaken.

No modification can be set up prior to these authorisations; unless changes are necessary to

prevent immediate hazards that would pose a threat to study patients, or they are changes

pertaining only to administrative/logistic aspects (e.g. change of CRA, of telephone number,

etc…).



The Sponsor or regulatory authorities may prematurely stop the study at any time.

A18.5.3 End of the study

Within 90 days of the end of the clinical trial the Sponsor shall notify the competent authorities,

ethics committee, and National Coordinating Centres that the clinical trial has ended. If the

clinical trial has been terminated early, this period shall be reduced to 15 days and the reasons

clearly explained.

The National Coordinating Centre of the participating country is responsible for the notification of

the end of the clinical trial to the competent authority and the EC/IRB in the respective country,

taking into account national laws.

A18.5.4 Publication rules

For publication of (or sections of) this study, the SIOPEN publication rules should be applied.

These are publically available.

A18.6 Central review

Central review is required for pathology and biology.

A18.6.1 Pathology

1 HE-stained section from each paraffin block needs to be sent to the responsible reference

pathologist within two weeks of finalisation of the local histopathological report.

For patients with an undifferentiated tumour one block or unstained sections are also required for

immunohistochemical studies.

The responsible reference pathologist will return his/her report to the local institution and

National Coordinating Centre within 4 weeks.

For more information see the Pathology Guidelines (Appendix 6, page 116).

A18.6.2 Biology

In case of surgical biopsies provide a fresh and frozen tumour sample. If the sample is small, it

should be frozen only.

In case of needle-core biopsies a minimum of 2 biopsies (preferably 3) snap-frozen in separate

microtubes are required.

The schedule for the results of the mandatory investigations (section A7.1) can be found in the

Biology Guidelines Appendix (Appendix 7, page 121).

For more information see the Biology Guidelines (Appendix 7, page 121).

A18.7 Removal from trial treatment

All patients no longer under a trial treatment strategy and/or receiving other treatment should be

followed until death, lost to follow-up or until they withdraw their consent for data collection.

Follow-up data for these patients should be submitted according to section A18.8.

A18.8 Follow-up

A report of the patient status using the appropriate CRF should be submitted every 6 months

(from diagnosis, i.e. the first follow-up should be completed 6 months following diagnosis,

regardless of whether or not the patient is still receiving treatment) for the first 5 years and then

annually until 10 years from diagnosis.

Information regarding long term effects or sequelae should be completed using the appropriate

CRF every 12 months from diagnosis.





APPENDIX 19. GUIDELINES FOR REPORTING TOXICITY AND SERIOUS

ADVERSE EVENTS (SAES)

The processing of Serious Adverse Events in a timely manner is a regulatory requirement. It is

also vital in order to safeguard the safety of patients entered into any trial. The treatment given

within this trial has well known side effects. Therefore, for the purposes of this trial only serious

adverse events (SAE) will be reported. The definitions of adverse event, adverse reaction and

unexpected adverse reaction are given below for reasons of completeness.

The information in this section is in line with ICH Good Clinical Practice Guidelines and the

European Directive 2001/20/EC.

A19.1 Definitions

A19.1.1 Adverse Event (AE)

Any untoward medical occurrence in a patient or clinical trial subject administered a medicinal

product and which does not necessarily have a causal relationship with this treatment.

That is, any unfavourable and unintended sign (including abnormal laboratory findings),

symptom, or disease temporally associated with the use of a medicinal product, whether or not it

is considered related to the medicinal product.

Please also note the following tips:

Adverse Events include:

Exacerbation/worsening of a pre-existing condition.

Increased frequency or intensity of a pre-existing episodic event or condition.

Conditions diagnosed after the start of the study even though they were possibly present

prior to the start of the study.

Adverse Events do not include:

Medical or surgical procedures (the indication leading to the procedure is an AE).

Pre-existing diseases and conditions that do not change.

The disease being studied (e.g. disease progression).

Death (death is an outcome; the underlying cause is an event).

A19.1.2 Adverse Reaction (AR)

Any untoward and unintended medical occurrence in a patient or clinical trial subject

administered a medicinal product for which a causal relationship is suspected.

That is, all adverse events judged by the reporting physician as having a reasonable causal

relationship to the medicinal product qualify as adverse reactions. The expression “reasonable

causal relationship” is defined as the case where there is proof or argument to suggest a causal

relationship.

A19.1.3 Unexpected Adverse Reaction (UAR)

Any adverse drug reaction where the nature or severity is not consistent with the applicable

product information.

That is, any expected event with a more sever outcome or increased rate of occurrence. Or an

event judged by the reporting physician as having a reasonable causal relationship to the

medicinal product, but is not documented in the applicable product information.



A19.1.4 Serious Adverse Events (SAE) or Reaction (SAR)

Any adverse event or reaction that:

o results in death,

o is life threatening (at the time of the event),

o requires in-patient hospitalisation or prolongation of existing hospitalisation,

o results in persistent or significant disability/incapacity, or

o is a congenital anomaly/birth defect.

A19.1.5 Suspected Expected Serious Adverse Reaction (SESAR)

Any serious adverse reaction that is documented in the applicable product information.

A19.1.6 Suspected Unexpected Serious Adverse Reaction (SUSAR)

Any serious adverse reaction where a causal relationship can not be excluded and the nature or

severity is not consistent with the applicable product information.

A19.2 Documentation and reporting of events

Events, regardless of cause, can occur from the time the informed consent is signed up until 30

days after the last day of treatment or observation, for those patients who are observed, on the

LINES protocol. Patients who withdraw (for any reason) are also included within this time frame.

The evaluation of the severity of events is based on Common Terminology Criteria for Adverse

Events (CTCAE) version 4.0 (see Appendix 15, page159). For the definition of what is

considered a serious adverse event see A19.1.4.

The treatment given within this trial has well known side effects. Therefore, for the purposes of

this trial only serious adverse events (SAE) will be reported. All SAEs, except those listed as

expected in section A19.3, that occur within this trial will require expedited (immediate)

reporting. Expedited is defined as within 24 hours of knowledge of the event.

A19.3 Expected SAEs which are exempt from expedited reporting

The following SAEs are consistent with the profile of expected toxicities in this trial and should

not be reported, unless in the opinion of the local investigator are unexpectedly severe.

Hospitalisation for febrile neutropenia without hemodynamic compromise

Hospitalisation for red blood cell or platelet transfusion

Hospitalisation for uncomplicated infection

Hospitalisation for uncomplicated mucositis or diarrhoea

Hospitalisation for uncomplicated vomiting

Hospitalisation for enteral or parenteral nutritional support

The following events are also exempt from expedited reporting:

Relapse

Signs and symptoms of disease progression

Death from disease progression

Secondary malignancy

Relapse, progression, secondary malignancy and death from disease progression must still be

reported via the appropriate forms.

A19.4 Reporting procedures and time limits

A19.4.1 Local investigator responsibilities

The local investigator must report all serious adverse events (SAEs), except those which are

expected, within 24 hours of knowledge of the event. SAEs should be reported via the SIOPEN-



R-NET using the SAE form. The SAE form appears for reference in Appendix 20, page 182. The

SIOPEN-R-NET has the ability to notify the Sponsor that a SAE has been reported. However,

notification can not be guaranteed and as such the local investigator is responsible for confirming

that the SAE report has been received by the Sponsor. If the SIOPEN-R-NET does not work,

SAEs should be reported by FAX (+34 349 44 16) or email ([email protected]). The

local investigator is responsible for providing an assessment of the seriousness, severity and

causality (A19.5) of the SAE. The SAE form should be completed as much as possible; the local

investigator should not wait for full details prior to making the initial report. The initial report

should be promptly followed up with the full details, so that the Sponsor can report the event to

the regulatory authorities within the required time periods (see Sponsor responsibilities for more

details).

For reported deaths, the local investigator should promptly supply the Sponsor and the IRB/IEC

with any additional requested information (e.g. autopsy reports and terminal record reports).

The local investigator must also comply with local requirements related to the reporting of SAEs

to the regulatory authorities and IRB/IEC. The local investigator is responsible for the

distribution of any information supplied by the Sponsor the appropriate regulatory authorities and

local IRB/IEC, as required by local laws.

The local investigator must ensure that there is a clearly documented audit trail of the reporting to

the Sponsor and local regulatory authorities.

A19.4.2 Sponsor responsibilities

The Sponsor is responsible for the ongoing safety evaluation of the trial. All SAEs received by

the Sponsor must be reviewed in a timely manner to ensure that the requirements for the onward

reporting can be completed in a timely manner. It is the responsibility of the Sponsor, with input

from the appropriate Principal Investigator if required, to assess the expectedness of a reported

SAE. That is, to classify, or not, a SUSAR.

In the case of a SUSAR it must be determined whether or not the event is fatal or life-threatening.

Reporting requirements are different for those events which are fatal or life-threatening to those

which are not.

SUSARs which are fatal or life-threatening qualify for rapid reporting. The appropriate

regulatory authorities and investigators must be notified within seven calendar days after first

knowledge by the Sponsor that the event qualifies as a SUSAR. A complete report must be

supplied within a further eight calendar days.

SUSARs which are not fatal or life-threatening must be reported to the appropriate regulatory

authorities and investigators within 15 calendar days after first knowledge by the Sponsor that the

event qualifies as a SUSAR.

SESARs must be reported together with the annual report to the IRB/IEC yearly.

The Sponsor is responsibly for notifying all concerned investigators and regulatory authorities of

findings that could adversely affect the safety of patients, impact the conduct of the trial, or alter

the IRB/IEC opinion to continue the approval of the trial.

If the SAE is considered a major cause of concern in relation to other, or future, patients on a trial,

it maybe necessary to issue an urgent safety notice to all investigators pending a formal



amendment. The urgent safety notice must be sent from the Sponsor and contain a clear

explanation of the event and any treatment recommendations.

The Sponsor must keep a clearly documented audit trail of receipt of SAEs, clinical review

conducted and outcome, and onward reporting.

A19.5 Causality assessment

Relationship Description

Definitely There is clear evidence to suggest a casual relationship and other

possible contributing factors can be ruled out.

Probable There is evidence to suggest a casual relationship and the influence

of other factors is unlikely.

Possible There is some evidence to suggest a causal relationship (e.g.

because the event occurs within a reasonable time after

administration of the trial medication). However, the influence of

other factors may have contributed to the event (e.g. the patient’s

clinical condition, other concomitant treatments).

Unlikely There is little evidence to suggest there is a casual relationship (e.g.

the event did not occur with a reasonable time after administration

of the trial medication). There is another reasonable explanation

for the event (e.g. the patient’s clinical condition, other concomitant

treatments).

Unrelated There is no evidence of any causal relationship.

Unknown/Not

Assessable

There is insufficient or incomplete evidence to make a clinical

judgement of the causal relationship.


Version 5.0; 1 September 2015 Page 182 of 237

APPENDIX 20. SERIOUS ADVERSE EVENT REPORT FORM

Patient UID: [ __ __ - __ __ __ __ - __ __ ] Date of birth: [ __ __ __ __ - __ __ - __ __ ] (yyyy-mm-dd)

Report information

Initial report

Follow-up report

Final report

Date of this report: [ __ __ __ __ - __ __ - __ __ ] (yyyy-mm-dd)

Full name and institution of investigator:

Patient information

Study group: 1 – observation arm 1 – chemotherapy arm 2 3 4 5 6 7 8 9 10

Treatment at time of SAE:

CO VP/Carbo CADO

Radiotherapy Isotretinoin Surgery

Observational period (study group 1 –

observation arm or study group 4)

Other chemotherapy

Date of last day of treatment prior to SAE: [ __ __ __ __ - __ __ - __ __ ] (yyyy-mm-dd)

If the patient is being observed only (study group – observation arm or study group 4) then this

date should be left blank.

Other chemotherapy specify:

Previous study treatment (including number of cycles):

Seriousness information

results in death

life-threatening

caused/prolonged inpatient hospitalisation

results in persistent of significant

disability/incapacity

congenital anomaly or birth defect

other medically significant condition

If SAE results in death, date of death:

[ __ __ __ __ - __ __ - __ __ ] (yyyy-mm-

dd)

Cause of death:

As written on the death certificate

Autopsy performed?

Yes No

If yes, attach autopsy report



Serious adverse event (SAE)

SAE Diagnosis Use concise medical terms

CTCAE grade (version 4.0) Maximum

grade

Onset date (yyyy-mm-dd)

Outcome Current status 1 = recovered 2 = recovering/resolving 3 = not recovered/resolved 4 = recovered/resolved with sequelae 5 = fatal 6 = unknown

End date (yyyy-mm-dd)

Possible explanation for SAE 1 = chemotherapy related 2 = concomitant medication 3 = pre-existing condition 4 = other explanation

SAE description (including relevant laboratory and clinical findings):

Treatment of SAE

None Drug treatment Other

Details of treatment:



Study Chemotherapy (CO, VP/Carbo, CADO)

Drug name

Date of first administration (yyyy-mm-dd)

Date of last administration prior to SAE (yyyy-mm-dd)

Total daily dose and unit at last administration prior to SAE

Total cumulative dose and unit

Causal relationship to SAE 1 = definitely 2 = probable 3 = possible 4 = unlikely 5 = unrelated 6 = unknown

Action taken 1 = dose not changed 2 = dose reduced 3 = dose increased 4 = drug withdrawn 5 = unknown

If drug withdrawn: Has drug been readministered? 1 = yes 2 = no 3 = unknown

If drug was readministered: Did reaction reoccur on readministration? 1 = yes 2 = no 3 = unknown

Other Study Treatment (radiotherapy, isotretinoin)



Relevant concomitant medication

Drug name

Do not enter drugs

used to treat the

SAE

Date of first

administration

(yyyy-mm-dd)

Indication Date of last

administration

prior to SAE

(yyyy-mm-dd)

Total daily

dose and unit

at last

administration

prior to SAE

Total

cumulative

dose and

unit

Causal

relationship to

SAE

1 = definitely

2 = probable

3 = possible

4 = unlikely

5 = unrelated

6 = unknown

Action taken

1 = dose not

changed

2 = dose reduced

3 = dose increased

4 = drug withdrawn

5 = unknown

If drug withdrawn:

Has drug been

readministered?

1 = yes

2 = no

3 = unknown

If drug was

readministered:

Did reaction

reoccur on

readministration?

1 = yes

2 = no

3 = unknown

Relevant medical history

Details of relevant conditions/diseases (including date of diagnosis and possible causal relationship to SAE):

Further comments



APPENDIX 21. PARENT AND PATIENT INFORMATION

A21.1 Registration into LINES for Low and Intermediate Risk Study Groups

A21.1.1 Information for parents

INFORMATION FOR PARENTS/LEGAL REPRESENTATIVES OF CHILDREN FOR

REGISTRATION IN THE EUROPEAN STUDY

LINES CLINICAL TRIAL

N° EUDRACT: 2010-021396-81

The doctors looking after your child have determined that your child probably has a type of

childhood cancer called neuroblastoma.

We would like to invite your child to take part in an international clinical research study, which

aims to maintain or improve the already excellent success rates in treated cases of non high risk

neuroblastoma while reducing the overall side effects related to treatment. The study is called the

European Low and Intermediate Risk Neuroblastoma Study and is organised by the European

Neuroblastoma Research Group (SIOPEN).

Neuroblastoma is a rare cancer of specialised nerve cells called neural crest cells. Neuroblastoma

can occur almost anywhere in the body, but most often it occurs in one of the adrenal glands in the

abdomen (these are 2 specialised glands above the kidneys that release hormones to maintain

blood pressure and enable us to respond to stress) or in nerve tissue that runs alongside the spinal

cord in the neck, chest, abdomen or pelvis (called the sympathetic nervous system). These

tumours can stay in one place (localised), or may spread around the body (metastasise) to other

organs, such as bones, bone marrow, lymph nodes, lung, nervous system and/or the liver.

The diagnosis of neuroblastoma needs to be confirmed by obtaining a sample of the tumour. This

will either be from a biopsy when a small amount of the tumour is removed or from a resection of

most or the entire tumour. Tests on the tumour tissue are undertaken both to confirm the diagnosis

and to get extra information on the tumour (called biological studies) which will help to determine

which regimen will be best for your child. Occasionally these tests can be done on a sample of

your child’s bone marrow if the neuroblastoma cells are in the bone marrow. These proposed

additional studies don’t require further invasive investigations and the samples are obtained as

part of the standard evaluation.

To help the doctors looking after your child decide on the best treatment and care, we would like

your permission to collect and analyse samples of your child’s tumour, blood and bone marrow.

Special tests need to be carried out on these samples. Some of these tests will be done in the hospital



where will your child is being looked after, while other tests will be done at specialised laboratories.

It is possible that specimens from your child will have to be sent to another country so that these

special tests can be done. The special tests that need to be carried out are

Examining the appearance (histology) of the tumour cells under the microscope

Determining if there is amplification (extra copies) of a tumour gene called MYCN

Looking for specific changes or abnormalities of the DNA or genetic material of the tumour

cells for example to see if part of a chromosome is missing

In addition to carrying out these special tests on your child’s tumour sample we would need to collect

information about your child that will make it possible for the doctors to decide the best treatment.

This information includes:

Date of birth

An identification of your child

Information about how extensive the tumour is

Information about how your child has been feeling (these are called symptoms)

Information on the results of the special tests (for example biological studies)

All information we collect from your child will be entered in a secure database owned by

SIOPEN; the data is encrypted and encoded. SIOPEN is a nonprofit legal entity that has been

devoted to the study of neuroblastoma in Europe for years; it consists of physicians and

researchers from different European countries. All reports will be made anonymous by using an

alphanumeric code and will be properly guarded. Reports will not be transmitted outside the

European Union's environment and will be kept confidential and will not be accessible or

available except to the physician (or persons under their supervision), health authorities or

individuals in which the sponsor has been delegated. All information (individual clinical and

biological data) collected will be treated in accordance with the European Directive of good

clinical practice (GCP) and National Law 15/99 including on access, rectification and cancellation

request as established by regulations and Law 14/2007 of biomedical research.

We request your consent to conduct tests and collect the minimum data set required to register your

child in the SIOPEN LINES trial.



A21.1.2 Informed consent for parents (all study groups)

CONSENT FORM FOR REGISTRATION IN THE LINES STUDY

Do you confirm that you have read this letter about registration into the LINES

study and the tests that need to be undertaken prior to consenting for your child to

be entered into the LINES STUDY?

YES/NO

Do you give your permission for samples from your child’s tumour to undergo the

special tests as detailed above?

YES/NO

Do you agree for the tumour samples to be stored for future research?

YES/NO

Do you agree for information on your child including the results of the biological

tests on the neuroblastoma sample to be stored in the SIOPEN database?

YES/NO

If you agree for your child to be registered into this study when all the above tests have been

completed your child’s doctor will talk to you about the best treatment option for your child and you

will be asked to sign another consent form.



CONSENT FORM FOR PARENTS/GUARDIAN

Clinical trial:

I…………………………….( Name of Patient)

• I confirm that I have read and understood the information contained in the information sheet.

• I have had the opportunity to ask questions and had satisfactory answers to them.

• I have received enough information about the study

I have talked to:

............................................................................................ (Investigator´s name).

I understand that my child’s participation is voluntary and that she/he is free to withdraw at any time

without giving any reason and without any negative consequence for his/her medical care

In my presence, (name of participant)------------------------has received all the information according

to his/her level of understanding and she/he agrees to participate

And I hereby approve the participation of my son/daughter. .....................( name of participant) in the

study.

I hereby approve the participation of (paticipant´s name)------------------------------------ in this trial.

Parent/Guardian signature Investigator´s signature

Name: Name:

Date: Date



A21.2 Low Risk Study

A21.2.1 Information for parents

INFORMATION FOR PARENTS/LEGAL REPRESENTATIVES OF CHILDREN

PARTICIPATING IN THE EUROPEAN STUDY « LINES » FOR LOW RISK

NEUROBLASTOMA

CLINICAL TRIAL

N° EUDRACT: 2010-021396-81

SPONSOR: Fundación para la Investigacion Hospital La Fe

INTRODUCTION

We are inviting your child to take part in a clinical trial looking at the treatment of low risk

neuroblastoma. A clinical trial is a research study involving treatment of a disease in human

patients. This study is organised by SIOPEN, the European neuroblastoma research group. Before

you decide what is best for your child it is important for you to understand why this research is

being done and what it will involve. Please take time to read the following information carefully

and discuss it with anyone you wish including members of your child’s medical team.

Your child has a disease called neuroblastoma. This is a rare cancer of specialised nerve cells

called neural crest cells. Neuroblastoma can occur almost anywhere in the body, but most often it

occurs in one of the adrenal glands in the abdomen (these are 2 specialised glands above the

kidneys that release hormones to maintain blood pressure and enable us to respond to stress) or in

nerve tissue that runs alongside the spinal cord in the neck, chest, abdomen or pelvis (called the

sympathetic nervous system). These tumours can stay in one place (localised), or may spread

around the body (metastasise) to other organs, such as bones, bone marrow, lymph nodes, lung,

nervous system and/or the liver. Your doctor will explain to you what the ‘stage’ of your child’s

tumour is, that is, whether the tumour has remained localised or whether it has spread to other

parts of the body. Your doctor will recommend a treatment course that will be based on the stage

of your child’s tumour, his/her age at the time of diagnosis, and some other special features of the

tumour, that are called ‘biological features’. Some biological features are determined by

examining the appearance (also called, histology) of the tumour under the microscope. Other

biological features are determined by doing certain tests on the tumour tissue in a research

laboratory. These tests include measuring the amount of DNA or genetic material present in the

tumour tissue (also called, ‘ploidy’) as well as looking for specific changes or abnormalities

(something that is not normal) of the DNA, such as when part of a chromosome is missing. All of

these tests are done on a sample of tumour tissue (biopsy) that the surgeon has already removed.

Once your doctor has collected the information about your child’s tumour stage and biological

features, he/she can determine what ‘risk’ category your child’s tumour falls into, and how much

chemotherapy will be needed to treat the neuroblastoma. The term ‘risk’ refers to the chance of

the cancer coming back after treatment; this is known as relapse or recurrence.

Your doctor has determined that your child’s neuroblastoma is considered to be of ‘low risk’,

based on all the above features and the fact that the tumor does not have amplification of the

MYCN gene.



There are two groups of patients with neuroblastoma that are classified as low risk.

1. The first group are patients whose age is below 18 months at diagnosis, who have a

localised tumour (i.e. it has not spread to other areas of the body) which cannot be

removed surgically without causing a risk to the patient.

2. The second group are patients less than 12 months of age with a tumour spread to other

organs known as stage Ms. This Ms stage is characterised by patients who usually have a

small primary lump of tumour with metastatic disease in the skin, liver and/or bone

marrow (but not in the lung, brain or bone). The liver can become extremely large with

deposits of neuroblastoma in it.

In both groups, it is possible that your son / daughter has some symptoms of illness that

may be mild.

1 – What is the purpose of this study?

In this study, tumour from all patients will be analysed using a relatively new technique (either

“array-CGH” or “MLPA”) which is able to analyse whether there are chromosomal abnormalities

in the tumour cells. The result of this test will be used to determine if certain low risk patients

should have more or less treatment (this has not been done before in a study group such as this).

This is because it has been shown that when there are certain (called segmental) chromosomal

abnormalities present in tumour cells there is a slight increased risk of relapse or tumour

progression during treatment.

Patients with these abnormalities in their tumor cells will not have their chemotherapy treatment

reduced and others (not treated with chemotherapy) will now be treated if they present with these

alterations. Therefore, your son / daughter will be included in a treatment group based on all

results obtained. The European researchers have clear objectives for each group of treatment,

which are summarized below:

AIMS OF THE STUDY

1. To optimise the treatment of low risk patients less than 18 months of age with unresectable

localised disease by organising the treatment into separate study groups depending on the

presence or not of life threatening symptoms and the presence or not of adverse biological

features in the tumour.

2. For one sub-group of unresectable localised disease patients, there will be the option of

being randomised to being observed rather than being treated with conventional

chemotherapy. These patients have localised unresectable disease with no life threatening

symptoms and no evidence of segmental chromosomal abnormalities.

3. To optimise the treatment of low risk patients less than 12 months of age with Stage Ms

disease by organising the treatment into separate study groups depending on the presence

or not of life threatening symptoms and the presence or not of adverse biological features

in the tumour.

2 – Why has my child been chosen?

All children in Europe with low risk disease according to the eligibility criteria of this study will

be invited to take part in this study. Over 5 years it is anticipated that there will be a total of 410

patients eligible from 20 participating countries.

It is up to you to decide whether or not your child should take part. Your child does not have to



participate in order to receive treatment for their neuroblastoma. If you do decide that you would

like your child to take part, you will be given this information sheet to keep and be asked to sign a

consent form. You are free to withdraw your child from the study at any time and without giving a

reason. This will not affect the standard of care your child will receive and treatment will be given

as per the relevant previous protocols. Your doctor may also withdraw your child from the study

if he/she considers that to be appropriate.

3 – What are the different steps of this study?

This study is divided into the following steps:

1. Inclusion in the study and necessary investigations (imaging examinations, blood and

urine, bone marrow study and obtaining tumor samples) undertaken including the

biological tests looking for segmental chromosomal changes in the tumour cells.

2. Allocation to a treatment arm when all results are available

3. Specific treatment of the patient within the allocated study group

4. The treatment could last for 2-6 months (this depends on the treatment group that your

child is assigned to) with observation and follow-up for 5 years

4 – What does the treatment involve?

After a consent form has been signed all patients will be allocated into a treatment study group

when information regarding their tumour is known. This will include standard information on the

tumour, age, stage, and the presence or not of symptoms and also the new biological test that

detects segmental chromosomal changes in the tumour cells. This is the first European Study to

stratify treatment for low risk patients according to the presence of these segmental chromosomal

changes. Your child’s doctor will tell you which study group your child will be treated according

to.

Information on the 6 study groups

Study group 1 Randomised Study

Patients < 18 months with unresectable localised disease and no amplification of the tumour gene

MYCN, with no life threatening symptoms and no evidence of segmental chromosomal

abnormalities.

Background to randomised study

The treatment of localised neuroblastoma historically has been based on a surgical removal of the

primary tumour. However if there are radiological signs indicating the presence of preoperative

risk factors a surgical removal is not recommended as it can be dangerous. In the past low dose

chemotherapy was given to these patients to induce tumour shrinkage in order to try and make a

surgical excision possible. We now know that this surgery may not be necessary in many of these

cases as it is clear that these tumours often regress spontaneously, without any treatment being

given. This has been shown to be the case in nearly half of all such cases in a study performed

recently in Germany.

At present it is not possible to know precisely which children do not require chemotherapy so we

are proposing that this group of patients will have the option of being randomised to being

observed rather than being treated with conventional chemotherapy. All of these patients will be

closely monitored and after some years of follow-up we may be able to demonstrate that most of

this group of patients do not need to be treated with initial chemotherapy.

Today, it is impossible to know precisely the children that require chemotherapy or not, so we

propose that in this group of patients treatment will be randomised between two possible



Strategies (observation compared to initial chemotherapy). This means that your son / daughter

has a 50% chance of receiving or not chemotherapy. This decision is "random".

Patients who are found to have segmental chromosome abnormalities in their tumour cells will not

be eligible for this randomisation as that biological profile may indicate that those patients do

need to be treated with chemotherapy rather than being observed.

In this study

If your child is randomised to the observation arm they will be regularly examined clinically with

appropriate radiological investigations every 8 weeks until the tumour starts to shrink when the

investigations will be performed 12 weekly for one year. The tumour will be removed 1 year

from diagnosis unless there are persisting radiological signs indicating that there is a risk to

surgical removal.

If the tumour significantly increases in size, or if any symptoms develop, then moderate dose

chemotherapy, will be given.

If your child is randomised to receive chemotherapy, they will have up to 6 courses of

chemotherapy to try and get the tumour small enough to be safe to surgically remove. The tumour

will be reassessed after each 2 courses of chemotherapy.

The surgical removal for both groups in Study Group 1 (observed patients and up front

chemotherapy) will not be performed if either the tumour has disappeared completely or if

radiologically defined risk factors are present, making a surgical removal difficult with a definite

risk of side effects (for instance to the kidney or the intestine).

In some patients it may be necessary to start chemotherapy before the study group is known. Your

doctor will discuss this with you.

Study Groups 2 and 3

Study Group 2 Patients < 18 months with unresectable localised disease and no amplification of

the tumour gene MYCN, with life threatening symptoms and no evidence of segmental

chromosomal abnormalities.

Study Group 3 Patients < 18 months with unresectable localised disease and no amplification of

the tumour gene MYCN, with or without life threatening symptoms and evidence of segmental

chromosomal abnormalities.

Your child will receive 2-4 courses of chemotherapy and a surgical removal if there are no

radiologically defined risk factors making surgery difficult.

Study Group 4

Patients < 12months with Stage Ms disease and no amplification of the tumour gene MYCN,

without life threatening symptoms and no evidence of segmental chromosomal abnormalities.

Your child will be observed only and there is no indication for the primary tumour to be removed.

Study Groups 5 and 6

Study Group 5 Patients < 12months with Stage Ms disease and no amplification of the tumour

gene MYCN, with life threatening symptoms and no evidence of segmental chromosomal

abnormalities.

Study Group 6 Patients < 12months with Stage Ms disease and no amplification of the tumour

gene MYCN, with or without life threatening symptoms and evidence of segmental chromosomal

abnormalities.



Your child will receive 2-4 courses of chemotherapy. A surgical resection of the primary tumour

is not indicated for Study group 5. A surgical removal may be performed for the primary tumour

in study group 6 if there are if there are no radiologically defined risk factors rendering surgery

difficult.

During and at the end of treatment, regular follow-up will be performed, in order to see if the

tumour has got smaller or disappeared, and is not growing back and that the symptoms associated

with it have resolved (if there were symptoms at diagnosis). If the tumour does get larger new

treatments might be necessary for your child. During off-treatment follow-up, clinical

examination by your child’s doctor will be regularly performed, as well as radiological follow-up

depending on the site of the primary tumour. This follow-up will take place at least over 5 years

after treatment and in general children are followed until young adult age. This pattern of clinical

follow-up is routine in all children with neuroblastoma and not linked specifically to this trial.

5 – What are the possible advantages of participating in this trial?

This study uses scientifically valid treatment stratification for low risk cases of neuroblastoma.

Your child will benefit from having new molecular tests undertaken on their tumour cells to show

more or less favourable features that will then be used to determine treatment.

This trial proposes to adjust the intensity of the treatment your son/daughter receives according to

his/her disease biological aggressiveness. It is possible that the participation of your son/daughter

in the study may not provide any health benefit.

6 – What are the possible risks?

6.1. Chemotherapy

All the recommended drugs have been given to children with neuroblastoma for many years.

Chemotherapy causes temporary hair loss (rarely this can be permanent) and a reduction in the

number of blood cells which increases the risk of infection. Transfusions and antibiotics may be

required. If there is nausea and vomiting this will be treated with anti-sickness medications.

Occasionally there can be an allergic reaction to a drug and this would need treatment with anti-

allergy medications. Specific side effects are listed below. If your child’s doctor believes the

toxicity of treatment is too great or is not having enough effect on your child’s illness the

treatment will be modified and alternative treatments will be discussed with you.

For the administration of chemotherapy, blood sampling, and for administration of antibiotics or

other medications, it is often necessary to place a catheter in a big vein in the chest, called a

central venous catheter. This device can be used to administer intravenous (into the blood stream)

chemotherapy and take necessary blood samples without any discomfort to your child, but some

hygiene rules need to be followed in order to reduce the risk of infection.

Specific drug related side effects:

Vincristine has very little effect on the reduction in the number of blood cells. However other

well known side effects are abdominal pain, constipation, cramp like pain in the arms or legs

or in the lower part of the jaw, or very rarely an alteration of the sodium blood level which can

also lead to an altered level of conscience.

Cyclophosphamide may lead to a reduction in the number of red cells, white cells and platelets

in the blood which might mean that your child would need a blood or a platelet transfusion

and could be at an increased risk of getting an infection.



Etoposide leads to a fall of blood cells with an increased risk of infection and also a theoretical

risk of secondary leukaemia (even if the prescribed doses are lower than the doses known to

induce leukaemia).

Carboplatin also leads to a fall of blood cells, especially white blood cells and platelets.

Following administration of this drug often red blood cell or platelet transfusions are required.

It can also lead to kidney damage or hearing loss, and your child will be monitored for these

side effects should this drug be given.

6.2. The risk of observation as upfront treatment in the randomised arm of study group 1

Primary observation, without any upfront chemotherapy, exposes the child to a risk of local

disease progression, or distant disease progression. The latter is rare, and taking into account the

favourable biological factors of the tumour of your child the risk of distant disease progression is

certainly below 4%. The regular clinical and radiological follow-up will enable a progression to

be detected early should it occur, and then appropriate treatment can be started.

6.3. Surgery

Surgery is the mainstay of treatment in localised neuroblastoma, but it should only be performed

in certain conditions in order to avoid complications or postoperative sequelae. The European

group has defined radiological signs which aim to predict preoperative complications. When these

signs are present surgery to the primary tumour should not be performed. The surgeons know

about these risk signs and will take them into account prior to making a decision about surgery at

the appropriate time point in the protocol.

6.4. In any study, there are no foreseeable risks. Monitoring procedures of appropriate risks are

considered. Unexpected complications will be monitored.

7 – What are the alternatives? The alternative to inclusion in the clinical trial is for your child to have treatment according to the

standard treatment recommendations based on previous clinical trials. Your child’s doctor will

talk to you about these if you wish.

8 – In case of participation in this trial.

This study was designed by a group of European medical experts in Neuroblastoma (SIOPEN),

who since 1994 are conducting cooperative research in this disease. Research is done according to

the principles of the Declaration of Helsinki and Good Clinical Practice standards.

Taking part in this study is voluntary. You may choose to not allow your child to take part or may

remove your child from the study at any time. Deciding not to be in the study or leaving the study

will not result in any penalty or loss of benefits to which your child is entitled.

The researchers did not receive any compensation for their participation in the study

The national sponsor of this trial, ……………………………………, has taken all the precautions

provided by the law for the protection of patients and has subscribed to an insurance according to

the …..

This trial has been submitted for investigation and approval by the ethics committee

……………………..… with an aim to verify that all conditions necessary for the respect of your

child’s rights are maintained. The ethics committee has given approval for this trial. At the end of

this research you will be informed of the results, should you wish. Researchers undertake to



inform you of new relevant data that may appear throughout the study and influence their decision

to stay in it.

All information (personal, clinical and research data from the biological material) is treated in

accordance with Directive 95/46/EC of the European Parliament and the Council of 24 October

1995 on the protection of individuals with regard to the processing of personal data, and Law

15/1999 of December 13, Protection of personal Data and its implementing regulations, among

which is the Royal Decree 994 / 1999 of 11 June, that approves the Regulations on security

measures for automated files containing data of this character. Only authorised researchers,

national authorities, and ethics committees will have access to such data, in accordance with the

normative state.

The researcher will enter the data in a secure database owned by SIOPEN; the data is encrypted

and encoded. SIOPEN is a nonprofit legal entity that has been dedicated to the study of

neuroblastoma in Europe for years; it consits of physicians and researchers from different

European countries and is located in Vienna (Austria). Data will not be transmitted outside the

European Union. There is a national center coordinator (H. La Fe) responsible for developing the

study in Spain and a center of biological studies (F. de Medicina, U. Valencia) where biological

samples are stored and processed in accordance with the Biomedical Research Law (14/2007).

The reports of the study and other information will be referenced with an alphanumeric code, and

your name and patient’ identity will never be disclosed to anyone except to fulfill the purposes of

the study, and in case of a medical emergency or legal requirement.

You are entitled to access the data, rectify, cancel and object to them at any time by notifying the

study physician. Similarly, you have the right to request the destruction of stored samples for

biological studies, if desired, at any time; you should talk to the physician of the study about this.

9 – Whom should you contact in case of questions or problems?

For questions about this study or a research related problem contact

DR……………………………………………………....

Telephone contact ……………………….………

who will reply to all the questions that you wish to ask. This can be either prior to your decision to

consent to participation in this trial, or after your child has been enrolled in the study.

If during the study you wish to ask for a second opinion or to consult another physician, you

should indicate that your child is participating in this trial so that he/she can contact the trial

organisers should this be necessary.

10. Therefore, we ask you to give your consent to:

- The provision of the information from your child's medical notes confidentially to the National

Coordinator(H. La Fe) and the SIOPEN data base for analysis.

-The collection, storage and supply of biological and clinical information to the national registry

(H. La Fe).

-Inform the medical team of your son / daughter participation in this clinical trial.

-To participate in the study LINES and be treated according to treatment group No. ... ....



A21.2.2 Informed consent for parents (all low risk study groups with a genomic profile)

CONSENT FORM FOR THE SIOPEN LINES LOW RISK STUDY

Patient with a genomic profile result or patient requiring chemotherapy prior genomic

profile result is known.

EudraCT number:2010-021396-81

Clinical trial:


• I confirm that I have read and understood the information contained in the information

sheet.



I have talked to:

............................................................................................ (Investigator name).

I understand that my child’s participation is voluntary and that she/he is free to withdraw at any

time without giving any reason and without any negative consequence for his/her medical care

In my presence, (name of participant)------------------------has received all the information

according to his/her level of understanding and she/he agrees to participate

And I hereby approve the participation of my son/daughter. .....................( name of participant) in

the study.

-I agree that the blood or tissue samples obtained for the study will be used in the future for

further analysis related to the disease, unforeseen in the current protocol (excluding genetic tests,

provided that they are not part of the objectives the study):

Yes� No�

I hereby approve the participation of (paticipant´s name)------------------------------------ in this

trial.


Name: Name:

Date: Date



A21.2.3 Informed consent for parents (all low risk study groups without a genomic profile)

CONSENT FORM FOR THE SIOPEN LINES LOW RISK STUDY

Patient without a genomic profile result

Patients without a result of their genomic profile.

I understand that because there is no result on the genomic profile of my son / daughter, he / she

is not eligible to participate in this clinical trial. However, he / she will be registered in the

clinical trial database and the information concerning the treatment received (GROUP:) and how

he / she responds to treatment will be collected and sent in a confidential manner to the SIOPEN

database

Clinical trial:



sheet.



I have talked to:







trial.

EudraCT number:: 2010-021396-81


Name: Name:

Date: Date



A21.2.4 Informed consent for parents (additional for study group 1)

CONSENT FORM FOR THE SIOPEN LINES LOW RISK STUDY GROUP 1

Randomisation for Low Risk localised patients with no biological risk factors in

Study Group 1 between

1. Treatment with chemotherapy followed by surgery or

2. Observation only until spontaneous regression preceding possible surgery

Do you agree to your child taking part in the trial randomisation as

explained in the information sheet?

YES/NO

Name of Patient ………………………………………………………………..

Name of Parent/Guardian ……………………………………………………...

Signed ………………………………………………………………………….

Date …………………………………………………………………………….

Name of Investigator ………………………………………………………………..

Signed ………………………………………………………………………….

Date …………………………………………………………………………….

EudraCT number: 2010-021396-81



A21.3 Intermediate risk study

A21.3.1 Information for Parents

INFORMATION FOR PARENTS / LEGAL REPRESENTATIVE OF THE CHILD

PARTICIPATING IN THE EUROPEAN STUDY “LINES” FOR INTERMEDIATE RISK

NEUROBLASTOMA

N° EUDRACT : 2010-021396-81

SPONSOR: Fundación para la Investigacion Hospital La Fe

Introduction

Your child suffers from a disease called neuroblastoma. It is a tumour originating in the adrenal

gland or in the ganglia of the paravertebral sympathetic nervous system located in the neck,

thorax, abdomen or pelvis. According to his/her age, the extent of the tumour and the biological

features related to the tumour cells, a child with neuroblastoma has different chances of cure,

defined in our studies as the risk of relapse (or not cure), varying from low to intermediate or

high. In your child, according to previous European studies and to other international experiences,

your child’s risk is intermediate.

Therefore, in this study we treat patients with Intermediate Risk Neuroblastoma. This is a broad

category that includes different groups of patients and your child is included in one of them:

1st group: Group LINES 7

- Age older than 18 months

- Absence of metastases

- Unresectable tumour since the surgical resection could seriously damage organs and blood

vessels and nerves next to the tumour.

- Biology of the tumour showing an absence of “bad” genetic factors.

- Favourable histology: differentiating neuroblastoma or differentiating


In the previous European study (EUNS) all these children were cured with 6 courses of

chemotherapy followed by surgery. Analysis of the results shows that 6 chemotherapy courses

were too many for this type of histology. Therefore in this new study we will administer to your

child only 4 courses of chemotherapy (instead of the 6 as previously given), followed by a

surgical excision of the tumour. The aim of the study is to decrease the volume of the tumour and,

consequently, to make it resectable by treating with only 4 courses of chemotherapy. By using this

approach (therapy reduction) we think we will achieve the same rate of cure with less toxicity.

2nd group: Group LINES 8

- Age older than 18 months


- Unresectable tumour since the surgical resection could seriously damage organs and blood

vessels and nerves next to the tumour.

- Biology of the tumour showing an absence of “bad” genetic factors.

- Unfavourable histology: poorly differentiated or undifferentiated neuroblastoma OR

poorly differentiated or undifferentiated ganglioneuroblastoma nodular.

In the previous European study (EUNS) all these children were treated with 6 courses of

chemotherapy followed by surgery. The results we obtained were not encouraging because only

60% of children were definitely cured. In fact, some of these children who initially responded to

treatment presented thereafter with disease progression or relapse. The relapse in some cases



involved only the site of the primary tumour while in other cases there was a reappearance of

disease in distant sites (metastases). For this reason, in order to increase the cure rate, in this

study we will administer 6 courses of chemotherapy, surgery, a course of radiotherapy to the site

of the resected tumour followed by a course of differentiating treatment with 6 courses of 13-cis-

retinoic acid (13-cis-RA).

Radiotherapy is a modality of treatment effective in some tumours, included neuroblastoma,

directly addressed to the site of the disease, in order to increase the effect of chemotherapy and

minimise the risk of a recurrence of the tumour in that local area. You will be asked to sign a

separate consent form if your child requires radiotherapy.

13-cis-RA is a drug that comes from vitamin A and has been proved to be effective in patients

with metastatic neuroblastoma, who at the end of treatment have been left with a small amount of

malignant cells in the body. The activity of 13-cis-RA against neuroblastoma cells is characterised

by 2 effects, differentiation of tumour cells (e.g. maturation to a benign status) and apoptosis (e.g.

programmed death of the cells). It would be given to your child to try and prevent the

neuroblastoma recurring.

3rd group: Group LINES 9 - Any age


- Tumour completely resected

- Biology of the tumour cells characterised by amplification of MYCN oncogene.

Every gene is normally present in a cell in 2 copies. Amplification of a gene means that the copies

of the gene in a single cell are more than 2. The amplification of MYCN gene in neuroblastoma

cells makes the tumour more aggressive. In our previous study we treated these children with

surgery alone, but an extended international study has demonstrated that the prognosis of

neuroblastomas with amplification of MYCN gene, treated only with surgery is poor. For this

reason in this study we treat these cases with chemotherapy, radiotherapy and 13-cis-RA in order

to significantly increase the rate of cure.

4th group: Group LINES 10 - Age less than 12 months

- Presence of metastases in particular sites like bones, lungs or pleura and central nervous

system.

- Biology of the tumour showing an absence of “bad” genetic factors

For patients with these characteristics our European working group for neuroblastoma conducted

a clinical study in the years 1999-2004 obtaining a cure rate superior to 95%. In this study we will

use the same therapeutic schedule consisting of giving chemotherapy at a standard dose, resection

of the primary tumour without administering either intensive chemotherapy with autologous stem

cell transplantation, or radiotherapy.

Aims of this study

1st group (Group LINES 7). Patients with favourable histology localised neuroblastoma. The

aim of the study is to obtain the same successful previous rate of cure (100%) with less

chemotherapy than previously given (4 courses rather than 6) and, consequently, less drug related

toxicity.

2nd group (Group LINES 8). Patients with unfavourable histology localised neuroblastoma. The

aim of the study is to increase the previous rate of cure (60%) with a more aggressive treatment.



The addition of radiotherapy and 13-cis-RA should improve the rate of local and distant

progression/ relapse of disease.

3rd group (Group LINES 9). Patients with operated and resected (stage 1), MYCN amplified

neuroblastoma. The aim is to increase the rate of cure in this group of patients by adding an

adjuvant (post-surgery) treatment to the previous treatment which was based on surgery alone.

The treatment, consisting of chemotherapy, radiotherapy and 13-cis-RA, should improve the rate

of local and distant relapse of disease.

4th group (Group LINES 10). Infants (age less than 12 months) with metastatic neuroblastoma.

The aim is to obtain the same successful cure rate (>95%) obtained with our previous clinical

trial.

Side effects of the drugs utilised in this study

- Vincristine. If extravasation occurs risk of local necrosis. Alopecia. Abdominal pain,

constipation, arms and legs pain, jaw pain. Serious urinary retention followed by conscience

alteration is very rare.

- Cyclophosphamide. Slight reduction of blood cell count. Nausea and vomiting. Alopecia.

- Etoposide. Alopecia, headache, fever, hypotension, nausea, vomiting, anaphylactic reactions.

Reduction of blood cell count with increased risk of infections and need of transfusions of red

cells and platelets. This drug administered at cumulative doses higher than in this protocol, has

been described to cause secondary leukaemia.

- Carboplatin. Renal toxicity and ototoxicity. Nausea and vomiting.

- Doxorubicin. Reduction of blood cell count with increased risk of infections and need of

transfusions of red cells and platelets. If extravasation occurs risk of local necrosis.

Cardiotoxicity. Mucosal ulceration, nausea and vomiting, alopecia.

- 13-cis-RA. Dry skin and mucosa, cheilitis

Number of patients in this clinical study

A total of 200 patients from many European countries will be recruited in this study in a 5-year

time

Local treatment

- Groups LINES 7 and 8 . After chemotherapy (2-4-6 courses) an evaluation of the tumour

will be done by computed tomography or MRI. It will be important to define whether the

tumour could be operated or not without evident surgical risk. In the absence of risks a

surgical resection will be done. If a complete resection is considered to still be difficult and

hazardous as at the time of diagnosis, the surgeon will discuss with other experts in this field

to consider an incomplete excision. In cases with unfavourable histology the local treatment

will also include radiotherapy.

- Group LINES 9 . Surgical excision in these patients has been obtained at diagnosis. After the

last course of chemotherapy radiotherapy will be administered.

- Group LINES 10 . After chemotherapy (4-8 courses) an evaluation of the tumour will be

done by computed tomography or MRI. A decision will made on the whether the tumour can



be surgically resected without risk. In the absence of risks a surgical resection will be done. If

otherwise, a complete resection will be judged not feasible, and the child will not be operated

on.

- Follow up

During and after the end of treatment your child will be regularly followed up in order to control

the response to the treatment, the achievement of a complete remission, e.g. absence of tumour in

any site, the inactivity of the residual disease, if any, or the onset of relapse or progression of the

disease. This clinical and radiological follow up will continue for at least 5 years from diagnosis.

Duration of treatment

Four to 12 months depending on the group of patients and the response to treatment.

What are the alternatives?

The alternative to inclusion in the clinical trial is for your child to have treatment according to the

standard treatment recommendations based on previous clinical trials. Your child’s doctor will

talk to you about these if you wish.

What are your rights in case of participation in this trial?


who since 1994 are conducting cooperative research in this disease. Research is according to the

principles of the Declaration of Helsinki and Good Clinical Practice standards.

Taking part in this study is voluntary. You may choose to not allow your child to take part or may

remove your child from the study at any time. Deciding not to be in the study or leaving the study

will not result in any penalty or loss of benefits to which your child is entitled.

The researchers did not receive any compensation for their participation in the study

The national sponsor of this trial, ……………………………………, has taken all the precautions

provided by the law for the protection of patients and has subscribed to an insurance according to

the …..

This trial has been submitted for investigation and approval by the ethics committee

……………………..… with an aim to verify that all conditions necessary for the respect of your

child’s rights are maintained. The ethics committee has given approval for this trial. At the end of

this research you will be informed of the results, should you wish. Researchers undertake to

inform you of new relevant data that may appear throughout the study and influence their

decision to stay in it.

All information (personal, clinical and research data from the biological material) ) is treated in

accordance with Directive 95/46/EC of the European Parliament and and the Council of 24

October 1995 on the protection of individuals with regard to the processing of personal data, and

Law 15/1999 of December 13, Protection of personal Data and its implementing regulations,



among which is the Royal Decree 994 / 1999 of 11 June, that approves the Regulations on

security measures for automated files containing data of this character. Only authorised

researchers, national authorities and ethics committees will have access to such data, in

accordance with the normative stated.


and encoded. SIOPEN is a non-profit legal entity that has been dedicated to the study of



European Union. There is a national center coordinator (H. La Fe) responsible for developing the

study in Spain and a center of biological studies (F. de Medicina, U. Valencia) where biological

samples arew stored and processed in accordance with the Biomedical Research Law (14/2007).

The reports of the study and other information will be referenced by an alphanumeric code, and






Whom should you contact in case of questions or problems?

For questions about this study or a research related problem contact

DR……………………………………………………....

Telephone contact ……………………….………

who will reply to all the questions that you wish to ask. This can be either prior to your decision to

consent to participation in this trial, or after your child has been enrolled in the study.



A21.3.2 Informed consent for parents (all IR study groups)

CONSENT FORM FOR THE SIOPEN LINES INTERMEDIATE RISK STUDY

Clinical trial:



sheet.



I have talked to:







the study.




Yes� No�


trial.



Name: Name:

Date: Date



A21.3.3 Informed consent for parents (IR patients without a defined histotype)

CONSENT FORM FOR THE SIOPEN LINES INTERMEDIATE RISK STUDY

L2 Patient without a defined histotype

Patients without a defined histotype.

I understand that because there is not a defined histotype for my son / daughter, he / she is not

eligible to participate in this clinical trial. However, he / she will be registered in the clinical trial

database and the information concerning the treatment received (GROUP: ) and how he / she

responds to treatment will be collected and sent in a confidential manner to the SIOPEN database.

Clinical trial:



sheet.



I have talked to:







the study.




Yes� No�


trial.



Name: Name:

Date: Date



A21.3.4 Information for patients over 6 years

INFORMATION FOR PATIENTS OVER 6 YEARS PARTICIPATING IN THE REGISTRY

STUDY AND FOR THE INTERMEDIATE RISK (GROUPS 7, 8 AND 9).

Nºeudract:2010-021396-81

EUROPEAN STUDY “LINES” FOR INTERMEDIATE AND LOW RISK OF

NEUROBLASTOMA

SPONSOR: FUNDACION PARA LA INVESTIGACION HOSPITAL UNIVERSITARIO

LA FE

Assent for the registration in the LINES study for intermediate risk groups, groups 7, 8 and

9

Doctors have seen that you have a lump in your body that may be a disease called neuroblastoma.

We would like to invite you to participate in an international clinical trial that intends to improve

treatment for children like you that have a neuroblastoma that is not of high risk (meaning that it

is not very bad). The trial is called the European Neuroblastoma Study for low and intermediate

risk and it is organized by the European Neuroblastoma Research Group (SIOPEN).

Neuroblastoma is a rare cancer of special nerve cells and can appear in many parts of the body but

most often in the abdomen (belly) or chest (trunk). It can stay in the same location or spread to

other locations (also known as metastasis), in your case it is only in one place

In order to find out the treatment that we have to give you, we need to take a piece of it and

analyze it. Sometimes, if it is possible the entire tumor is removed, but often it cannot be removed

so we take a piece. To remove this piece we perform a surgical procedure called a biopsy. This

piece will be analyzed by several different tests and we will know if it is a neuroblastoma and

which type (good, fair or bad). In addition we will make further tests to see if it is somewhere

else: we will take a little bit of bone marrow (the stuff inside your bones) and we will make



imaging examinations, including MIBG. For all this, we will ask for your cooperation (for

example, you will have to take a few drops of iodine) and we will always try to make it as easy

and pain free as possible. If necessary, we will give you some medicine to help you be more

relaxed or asleep.

Your doctor and some more people who work with him/her will know all this information but it will

always be kept confidential. The information will also be entered into a secure SIOPEN database, but

you be given a special number so no one will know who you are. This means that no one will be able

to know that you are the sick, but doctors will be able to learn more about the disease you have to

continue curing more children like you in the future. If at any time, you do not want your data to be

use, you can tell us and we will not use it.

You know that you can ask whatever you want and your doctor ... ... ... ... ... ... ... ... ... ... ... ... ..

will answer all your questions and doubts.

You know that you can ask whatever you want and your doctor ... ... ... ... ... ... ... ... ... ... ... ... ..

he/she will answer to all your questions and doubts.

So, we ask you to give us your consent (you agree) to:

Register in the LINESstudy

Perform the necessary tests to study your lump

Keep your piece of the tumor to study in accordance with the rules of trial

Save the data in the SIOPENdatabase

Receive treatment depending on the tumour you have:

Group 7: tumour in only one site, it is adviced to give you medications to make the tumour smaller

and depending on the response to remove it if possible

Group 8: tumour in only one site, from which only one piece has been analysed and can behave

regularly-badly: drugs to make the tumor smaller, then remove it if possible, then take a special

vitamin and radiotherapy. Treatment can take six to nine months.

or Group 9: tumour that has been completely removed and it is a bad neuroblastoma: it is adviced to

give you medications to prevent cells that we do not see but they can still grow in the future and

make the tumour reappear. We will also give you radiatherapy in the area and some special vitamins.

Treatment can last between 6 and 9 months.

PATIENT´S NAME: PHYSICIAN:

SIGNATURE SIGNATURE



A21.4 Sample of consent form for radiotherapy

CONSENT FOR RADIOTHERAPY EUDRACT NUMBER : 2010-021396-81

Radiotherapy for neuroblastoma

We give radiation treatment (radiotherapy) in addition to surgery and chemotherapy because the

disease may come back if we do not. The treatment has to be planned with great care .

The total radiation dose has to be spread out over quite a long time to make the individual

treatments safe. We give treatment every day for about 3 to 4 weeks (usually excluding weekends

and bank holidays). At each session, the patient spends about ten minutes in the treatment room

but most of this time is spent getting him/her into the right position.

Radiotherapy does not hurt, the machine does not touch the patient and it is rather like having an

ordinary x-ray. We have to treat the whole area accurately, which means that the patient must lie

very still. We decide whether the patient lies on their front or their back depending on the best

way of giving the treatment. To make lying still easier, we often surround the abdomen and chest

with a bag of polystyrene beads. Lying still can be very difficult for some children, particularly

young ones, so occasionally we have to use a general anaesthetic.

Side effects

There are always side effects. We can relieve most of them but sometimes not completely. Some

of the side effects happen immediately, some of them happen soon after we finish treatment and

some take a longer time to show.

Immediate side effects

• Nausea and vomiting may occur with this treatment but these can usually be well

controlled with anti-sickness drugs.

• Mild diarrhoea may occur but is unusual.

Side effects after treatment

• A course of radiotherapy often makes patients tired. This is at its worst near the end of the

treatment and just afterwards.

Long-term side effects

• These are the hardest to predict and unfortunately, when they do happen, they are

permanent.

• The radiation may affect the growth of the vertebral bodies (back bones) and ribs in the

radiation field. As only a small part of the spine is treated it is unlikely to have a significant effect

on final height. However because only one side of the tummy is treated one loin can look thinner

than the other and the waist may be narrow.

• The testicles are usually well away from the treated area and in most cases will be

unaffected by the radiotherapy. The ovaries can be close to or occasionally within the treatment



area and, if they are, fertility may be affected. This will need to be discussed on an individual

basis.

• Both radiotherapy and chemotherapy may affect the kidney. One kidney is usually (but not

always) in the area we need to treat and will receive a dose that in most cases will stop it working.

It may take 1-2 years after treatment finishes for the kidney to fail but once it does it will not work

again. We are careful to minimize the risk to the other kidney. Provided that the other kidney

continues to work normally there should not be any symptoms. The function of the kidney and

blood pressure will be carefully monitored at follow-up as problems can develop many years after

treatment.

• The radiation dose we give is low and we would not expect any long-term bowel

problems. However sometimes after surgery and radiotherapy the tissue surrounding the bowel

can stick together (adhesions) and this can sometimes cause symptoms requiring surgery.

• We are finding that, very rarely, children who have had treatment for one tumour can

develop another type of tumour some years later. Smoking significantly increases the risks of

developing many tumours.

During follow-up we will be looking out for all of these problems.

What are the alternatives to this treatment?

Your son / daughter is participating in the LINES trial, intermediate-risk group (group 8 or group

9). At this point, he/she will receive radiation therapy. The alternative to such treatment is to be

treated in accordance with standard recommendations based on previous trials (do not administer

radiation therapy to these two groups of patients). Your doctor will inform you in detail about this

if you desire.

What are your rights in case of participation in the trial?


which since 1994 has been conducting cooperative research in this disease. Research is done

according to the principles of the Declaration of Helsinki and Good Clinical Practice standards.

Taking part in this trial is voluntary. You can choose not to include your son / daughter in the

study or withdraw at any time if you want, without any explanation. You can decide not to be in

the study or leave it, and this will not be a prejudice to the rights your child has. You will not

receive financial remuneration for participating and you will not have to pay for the treatments

used in the study.

The researchers do not receive any compensation for their participation in the study.

The national sponsor of the trial has taken all measures and precautions that dictates the law for

the protection of patients and has signed an insurance policy with the company

……………………………………………………policy number as…………………………..

The study was approved by the Ethics Committees of participating centers. These Ethics

Committees have verified the conditions necessary to respect and fulfill your child’s rights. This

Committee has approved the trial. At the end of the research, we will inform the authorities, ethics

committees and you, if desired about the results. In addtion, the results will be communicated to

the scientific community in the form of publication. Researchers undertake to inform you of new

relevant data that may appear throughout the study and influence their decision to stay in it.




and encoded. SIOPEN is a non-profit legal entity that has been dedicated to the study of



European Union.

There is a national center coordinator (H. La Fe) responsible for developing the study in Spain

and a center of biological studies (F. de Medicina, U. Valencia) where biological samples arew

stored and processed in accordance with the Biomedical Research Law (14/2007).

The reports of the study and other information will be referenced by an alphanumeric code, and








A21.5 Neonatal suprarenal masses: an observational approach

A21.5.1 Information sheet for parents

INFORMATION SHEET FOR AN OBSERVATIONAL APPROACH IN

INFANTS WITH SUPRARENAL MASSES

NAME PRINCIPAL INVESTIGATOR:

CONTACT PHONE:

EMAIL:

CENTRE:

We are inviting your child to take part in a clinical trial looking at the treatment of suprarenal

masses in infants. A clinical trial is a research study involving treatment of a disease in human

patients. This study is organised by SIOPEN, the European neuroblastoma research group. Before

you decide what is best for your child it is important for you to understand why this research is

being done and what it will involve. Please take time to read the following information carefully

and discuss it with anyone you wish including members of your child’s medical team. This kind

of study includes only patients who are willing to participate.

WHY ARE WE PERFORMING THIS STUDY?

We would like to include your baby in this study since he/she has a small solid or cystic tumour or

mass in the adrenal gland. Your baby’s exhaustive evaluation has not revealed any other

abnormalities. It is very likely that this tumour will be a neuroblastoma, although it may also

represent a lung malformation (known as pulmonary sequestration) or an adrenal haemorrhage or

other abnormality. Neuroblastoma is a rare cancer of specialised nerve cells called neural crest

cells. Neuroblastoma can occur almost anywhere in the body, but most often it occurs in one of

the adrenal glands in the abdomen (these are 2 specialised glands above the kidneys that release

hormones to maintain blood pressure and enable us to respond to stress) or in nerve tissue that

runs alongside the spinal cord in the neck, chest, abdomen or pelvis (called the sympathetic

nervous system). These tumours can stay in one place (localised), or may spread around the body

(metastasise) to other organs, such as bones, bone marrow, lymph nodes, lung, nervous system

and/or the liver. The most common treatment for neuroblastoma is to perform a surgical operation

to remove it.

In the past we have learned a lot about neuroblastoma. One of the most important features is that

it behaves very differently in infants or in older children. Recent studies suggest that most (33-

100%) of these small tumours in infants’ adrenal gland disappear spontaneously, without any

treatment. This benign “behaviour” (including the possibility of total disappearance) makes us

willing to investigate whether we can avoid surgery. We will monitor them closely by ultrasound

and urine analysis in order to follow the tumoural activity and evolution. Abdominal surgery in

small babies has a 10% rate of complications and a 2% mortality rate. A therapeutic approach that

diminishes the frequency of surgery in these babies without jeopardising survival will be an

important improvement. Moreover, the adrenal gland cannot regenerate. Therefore, the fact of

excising one of the two human adrenal glands increases the risk of suffering an adrenal

insufficiency (if this condition happens, it requires a permanent hormonal supplementation) if the

non-excised gland fails during adulthood.



Therefore, this study is designed to answer the question: “IS IT SAFE TO AVOID SURGICAL

EXCISION OF A SMALL ADRENAL TUMOUR IN AN INFANT, BY EXHAUSTIVE

MONITORING THE TUMORAL BEHAVIOUR?”

If you agree in your child’s participation in the study, he/she will not have surgery initially. We

will watch closely by urine analysis, ultrasounds and other imaging tests. If there is an important

growth of the tumour or a change in its functioning, detected by the urine analysis, we will

recommend surgery. If the tumour stays stable or decreases, we will keep on watching until the

end of the observation period (12 months).

In the case we recommend surgery due to tumoural growth or changes in function, you should

know that neuroblastomas in infants behave very favourably. In spite of its growth, it is possible

that surgery will be the only treatment your child needs. It is very unusual and infrequent that in

infants, neuroblastoma will extend to other parts of the body. Infants need chemotherapy or

radiotherapy very seldom, since most infant neuroblastomas disappear with no treatments. If your

child has surgery, we will perform the biological analysis mandatory in neuroblastoma; and, if it

is shown that it is an aggressive tumour (very unusual in infants), your child may need treatment

with chemo and radiotherapy.

HOW MANY PATIENTS WILL PARTICIPATE IN THE STUDY?

The number of patients anticipated to be recruited to this observational approach over 5 year is

100 in the participating European countries. This number has been calculated using data from the

former European Infant Neuroblastoma European Study (INES) database. We don’t know the real

incidence of suprarenal masses in babies since nobody has conducted such a study in Europe. We

intend to maintain a survival of 95%, similar to the INES study.

WHAT DOES IT MEAN FOR YOUR CHILD TO PARTICIPATE IN THE STUDY?

It means that we will perform different tests on your child over the period of a year.

These tests are the following:

Physical examination.

Blood test.

Urine tests.

Abdominal ultrasounds.

MRI at week 6 from diagnosis, week 18 and week 42.

MIBG, which is a nuclear medicine test: we will inject a radioactive product with high

affinity for neuroblastoma. It will be done in the first 6 weeks of life. Your baby might be

sedated for the test and will have iodine given by mouth, which will help protect the

thyroid gland from future hypothyroidism. This test is appropriate to confirm that this is a

neuroblastoma and to know for sure whether it is located ONLY in the adrenal gland and

not in other organs.

Urine analysis and ultrasound will be repeated every 3 weeks according to the following table:

Follow-up (weeks) 0 3 6 9 12 18 30 48

Ultrasound + + + + + + + +

Catecholamines + + + + + + + +

MRI(*) + +

MIBG (*)



After week 12, if a tumour is regressing (i.e. it is decreasing in size), your child will have visits to

the clinic at week 18 and 42. If it is not regressing, visits will continue every 4 weeks until the

beginning of regression or the end of the observation period.

HOW LONG WILL THE STUDY LAST?

Your child will be in the study for 3 years, with annual follow-up. Your doctor will recommend

surgery if the tumour increases in volume to more than 50% from the initial size or if the urine

analysis suggests the tumour is behaving more aggressively, in the absence of any volume

increase. We will also recommend surgery if at the end of the observation period the tumour or

mass remains. You can withdraw your child from the study at anytime, and your decision will not

influence his/her medical care.

WHAT ARE THE RISK OF PARTICIPATING?

This study has been designed to minimise the risks for your child. In spite of this, there is a

potential risk of tumour progression. This is due to the fact that neuroblastoma in older children

can grow, and eventually, extend to other body parts. It is possible that a tumour like this in your

baby could have the same behaviour. There are no published cases or reports of localised tumours

in babies with favourable catecholamines that have progressed to more advanced stages without

having showed important local growth first. Although it is expected, and it is more likely, that

your baby’s tumour decreases or, at least, remains stable in size during the study, there is a small

risk that it may growth rapidly. If the tumour is bigger at the time of surgery, this procedure can

be more difficult and your child exposed to more complications than if he/she were operated with

a smaller tumour. Depending on the size and the biological characteristics, he/she may need other

treatments (chemotherapy and/or radiotherapy). This last is very unlikely, but this remote

possibility still exists.

WHAT ARE THE BENEFITS OF TAKING PART IN THIS STUDY?

If the tumour or mass disappears spontaneously, your child will be cured without going through

the anaesthetic or surgical risks that are involved in an operation. There is no other direct benefit

for him/her. We expect the information obtained by the investigators will help and benefit other

patients like your son/daughter in the future.

WHAT ARE THE OTHER OPTIONS OF TREATMENTS?

There are other options if you don’t want your child to participate:

Standard treatment for suprarenal masses: immediate surgery.

Any other research therapies you can be offered.

You are expected to talk to your doctor to discuss all the options.

WHAT ABOUT CONFIDENTIALITY?

All the information (individual, clinical and biological data) collected from your child will be

treated according to EU Directive 95/46/CE, 24 October 1995, concerning to protection of

individual rights regarding to confidential data, and to the national laws within your own country.



The investigator will enter your child’s individual data into a secure database. All the reports will

be anonymised by using an alphanumeric code. Your child’s name will never appear and his/her

identity will never be revealed, except in case of emergency or legal requirement. The results of

the study will be conveyed to health authorities and to the scientific community (publications).

You are entitled to access the data, rectify, cancel and object to them at any time notifying to the

study physician.

WHAT ARE THE COSTS?

To take part in this study will not represent an extra cost for your child’s insurance company or

health institutions, since all the tests are regularly performed in the clinical setting. You will not

receive payment for participation in this study.

WHAT ARE MY CHILD’S RIGHTS?

To take part in this study is voluntary. You can choose whether your child participates or not or to

withdraw from the study any time. Before withdrawing your child from this study you are

encouraged to speak to your child’s doctor. If your child withdraws from the study, this will not

influence his/her further care.

We will keep you informed about the progress of the study and results, and, of course, all those

facts that can influence in your child’s well-being.

IF YOU HAVE ANY QUESTIONS, PLEASE ASK YOUR DOCTOR.

DOCTOR´S NAME:

ADDRESS:

PHONE:



A21.5.2 Informed consent for parents

CONSENT FORM FOR AN OBSERVATIONAL APPROACH IN INFANTS WITH

SUPRARENAL MASSES



sheet.



I have talked to:







the study.




Yes� No�


trial.



Name: Name:

Date: Date



APPENDIX 22. CONTACT INFORMATION

A22.1 Principal Investigators LOW RISK (LR) STUDY

Gudrun SCHLEIERMACHER (PI)

Email: [email protected]

France

Kate WHEELER (coPI)


UK

INTERMEDIATE RISK (IR) STUDY

Andrea DI CATALDO (PI)


Italy

Adela CANETE (coPI)


Spain

NEONATAL SUPRARENAL MASS (NSM) STUDY

Adela CANETE (PI)


Spain

Vassilios PAPADAKIS (coPI)


Greece

A22.2 National Coordinators

Toby TRAHAIR


Australia

Ruth LADENSTEIN


Austria

Bénédicte BRICHARD


Belgium

Josef MALIS


Czech Republic

Henrik SCHROEDER


Denmark

Gudrun Schleiermacher


France (LR Study)

Anne-Sophie Defachelles


France (IR Study)

Dominique Plantaz


France (NSM Study)

Vassilios PAPADAKIS


Greece

Margarita Baka Email:[email protected] Greece

Miklós GARAMI


Hungary

Anne O’MEARA

Email: Anne.O'[email protected]

Ireland

Michael CAPRA


Ireland

Shifra ASH


Israel



Massimo CONTE


Italy (LR Study)

Andrea DI CATALDO

Email: Email: [email protected]

Italy (IR Study)

Massimo CONTE


Italy (NSE Study)

Maria W GUNNES


Norway (LR Study)

Ellen RUUD


Norway (IR Study)

Maria W GUNNES


Norway (NSM Study)

Walentyna BALWIERZ


Poland

Anna FORJAZ DE LACERDA


Portugal

Dragana VUJIC


Serbia

Pavel BICIAN


Slovakia

Adela CANETE


Spain (LR Study)

Victoria CASTEL


Spain (IR Study)

Adela CANETE


Spain (NSM Study)

Per KOGNER


Sweden (including Iceland)

Maja BECK-POPOVIC


Switzerland

Kate WHEELER


UK

A22.3 Data Monitoring Committee

Angelika EGGERT, Prof

Universitätsklinikum Essen Paediatric Haematology/Oncology and Endocrinology Zentrum für Kinderheilkunde Hufelandstraße 55 45122 Essen Tel: +49-0-201-723-3755 Fax: +49-0-201-723-4694 Email: [email protected]

Germany



Lisa DILLER, Associate Prof Dana-Farber Cancer Institute Department of Pediatric Oncology 44 Binney Street Boston MA 02115 Tel: +1-617-632-5642 Fax: +1-617-5828218 Email: [email protected]

USA

Richard SPOSTO, Prof

Children’s Hospital Los Angeles Children's Center for Cancer and Blood Diseases 4650 Sunset Blvd. Los Angeles, CA 90027 Tel: +1-323-644-8582 Fax: +1-323-671-1803 Email: [email protected]

USA

A22.4 Biology Committee

Toby TRAHAIR


Australia

Inge AMBROS


Austria

Peter AMBROS


Austria

Frank SPELEMAN


Belgium

Nadine VAN ROY


Belgium

Ales VICHA, Dr.

Department of Paediatric Haematology and Oncology Charles University 2nd Faculty of Medicine V uvalu 84 Prague 5- Motol 15008 Tel: +420-224436470, 6494 Fax: +420-224436417 Email: [email protected], [email protected]

Czech Republic

Jean BÉNARD


France



Valérie COMBARET, Dr.

Laboratoire de Recherche Translationnelle Batiment CHENEY D 1er étage Centre Léon BERARD 28, rue Laennec 69373 LYON cedex 08 Email: [email protected]

France

Jérome COUTURIER

Département d'Oncologie Pédiatrique Institut Curie 26 rue d'Ulm 75248 Paris Cedex 05 Tel.: +33 1 56 24 67 28 Fax: +33 1 53 10 26 47 Email: [email protected]

France

Gudrun SCHLEIERMACHER

Département d'Oncologie Pédiatrique Institut Curie 26 rue d'Ulm 75248 Paris Cedex 05

Tel.: +33 1 44 32 45 54, +33 1 44 32 45 55 Email: [email protected]

France

Alexander VALENT


France

Raymond STALLINGS


Ireland

Marta JEISON, Dr.

Head of the Ca-Cytogenetic and Molecular Cytogenetic Lab. Pediatric Hematology Oncology Dept. Schneider Children's Medical Center of Israel Kaplan 14

49202- Petah Tikva. Tel: 972-3-9253185

Fax: 972-3-9253042


Israel

Raffaella DEFFERRARI


Italy

Katia MAZZOCCO


Italy



Gian Paolo TONINI, Dr.

Director Translational Paediatric Oncology National Cancer Research Institute (IST) L.go R. Benzi, 10 16132 Genoa Tel: ++39-010-5737381/487/463/490/430 Fax: ++39-010-5737487 Email: [email protected]

Italy

Klaus BEISKE, Dr.

Department of Pathology Oslo University Hospital Rikshospitalet Sognsvannsveien 20 N-0027 Oslo

Tel: +47 2307 1400 (secretary) +47 2307 4076 (office) Fax: +47 2307 1410 Email: [email protected] [email protected]

Norway

Bárbara MARQUES, Dr. Ricardo JORGE, Dr.

Departamento de Genética Instituto Nacional de Saúde, IP Av. Padre Cruz, 1649-016 Lisboa Tel: (+351) 21 752 64 11 Fax: (+351) 21 752 64 10 Email: [email protected]

Portugal

Rosa NOGUERA, Profª. Marta PIQUERAS Eva VILLAMÓN

Department of Pathology Medical School of Valencia University of Valencia Avda. Blasco Ibañez 15 46010 Valencia Tel: +34 96 398 3948, +34 96 398 3626 Tel Molecular Pathology Lab : +34 96 386 4100 Ext: 52467 Fax: +34 96 398 3226 Email: [email protected], [email protected], [email protected]

Spain

Eva VILLAMÓN


Spain

mailto:[email protected]



Tommy MARTINSSON, Prof.

Department of Clinical Genetics The Sahlgrenska Academy at the University of Gothenburg Sahlgrenska University Hospital S-41345 Gothenburg Tel: +46 31 343 4803 Fax : +46 31 842 160 Email: [email protected]

Sweden

Nicole GROSS, Dr.

Chef de Secteur de Recherche Recherche en Oncologie Pédiatrique University Hospital CHUV BH-11-951 Rue du Bugnon 46 CH-1011 Lausanne Tel:+41 21 31 41 746 Fax: +41 21 31 43 664 Email: [email protected]

Switzerland

Clare BEDWELL


UK

Nick BOWN, Dr.

Cytogenetics Unit Northern Genetics Service Institute for Human Genetics Central Parkway Newcastle upon Tyne NE1 3BZ Tel: +44 191-241-8703 Fax: +44 191-241-8713 Email: [email protected]

UK

John LUNEC, Prof.

Gillespie Chair of Molecular Oncology Northern Institute for Cancer Research Medical School Newcastle University Framlington Place Newcastle upon Tyne NE2 4HH Tel: +44 191 246 4420 Fax:+44 191 246 4301 Email: [email protected]

UK

Deborah TWEDDLE, Dr. Email: [email protected]

UK



A22.5 Nuclear Medicine and Physic Committee Coordinator

Ariane Boubaker, Dr.

Sevice de Médecine Nucléaire BH-07 Centre Hospitalier Universitaire Vaudois rue du Bougnon CH - 1011 Lausanne Phone: +41 21 131 44353 Fax: +41 21 314 43 43 Email: [email protected]

Switzerland

A22.6 Bone Marrow Committee Coordinator

Klaus BEISKE, Dr.


Norway

A22.7 Pathology Committee

Gabriele AMANN, Dr.

Clinical Institute of Pathology Medical University of Vienna, AKH Waehringer Guertel 18-20 1090 Wien

Austria

Michel PEUCHMAUR, Prof.

Service de Pathologie APHP et Université Paris 7 Hôpital Robert Debré, 48 Boulevard Sérurier 75019 Paris

France

Emanuele D’AMORE, Dr.

UO di Anatomia Patologica Ospedale San Bortolo Viale F. Rodolfi n.37 36100 Vicenza

Italy

Claudio GAMBINI, Dr.

U.O. di Anatomia Patologica Istituto G. Gaslini Largo G. Gaslini 5 1648 Genova

Italy

Klaus BEISKE, Dr.


Norway

Samuel NAVARRO, Dr.

Departamento de Patologia Facultad de Medicina AVDA Blasco Ibanez 17 E-4610 Valencia

Spain



Catherine CULLINANE, Dr.

Department of Histopathology, Level 5, Bexley Wing, St. James’s University Hospital, Beckett Street, Leeds LS9 7TF

UK

A22.8 Radiology Committee

Claudio GRANTA, Dr.

Italy

Roisin HAYES, Dr.


Ireland

Liora KORENRIECH, Dr.

Israel

Ingegerd AAGENÆS, Dr. Department of Radiology Oslo University Hospital Email: [email protected]

Norway

Katerina NOVOTNA, Dr.

Tel: +421-904-047 234 Email: [email protected]

Slovakia

SANGÜESA, Dr.

H.I.LA FE. AVDA DE CAMPANAR 21 46009 VALENCIA

Spain

MURO, Dr.

H.I.LA FE. AVDA DE CAMPANAR 21 46009 VALENCIA Email: [email protected]

Spain

Fiona DICKINSON, Dr.

UK

A22.9 Radiotherapy Committee

Mark GAZE, Dr.

University College London Hospitals NHS Foundation Trust First Floor Central 250 Euston Road London, NW1 2PG Email: [email protected]

UK

Tom BOTERBERG, Dr.


Belgium

Catriona O’SULLIVAN, Dr.


Ireland



Nili RAMU, Dr.

Israel

Knut LOTE, Dr. Department of Oncology Radiumhospitalet Oslo University Hospital Email: [email protected]

Norway

Branislav SEPESI, Dr.

Tel: +421-48-4413260 Email: [email protected]

Slovakia

BABAL, Dr.


Spain

A22.10 Statistics Committee

Veronique MOSSERI, Dr.


France

José BERMÚDEZ, Dr.


Spain

A22.11 Surgery Committee

Sabine Sarnacki, Dr.


France

Martin CORBALLY, Dr.


Ireland

Ran STEINBERG, Dr.

Israel

Kristin BJØRNLAND, Dr. Department of Pediatric Surgery Oslo University Hospital Email: [email protected]

Norway

Jan NOVOTNY, Dr.

Tel: +421-915-838414 Email: [email protected]

Slovakia

MARCO, Dr.


Spain

COSTA, Dr.

H.I.LA FE. AVDA DE CAMPANAR 21 46009 VALENCIA

Spain

Roly SQUIRE, Dr.


UK



APPENDIX 23. DECLARATION OF HELSINKI

WORLD MEDICAL ASSOCIATION DECLARATION OF HELSINKI

Ethical Principles for Medical Research Involving Human Subjects

Adopted by the 18th WMA General Assembly Helsinki, Finland, June 1964 and amended by the

29th WMA General Assembly, Tokyo, Japan, October 1975

35th WMA General Assembly, Venice, Italy, October 1983

41st WMA General Assembly, Hong Kong, September 1989

48th WMA General Assembly, Somerset West, Republic of South Africa, October 1996

52nd WMA General Assembly, Edinburgh, Scotland, October 2000

53rd WMA General Assembly, Washington 2002 (Note of Clarification on paragraph 29 added)

55th WMA General Assembly, Tokyo 2004 (Note of Clarification on Paragraph 30 added)

59th WMA General Assembly, Seoul, October 2008

A23.1 Introduction

1. The World Medical Association (WMA) has developed the Declaration of Helsinki as a

statement of ethical principles for medical research involving human subjects, including research

on identifiable human material and data.

The Declaration is intended to be read as a whole and each of its constituent paragraphs should

not be applied without consideration of all other relevant paragraphs.

2. Although the Declaration is addressed primarily to physicians, the WMA encourages other

participants in medical research involving human subjects to adopt these principles.

3. It is the duty of the physician to promote and safeguard the health of patients, including those

who are involved in medical research. The physician's knowledge and conscience are dedicated to

the fulfilment of this duty.

4. The Declaration of Geneva of the WMA binds the physician with the words, “The health of my

patient will be my first consideration,” and the International Code of Medical Ethics declares that,

“A physician shall act in the patient's best interest when providing medical care.”

5. Medical progress is based on research that ultimately must include studies involving human

subjects. Populations that are underrepresented in medical research should be provided

appropriate access to participation in research.

6. In medical research involving human subjects, the well-being of the individual research subject

must take precedence over all other interests.

7. The primary purpose of medical research involving human subjects is to understand the causes,

development and effects of diseases and improve preventive, diagnostic and therapeutic

interventions (methods, procedures and treatments). Even the best current interventions must be

evaluated continually through research for their safety, effectiveness, efficiency, accessibility and

quality.

8. In medical practice and in medical research, most interventions involve risks and burdens.

9. Medical research is subject to ethical standards that promote respect for all human subjects and

protect their health and rights. Some research populations are particularly vulnerable and need



special protection. These include those who cannot give or refuse consent for themselves and

those who may be vulnerable to coercion or undue influence.

10. Physicians should consider the ethical, legal and regulatory norms and standards for research

involving human subjects in their own countries as well as applicable international norms and

standards. No national or international ethical, legal or regulatory requirement should reduce or

eliminate any of the protections for research subjects set forth in this Declaration.

A23.2 Principles for All Medical Research

11. It is the duty of physicians who participate in medical research to protect the life, health,

dignity, integrity, right to self-determination, privacy, and confidentiality of personal information

of research subjects.

12. Medical research involving human subjects must conform to generally accepted scientific

principles, be based on a thorough knowledge of the scientific literature, other relevant sources of

information, and adequate laboratory and, as appropriate, animal experimentation. The welfare of

animals used for research must be respected.

13. Appropriate caution must be exercised in the conduct of medical research that may harm the

environment.

14. The design and performance of each research study involving human subjects must be clearly

described in a research protocol. The protocol should contain a statement of the ethical

considerations involved and should indicate how the principles in this Declaration have been

addressed. The protocol should include information regarding funding, sponsors, institutional

affiliations, other potential conflicts of interest, incentives for subjects and provisions for treating

and/or compensating subjects who are harmed as a consequence of participation in the research

study. The protocol should describe arrangements for post-study access by study subjects to

interventions identified as beneficial in the study or access to other appropriate care or benefits.

15. The research protocol must be submitted for consideration, comment, guidance and approval

to a research ethics committee before the study begins. This committee must be independent of

the researcher, the sponsor and any other undue influence. It must take into consideration the laws

and regulations of the country or countries in which the research is to be performed as well as

applicable international norms and standards but these must not be allowed to reduce or eliminate

any of the protections for research subjects set forth in this Declaration. The committee must have

the right to monitor ongoing studies. The researcher must provide monitoring information to the

committee, especially information about any serious adverse events. No change to the protocol

may be made without consideration and approval by the committee.

16. Medical research involving human subjects must be conducted only by individuals with the

appropriate scientific training and qualifications. Research on patients or healthy volunteers

requires the supervision of a competent and appropriately qualified physician or other health care

professional. The responsibility for the protection of research subjects must always rest with the

physician or other health care professional and never the research subjects, even though they have

given consent.

17. Medical research involving a disadvantaged or vulnerable population or community is only

justified if the research is responsive to the health needs and priorities of this population or



community and if there is a reasonable likelihood that this population or community stands to

benefit from the results of the research.

18. Every medical research study involving human subjects must be preceded by careful

assessment of predictable risks and burdens to the individuals and communities involved in the

research in comparison with foreseeable benefits to them and to other individuals or communities

affected by the condition under investigation.

19. Every clinical trial must be registered in a publicly accessible database before recruitment of

the first subject.

20. Physicians may not participate in a research study involving human subjects unless they are

confident that the risks involved have been adequately assessed and can be satisfactorily managed.

Physicians must immediately stop a study when the risks are found to outweigh the potential

benefits or when there is conclusive proof of positive and beneficial results.

21. Medical research involving human subjects may only be conducted if the importance of the

objective outweighs the inherent risks and burdens to the research subjects.

22. Participation by competent individuals as subjects in medical research must be voluntary.

Although it may be appropriate to consult family members or community leaders, no competent

individual may be enrolled in a research study unless he or she freely agrees.

23. Every precaution must be taken to protect the privacy of research subjects and the

confidentiality of their personal information and to minimize the impact of the study on their

physical, mental and social integrity.

24. In medical research involving competent human subjects, each potential subject must be

adequately informed of the aims, methods, sources of funding, any possible conflicts of interest,

institutional affiliations of the researcher, the anticipated benefits and potential risks of the study

and the discomfort it may entail, and any other relevant aspects of the study. The potential subject

must be informed of the right to refuse to participate in the study or to withdraw consent to

participate at any time without reprisal. Special attention should be given to the specific

information needs of individual potential subjects as well as to the methods used to deliver the

information. After ensuring that the potential subject has understood the information, the

physician or another appropriately qualified individual must then seek the potential subject’s

freely-given informed consent, preferably in writing. If the consent cannot be expressed in

writing, the non-written consent must be formally documented and witnessed.

25. For medical research using identifiable human material or data, physicians must normally seek

consent for the collection, analysis, storage and/or reuse. There may be situations where consent

would be impossible or impractical to obtain for such research or would pose a threat to the

validity of the research. In such situations the research may be done only after consideration and

approval of a research ethics committee.

26. When seeking informed consent for participation in a research study the physician should be

particularly cautious if the potential subject is in a dependent relationship with the physician or

may consent under duress. In such situations the informed consent should be sought by an

appropriately qualified individual who is completely independent of this relationship.



27. For a potential research subject who is incompetent, the physician must seek informed consent

from the legally authorized representative. These individuals must not be included in a research

study that has no likelihood of benefit for them unless it is intended to promote the health of the

population represented by the potential subject, the research cannot instead be performed with

competent persons, and the research entails only minimal risk and minimal burden.

28. When a potential research subject who is deemed incompetent is able to give assent to

decisions about participation in research, the physician must seek that assent in addition to the

consent of the legally authorized representative. The potential subject’s dissent should be

respected.

29. Research involving subjects who are physically or mentally incapable of giving consent, for

example, unconscious patients, may be done only if the physical or mental condition that prevents

giving informed consent is a necessary characteristic of the research population. In such

circumstances the physician should seek informed consent from the legally authorized

representative. If no such representative is available and if the research cannot be delayed, the

study may proceed without informed consent provided that the specific reasons for involving

subjects with a condition that renders them unable to give informed consent have been stated in

the research protocol and the study has been approved by a research ethics committee. Consent to

remain in the research should be obtained as soon as possible from the subject or a legally

authorized representative.

30. Authors, editors and publishers all have ethical obligations with regard to the publication of

the results of research. Authors have a duty to make publicly available the results of their research

on human subjects and are accountable for the completeness and accuracy of their reports. They

should adhere to accepted guidelines for ethical reporting. Negative and inconclusive as well as

positive results should be published or otherwise made publicly available. Sources of funding,

institutional affiliations and conflicts of interest should be declared in the publication. Reports of

research not in accordance with the principles of this Declaration should not be accepted for

publication.

A23.3 Additional Principles for Medical Research Combined with Medical Care

31. The physician may combine medical research with medical care only to the extent that the

research is justified by its potential preventive, diagnostic or therapeutic value and if the physician

has good reason to believe that participation in the research study will not adversely affect the

health of the patients who serve as research subjects.

32. The benefits, risks, burdens and effectiveness of a new intervention must be tested against

those of the best current proven intervention, except in the following circumstances:

The use of placebo, or no treatment, is acceptable in studies where no current proven

intervention exists; or

Where for compelling and scientifically sound methodological reasons the use of placebo

is necessary to determine the efficacy or safety of an intervention and the patients who

receive placebo or no treatment will not be subject to any risk of serious or irreversible

harm. Extreme care must be taken to avoid abuse of this option.

33. At the conclusion of the study, patients entered into the study are entitled to be informed about

the outcome of the study and to share any benefits that result from it, for example, access to

interventions identified as beneficial in the study or to other appropriate care or benefits.



34. The physician must fully inform the patient which aspects of the care are related to the

research. The refusal of a patient to participate in a study or the patient’s decision to withdraw

from the study must never interfere with the patient-physician relationship.

35. In the treatment of a patient, where proven interventions do not exist or have been ineffective,

the physician, after seeking expert advice, with informed consent from the patient or a legally

authorized representative, may use an unproven intervention if in the physician's judgement it

offers hope of saving life, re-establishing health or alleviating suffering. Where possible, this

intervention should be made the object of research, designed to evaluate its safety and efficacy. In

all cases, new information should be recorded and, where appropriate, made publicly available.



APPENDIX 24. REFERENCE LIST

[1] Brodeur GM, Pritchard J, Berthold F, Carlsen NL, Castel V, Castelberry RP, et al. Revisions of the international criteria for neuroblastoma diagnosis, staging, and response to treatment. J Clin Oncol. 1993 Aug;11(8):1466-77. [2] Cohn SL, Pearson AD, London WB, Monclair T, Ambros PF, Brodeur GM, et al. The International Neuroblastoma Risk Group (INRG) classification system: an INRG Task Force report. J Clin Oncol. 2009 Jan 10;27(2):289-97. [3] De Bernardi B, Gerrard M, Boni L, Rubie H, Canete A, Di Cataldo A, et al. Excellent outcome with reduced treatment for infants with disseminated neuroblastoma without MYCN gene amplification. J Clin Oncol. 2009 Mar 1;27(7):1034-40. [4] Janoueix-Lerosey I, Schleiermacher G, Michels E, Mosseri V, Ribeiro A, Lequin D, et al. Overall genomic pattern is a predictor of outcome in neuroblastoma. J Clin Oncol. 2009 Mar 1;27(7):1026-33. [5] Schleiermacher G, Michon J, Huon I, d'Enghien CD, Klijanienko J, Brisse H, et al. Chromosomal CGH identifies patients with a higher risk of relapse in neuroblastoma without MYCN amplification. British journal of cancer. 2007 Jul 16;97(2):238-46. [6] Cecchetto G, Mosseri V, De Bernardi B, Helardot P, Monclair T, Costa E, et al. Surgical risk factors in primary surgery for localized neuroblastoma: the LNESG1 study of the European International Society of Pediatric Oncology Neuroblastoma Group. J Clin Oncol. 2005 Nov 20;23(33):8483-9. [7] Monclair T, Brodeur GM, Ambros PF, Brisse HJ, Cecchetto G, Holmes K, et al. The International Neuroblastoma Risk Group (INRG) staging system: an INRG Task Force report. J Clin Oncol. 2009 Jan 10;27(2):298-303. [8] De Bernardi B, Mosseri V, Rubie H, Castel V, Foot A, Ladenstein R, et al. Treatment of localised resectable neuroblastoma. Results of the LNESG1 study by the SIOP Europe Neuroblastoma Group. British journal of cancer. 2008 Oct 7;99(7):1027-33. [9] Bagatell R, Beck-Popovic M, London WB, Zhang Y, Pearson AD, Matthay KK, et al. Significance of MYCN amplification in international neuroblastoma staging system stage 1 and 2 neuroblastoma: a report from the International Neuroblastoma Risk Group database. J Clin Oncol. 2009 Jan 20;27(3):365-70. [10] Canete A, Gerrard M, Rubie H, Castel V, Di Cataldo A, Munzer C, et al. Poor survival for infants with MYCN-amplified metastatic neuroblastoma despite intensified treatment: the International Society of Paediatric Oncology European Neuroblastoma Experience. J Clin Oncol. 2009 Mar 1;27(7):1014-9. [11] Hero B, Simon T, Spitz R, Ernestus K, Gnekow AK, Scheel-Walter HG, et al. Localized infant neuroblastomas often show spontaneous regression: results of the prospective trials NB95-S and NB97. J Clin Oncol. 2008 Mar 20;26(9):1504-10. [12] Rubie H, Michon J, Plantaz D, Peyroulet MC, Coze C, Frappaz D, et al. Unresectable localized neuroblastoma: improved survival after primary chemotherapy including carboplatin-etoposide. Neuroblastoma Study Group of the Societe Francaise d'Oncologie Pediatrique (SFOP). British journal of cancer. 1998 Jun;77(12):2310-7. [13] Castel V, Badal MD, Bezanilla JL, Llombart A, Ruiz-Jimenez JI, Sanchez de Toledo J, et al. Treatment of stage III neuroblastoma with emphasis on intensive induction chemotherapy: a report from the Neuroblastoma Group of the Spanish Society of Pediatric Oncology. Med Pediatr Oncol. 1995 Jan;24(1):29-35. [14] Garaventa A, De Bernardi B, Pianca C, Donfrancesco A, Cordero di Montezemolo L, Di Tullio MT, et al. Localized but unresectable neuroblastoma: treatment and outcome of 145 cases. Italian Cooperative Group for Neuroblastoma. J Clin Oncol. 1993 Sep;11(9):1770-9. [15] Garaventa A, Boni L, Lo Piccolo MS, Tonini GP, Gambini C, Mancini A, et al. Localized unresectable neuroblastoma: results of treatment based on clinical prognostic factors. Ann Oncol. 2002 Jun;13(6):956-64.



[16] Castleberry RP, Kun LE, Shuster JJ, Altshuler G, Smith IE, Nitschke R, et al. Radiotherapy improves the outlook for patients older than 1 year with Pediatric Oncology Group stage C neuroblastoma. J Clin Oncol. 1991 May;9(5):789-95. [17] Modak S, Kushner BH, LaQuaglia MP, Kramer K, Cheung NK. Management and outcome of stage 3 neuroblastoma. Eur J Cancer. 2009 Jan;45(1):90-8. [18] Park JR, Villablanca JG, London WB, Gerbing RB, Haas-Kogan D, Adkins ES, et al. Outcome of high-risk stage 3 neuroblastoma with myeloablative therapy and 13-cis-retinoic acid: a report from the Children's Oncology Group. Pediatric blood & cancer. 2009 Jan;52(1):44-50. [19] Matthay KK, Villablanca JG, Seeger RC, Stram DO, Harris RE, Ramsay NK, et al. Treatment of high-risk neuroblastoma with intensive chemotherapy, radiotherapy, autologous bone marrow transplantation, and 13-cis-retinoic acid. Children's Cancer Group. The New England journal of medicine. 1999 Oct 14;341(16):1165-73. [20] Brodeur GM, Maris JM, Yamashiro DJ, Hogarty MD, White PS. Biology and genetics of human neuroblastomas. J Pediatr Hematol Oncol. 1997 Mar-Apr;19(2):93-101. [21] Maris JM, Matthay KK. Molecular biology of neuroblastoma. J Clin Oncol. 1999 Jul;17(7):2264-79. [22] Brodeur GM. Neuroblastoma: biological insights into a clinical enigma. Nature reviews. 2003 Mar;3(3):203-16. [23] Maris JM. The biologic basis for neuroblastoma heterogeneity and risk stratification. Current opinion in pediatrics. 2005 Feb;17(1):7-13. [24] Seeger RC, Brodeur GM, Sather H, Dalton A, Siegel SE, Wong KY, et al. Association of multiple copies of the N-myc oncogene with rapid progression of neuroblastomas. The New England journal of medicine. 1985 Oct 31;313(18):1111-6. [25] Ladenstein R, Ambros IM, Potschger U, Amann G, Urban C, Fink FM, et al. Prognostic significance of DNA di-tetraploidy in neuroblastoma. Med Pediatr Oncol. 2001 Jan;36(1):83-92. [26] Look AT, Hayes FA, Shuster JJ, Douglass EC, Castleberry RP, Bowman LC, et al. Clinical relevance of tumor cell ploidy and N-myc gene amplification in childhood neuroblastoma: a Pediatric Oncology Group study. J Clin Oncol. 1991 Apr;9(4):581-91. [27] Caron H, van Sluis P, de Kraker J, Bokkerink J, Egeler M, Laureys G, et al. Allelic loss of chromosome 1p as a predictor of unfavorable outcome in patients with neuroblastoma. The New England journal of medicine. 1996 Jan 25;334(4):225-30. [28] Luttikhuis ME, Powell JE, Rees SA, Genus T, Chughtai S, Ramani P, et al. Neuroblastomas with chromosome 11q loss and single copy MYCN comprise a biologically distinct group of tumours with adverse prognosis. British journal of cancer. 2001 Aug 17;85(4):531-7. [29] Schleiermacher G, Delattre O, Peter M, Mosseri V, Delonlay P, Vielh P, et al. Clinical relevance of loss heterozygosity of the short arm of chromosome 1 in neuroblastoma: a single-institution study. International journal of cancer. 1996 Apr 22;69(2):73-8. [30] Attiyeh EF, London WB, Mosse YP, Wang Q, Winter C, Khazi D, et al. Chromosome 1p and 11q deletions and outcome in neuroblastoma. The New England journal of medicine. 2005 Nov 24;353(21):2243-53. [31] Brinkschmidt C, Christiansen H, Terpe HJ, Simon R, Lampert F, Boecker W, et al. Distal chromosome 17 gains in neuroblastomas detected by comparative genomic hybridization (CGH) are associated with a poor clinical outcome. Med Pediatr Oncol. 2001 Jan;36(1):11-3. [32] Lastowska M, Cullinane C, Variend S, Cotterill S, Bown N, O'Neill S, et al. Comprehensive genetic and histopathologic study reveals three types of neuroblastoma tumors. J Clin Oncol. 2001 Jun 15;19(12):3080-90. [33] Plantaz D, Mohapatra G, Matthay KK, Pellarin M, Seeger RC, Feuerstein BG. Gain of chromosome 17 is the most frequent abnormality detected in neuroblastoma by comparative genomic hybridization. The American journal of pathology. 1997 Jan;150(1):81-9. [34] Bown N, Lastowska M, Cotterill S, O'Neill S, Ellershaw C, Roberts P, et al. 17q gain in neuroblastoma predicts adverse clinical outcome. U.K. Cancer Cytogenetics Group and the U.K. Children's Cancer Study Group. Med Pediatr Oncol. 2001 Jan;36(1):14-9. [35] Brinkschmidt C, Poremba C, Christiansen H, Simon R, Schafer KL, Terpe HJ, et al. Comparative genomic hybridization and telomerase activity analysis identify two biologically different groups of 4s neuroblastomas. British journal of cancer. 1998 Jun;77(12):2223-9.



[36] Plantaz D, Vandesompele J, Van Roy N, Lastowska M, Bown N, Combaret V, et al. Comparative genomic hybridization (CGH) analysis of stage 4 neuroblastoma reveals high frequency of 11q deletion in tumors lacking MYCN amplification. International journal of cancer. 2001 Mar 1;91(5):680-6. [37] Vandesompele J, Baudis M, De Preter K, Van Roy N, Ambros P, Bown N, et al. Unequivocal delineation of clinicogenetic subgroups and development of a new model for improved outcome prediction in neuroblastoma. J Clin Oncol. 2005 Apr 1;23(10):2280-99. [38] Vandesompele J, Speleman F, Van Roy N, Laureys G, Brinskchmidt C, Christiansen H, et al. Multicentre analysis of patterns of DNA gains and losses in 204 neuroblastoma tumors: how many genetic subgroups are there? Med Pediatr Oncol. 2001 Jan;36(1):5-10. [39] Vandesompele J, Van Roy N, Van Gele M, Laureys G, Ambros P, Heimann P, et al. Genetic heterogeneity of neuroblastoma studied by comparative genomic hybridization. Genes, chromosomes & cancer. 1998 Oct;23(2):141-52. [40] Mosse YP, Diskin SJ, Wasserman N, Rinaldi K, Attiyeh EF, Cole K, et al. Neuroblastomas have distinct genomic DNA profiles that predict clinical phenotype and regional gene expression. Genes, chromosomes & cancer. 2007 Oct;46(10):936-49. [41] Janoueix-Lerosey I, Hupe P, Maciorowski Z, La Rosa P, Schleiermacher G, Pierron G, et al. Preferential occurrence of chromosome breakpoints within early replicating regions in neuroblastoma. Cell cycle (Georgetown, Tex. 2005 Dec;4(12):1842-6. [42] Schleiermacher G, Bourdeaut F, Combaret V, Picrron G, Raynal V, Aurias A, et al. Stepwise occurrence of a complex unbalanced translocation in neuroblastoma leading to insertion of a telomere sequence and late chromosome 17q gain. Oncogene. 2005 May 5;24(20):3377-84. [43] Ambros IM, Benard J, Boavida M, Bown N, Caron H, Combaret V, et al. Quality assessment of genetic markers used for therapy stratification. J Clin Oncol. 2003 Jun 1;21(11):2077-84. [44] Ambros PF, Ambros IM, Brodeur GM, Haber M, Khan J, Nakagawara A, et al. International consensus for neuroblastoma molecular diagnostics: report from the International Neuroblastoma Risk Group (INRG) Biology Committee. British journal of cancer. 2009 May 5;100(9):1471-82. [45] Brodeur GM, Seeger RC, Barrett A, Berthold F, Castleberry RP, D'Angio G, et al. International criteria for diagnosis, staging, and response to treatment in patients with neuroblastoma. J Clin Oncol. 1988 Dec;6(12):1874-81. [46] Shimada H, Ambros IM, Dehner LP, Hata J, Joshi VV, Roald B, et al. The International Neuroblastoma Pathology Classification (the Shimada system). Cancer. 1999 Jul 15;86(2):364-72. [47] Shimada H, Ambros IM, Dehner LP, Hata J, Joshi VV, Roald B. Terminology and morphologic criteria of neuroblastic tumors: recommendations by the International Neuroblastoma Pathology Committee. Cancer. 1999 Jul 15;86(2):349-63. [48] Kerbl R, Urban CE, Ambros IM, Dornbusch HJ, Schwinger W, Lackner H, et al. Neuroblastoma mass screening in late infancy: insights into the biology of neuroblastic tumors. J Clin Oncol. 2003 Nov 15;21(22):4228-34. [49] Kerbl R, Urban CE, Lackner H, Hofler G, Ambros IM, Ratschek M, et al. Connatal localized neuroblastoma. The case to delay treatment. Cancer. 1996 Apr 1;77(7):1395-401. [50] Schwab M, Westermann F, Hero B, Berthold F. Neuroblastoma: biology and molecular and chromosomal pathology. The lancet oncology. 2003 Aug;4(8):472-80. [51] Nadler EP, Barksdale EM. Adrenal masses in the newborn. Seminars in pediatric surgery. 2000 Aug;9(3):156-64. [52] Donoghue V, Ryan S, Twomey E. Perinatal tumours: the contribution of radiology to management. Pediatric radiology. 2008 Jun;38 Suppl 3:S477-83. [53] Mahony R, McParland P. Approaches to the management of antenatally diagnosed congenital tumours. Pediatric radiology. 2009 Nov;39(11):1173-8. [54] Avni FE, Massez A, Cassart M. Tumours of the fetal body: a review. Pediatric radiology. 2009 Nov;39(11):1147-57.



[55] Deeg KH, Bettendorf U, Hofmann V. Differential diagnosis of neonatal adrenal haemorrhage and congenital neuroblastoma by colour coded Doppler sonography and power Doppler sonography. European journal of pediatrics. 1998 Apr;157(4):294-7. [56] Nuchtern JG. Perinatal neuroblastoma. Seminars in pediatric surgery. 2006 Feb;15(1):10-6. [57] Noguchi S, Masumoto K, Taguchi T, Takahashi Y, Tsuneyoshi M, Suita S. Adrenal cytomegaly: two cases detected by prenatal diagnosis. Asian journal of surgery / Asian Surgical Association. 2003 Oct;26(4):234-6. [58] Barrette S, Bernstein ML, Leclerc JM, Champagne MA, Samson Y, Brossard J, et al. Treatment complications in children diagnosed with neuroblastoma during a screening program. J Clin Oncol. 2006 Apr 1;24(10):1542-5. [59] Suita S, Tajiri T, Higashi M, Tanaka S, Kinoshita Y, Takahashi Y, et al. Insights into infant neuroblastomas based on an analysis of neuroblastomas detected by mass screening at 6 months of age in Japan. Eur J Pediatr Surg. 2007 Feb;17(1):23-8. [60] Chen CP, Chen SH, Chuang CY, Lee HC, Hwu YM, Chang PY, et al. Clinical and perinatal sonographic features of congenital adrenal cystic neuroblastoma: a case report with review of the literature. Ultrasound Obstet Gynecol. 1997 Jul;10(1):68-73. [61] Daneman A, Baunin C, Lobo E, Pracros JP, Avni F, Toi A, et al. Disappearing suprarenal masses in fetuses and infants. Pediatric radiology. 1997 Aug;27(8):675-81. [62] Granata C, Fagnani AM, Gambini C, Boglino C, Bagnulo S, Cecchetto G, et al. Features and outcome of neuroblastoma detected before birth. Journal of pediatric surgery. 2000 Jan;35(1):88-91. [63] Lee SY, Chuang JH, Huang CB, Hsiao CC, Wan YL, Ng SH, et al. Congenital bilateral cystic neuroblastoma with liver metastases and massive intracystic haemorrhage. The British journal of radiology. 1998 Nov;71(851):1205-7. [64] Nishi M, Miyake H, Takeda T, Yonemori H, Hanai J, Kikuchi Y, et al. A trial to discriminate spontaneous regression from non-regression cases during mass screening for neuroblastoma. Japanese journal of clinical oncology. 1994 Oct;24(5):247-51. [65] Schwarzler P, Bernard JP, Senat MV. Neonatal adrenal hemorrhage presenting as a multiloculated cystic neuroblastoma. Acta Radiol Diagn. 1986;27:3-10. [66] Masiakos PT, Gerstle JT, Cheang T, Viero S, Kim PC, Wales P. Is surgery necessary for incidentally discovered adrenal masses in children? Journal of pediatric surgery. 2004 May;39(5):754-8. [67] Oue T, Inoue M, Yoneda A, Kubota A, Okuyama H, Kawahara H, et al. Profile of neuroblastoma detected by mass screening, resected after observation without treatment: results of the Wait and See pilot study. Journal of pediatric surgery. 2005 Feb;40(2):359-63. [68] Tsuchida Y, Ikeda H, Iehara T, Toyoda Y, Kawa K, Fukuzawa M. Neonatal neuroblastoma: incidence and clinical outcome. Med Pediatr Oncol. 2003 Jun;40(6):391-3. [69] Sauvat F, Sarnacki S, Brisse H, Medioni J, Rubie H, Aigrain Y, et al. Outcome of suprarenal localized masses diagnosed during the perinatal period: a retrospective multicenter study. Cancer. 2002 May 1;94(9):2474-80. [70] Crofton PM, Squires N, Davidson DF, Henderson P, Taheri S. Reliability of urine collection pads for routine and metabolic biochemistry in infants and young children. European journal of pediatrics. 2008 Nov;167(11):1313-9. [71] Kellie SJ, Clague AE, McGeary HM, Smith PJ. The value of catecholamine metabolite determination on untimed urine collections in the diagnosis of neural crest tumours in children. Australian paediatric journal. 1986 Nov;22(4):313-5. [72] Pussard E, Neveux M, Guigueno N. Reference intervals for urinary catecholamines and metabolites from birth to adulthood. Clinical biochemistry. 2009 Apr;42(6):536-9. [73] Snow AB, Khalyfa A, Serpero LD, Capdevila OS, Kim J, Buazza MO, et al. Catecholamine alterations in pediatric obstructive sleep apnea: effect of obesity. Pediatric pulmonology. 2009 Jun;44(6):559-67. [74] Combaret V, Audoynaud C, Iacono I, Favrot MC, Schell M, Bergeron C, et al. Circulating MYCN DNA as a tumor-specific marker in neuroblastoma patients. Cancer research. 2002 Jul 1;62(13):3646-8.



[75] Combaret V, Bergeron C, Noguera R, Iacono I, Puisieux A. Circulating MYCN DNA predicts MYCN-amplification in neuroblastoma. J Clin Oncol. 2005 Dec 1;23(34):8919-20; author reply 20. [76] Gotoh T, Hosoi H, Iehara T, Kuwahara Y, Osone S, Tsuchiya K, et al. Prediction of MYCN amplification in neuroblastoma using serum DNA and real-time quantitative polymerase chain reaction. J Clin Oncol. 2005 Aug 1;23(22):5205-10. [77] Therasse P, Arbuck SG, Eisenhauer EA, Wanders J, Kaplan RS, Rubinstein L, et al. New guidelines to evaluate the response to treatment in solid tumors. European Organization for Research and Treatment of Cancer, National Cancer Institute of the United States, National Cancer Institute of Canada. Journal of the National Cancer Institute. 2000 Feb 2;92(3):205-16. [78] Ambros PF, Ambros IM, Kerbl R, Luegmayr A, Rumpler S, Ladenstein R, et al. Intratumoural heterogeneity of 1p deletions and MYCN amplification in neuroblastomas. Med Pediatr Oncol. 2001 Jan;36(1):1-4. [79] Stark DD, Moss AA, Brasch RC, deLorimier AA, Albin AR, London DA, et al. Neuroblastoma: diagnostic imaging and staging. Radiology. 1983 Jul;148(1):101-5. [80] Brody AS, Frush DP, Huda W, Brent RL. Radiation risk to children from computed tomography. Pediatrics. 2007 Sep;120(3):677-82. [81] Semelka RC, Armao DM, Elias J, Jr., Huda W. Imaging strategies to reduce the risk of radiation in CT studies, including selective substitution with MRI. J Magn Reson Imaging. 2007 May;25(5):900-9. [82] Siegel MJ, Jaju A. MR imaging of neuroblastic masses. Magnetic resonance imaging clinics of North America. 2008 Aug;16(3):499-513, vi. [83] Sofka CM, Semelka RC, Kelekis NL, Worawattanakul S, Chung CJ, Gold S, et al. Magnetic resonance imaging of neuroblastoma using current techniques. Magnetic resonance imaging. 1999 Feb;17(2):193-8. [84] Lonergan GJ, Schwab CM, Suarez ES, Carlson CL. Neuroblastoma, ganglioneuroblastoma, and ganglioneuroma: radiologic-pathologic correlation. Radiographics. 2002 Jul-Aug;22(4):911-34. [85] Brisse HJ, Aubert B. [CT exposure from pediatric MDCT: results from the 2007-2008 SFIPP/ISRN survey]. Journal de radiologie. 2009 Feb;90(2):207-15. [86] Goske MJ, Applegate KE, Boylan J, Butler PF, Callahan MJ, Coley BD, et al. The Image Gently campaign: working together to change practice. Ajr. 2008 Feb;190(2):273-4. [87] Boglino C, Martins AG, Ciprandi G, Sousinha M, Inserra A. Spinal cord vascular injuries following surgery of advanced thoracic neuroblastoma: an unusual catastrophic complication. Med Pediatr Oncol. 1999 May;32(5):349-52. [88] Ou P, Schmit P, Layouss W, Sidi D, Bonnet D, Brunelle F. CT angiography of the artery of Adamkiewicz with 64-section technology: first experience in children. Ajnr. 2007 Feb;28(2):216-9. [89] Goo HW, Choi SH, Ghim T, Moon HN, Seo JJ. Whole-body MRI of paediatric malignant tumours: comparison with conventional oncological imaging methods. Pediatric radiology. 2005 Aug;35(8):766-73. [90] Uhl M, Altehoefer C, Kontny U, Il'yasov K, Buchert M, Langer M. MRI-diffusion imaging of neuroblastomas: first results and correlation to histology. European radiology. 2002 Sep;12(9):2335-8. [91] Mendichovszky IA, Marks SD, Simcock CM, Olsen OE. Gadolinium and nephrogenic systemic fibrosis: time to tighten practice. Pediatric radiology. 2008 May;38(5):489-96; quiz 602-3. [92] Eisenhauer EA, Therasse P, Bogaerts J, Schwartz LH, Sargent D, Ford R, et al. New response evaluation criteria in solid tumours: revised RECIST guideline (version 1.1). Eur J Cancer. 2009 Jan;45(2):228-47. [93] Giammarile F, Boneu A, Edeline V, Lumbroso J, Siles S, Wioland M. Guide de realisation de la scintigraphie à la méta-iodobenzylguanidine (mIBG) en oncologie pédiatrique. Médecine nucléaire. 2000;24(1):35-42. [94] Khafagi FA, Shapiro B, Fischer M, Sisson JC, Hutchinson R, Beierwaltes WH. Phaeochromocytoma and functioning paraganglioma in childhood and adolescence: role of iodine 131 metaiodobenzylguanidine. European journal of nuclear medicine. 1991;18(3):191-8.



[95] Leung A, Shapiro B, Hattner R, Kim E, de Kraker J, Ghazzar N, et al. Specificity of radioiodinated MIBG for neural crest tumors in childhood. J Nucl Med. 1997 Sep;38(9):1352-7. [96] Shapiro B, Copp JE, Sisson JC, Eyre PL, Wallis J, Beierwaltes WH. Iodine-131 metaiodobenzylguanidine for the locating of suspected pheochromocytoma: experience in 400 cases. J Nucl Med. 1985 Jun;26(6):576-85. [97] Shulkin BL, Shapiro B. Current concepts on the diagnostic use of MIBG in children. J Nucl Med. 1998 Apr;39(4):679-88. [98] Sisson JC, Shulkin BL. Nuclear medicine imaging of pheochromocytoma and neuroblastoma. Q J Nucl Med. 1999 Sep;43(3):217-23. [99] Wieland DM, Wu J, Brown LE, Mangner TJ, Swanson DP, Beierwaltes WH. Radiolabeled adrenergi neuron-blocking agents: adrenomedullary imaging with [131I]iodobenzylguanidine. J Nucl Med. 1980 Apr;21(4):349-53. [100] Khafagi FA, Shapiro B, Gross MD. The adrenal gland. In: Maisey MN, Britton KE, Gilday DL, eds. Clinical Nuclear Medicine. 2nd ed: London Chapman & Hall 1989:271-91. [101] Wilson LM, Draper GJ. Neuroblastoma, its natural history and prognosis: a study of 487 cases. British medical journal. 1974 Aug 3;3(5926):301-7. [102] Young JL, Jr., Ries LG, Silverberg E, Horm JW, Miller RW. Cancer incidence, survival, and mortality for children younger than age 15 years. Cancer. 1986 Jul 15;58(2 Suppl):598-602. [103] Gelfand MJ. Meta-iodobenzylguanidine in children. Seminars in nuclear medicine. 1993 Jul;23(3):231-42. [104] Jacobs A, Delree M, Desprechins B, Otten J, Ferster A, Jonckheer MH, et al. Consolidating the role of *I-MIBG-scintigraphy in childhood neuroblastoma: five years of clinical experience. Pediatric radiology. 1990;20(3):157-9. [105] Lumbroso JD, Guermazi F, Hartmann O, Coornaert S, Rabarison Y, Leclere JG, et al. Meta-iodobenzylguanidine (mIBG) scans in neuroblastoma: sensitivity and specificity, a review of 115 scans. Progress in clinical and biological research. 1988;271:689-705. [106] Parisi MT, Greene MK, Dykes TM, Moraldo TV, Sandler ED, Hattner RS. Efficacy of metaiodobenzylguanidine as a scintigraphic agent for the detection of neuroblastoma. Investigative radiology. 1992 Oct;27(10):768-73. [107] Perel Y, Conway J, Kletzel M, Goldman J, Weiss S, Feyler A, et al. Clinical impact and prognostic value of metaiodobenzylguanidine imaging in children with metastatic neuroblastoma. J Pediatr Hematol Oncol. 1999 Jan-Feb;21(1):13-8. [108] Moyes J, McCready VR, Fullbrook AC. Neuroblastoma MIBG in its diagnosis and management: Springer-Verlag Berlin and Heidelberg GmbH & Co. K 1989. [109] Hoefnagel CA, De Kraker J, Valdes Olmos RA, Voute PA. [131I]MIBG as a first line treatment in advanced neuroblastoma. Q J Nucl Med. 1995 Dec;39(4 Suppl 1):61-4. [110] Lumbroso J, Hartmann O, Schlumberger M. Therapeutic use of [131I]metaiodobenzylguanidine in neuroblastoma: a phase II study in 26 patients. "Societe Francaise d'Oncologie Pediatrique" and Nuclear Medicine Co-investigators. J Nucl Biol Med. 1991 Oct-Dec;35(4):220-3. [111] Mairs RJ. Neuroblastoma therapy using radiolabelled [131I]meta-iodobenzylguanidine ([131I]MIBG) in combination with other agents. Eur J Cancer. 1999 Aug;35(8):1171-3. [112] Gordon I, Peters AM, Gutman A, Morony S, Dicks-Mireaux C, Pritchard J. Skeletal assessment in neuroblastoma--the pitfalls of iodine-123-MIBG scans. J Nucl Med. 1990 Feb;31(2):129-34. [113] Solanki KK, Bomanji J, Moyes J, Mather SJ, Trainer PJ, Britton KE. A pharmacological guide to medicines which interfere with the biodistribution of radiolabelled meta-iodobenzylguanidine (MIBG). Nuclear medicine communications. 1992 Jul;13(7):513-21. [114] Piepsz A, Hahn K, Roca I, Ciofetta G, Toth G, Gordon I, et al. A radiopharmaceuticals schedule for imaging in paediatrics. Paediatric Task Group European Association Nuclear Medicine. European journal of nuclear medicine. 1990;17(3-4):127-9. [115] Stabin MG, Gelfand MJ. Dosimetry of pediatric nuclear medicine procedures. Q J Nucl Med. 1998 Jun;42(2):93-112. [116] Pintelon H, Jonckheer MH, Piepsz A. Paediatric nuclear medicine procedures: routine sedation or management of anxiety? Nuclear medicine communications. 1994 Aug;15(8):664-6.



[117] Gelfand MJ, Elgazzar AH, Kriss VM, Masters PR, Golsch GJ. Iodine-123-MIBG SPECT versus planar imaging in children with neural crest tumors. J Nucl Med. 1994 Nov;35(11):1753-7. [118] Rufini V, Fisher GA, Shulkin BL, Sisson JC, Shapiro B. Iodine-123-MIBG imaging of neuroblastoma: utility of SPECT and delayed imaging. J Nucl Med. 1996 Sep;37(9):1464-8. [119] Rufini V, Giordano A, Di Giuda D, Petrone A, Deb G, De Sio L, et al. [123I]MIBG scintigraphy in neuroblastoma: a comparison between planar and SPECT imaging. Q J Nucl Med. 1995 Dec;39(4 Suppl 1):25-8. [120] Bonnin F, Lumbroso J, Tenenbaum F, Hartmann O, Parmentier C. Refining interpretation of MIBG scans in children. J Nucl Med. 1994 May;35(5):803-10. [121] Lumbroso J, Giammarile F, Hartmann O, Bonnin F, Parmentier C. Upper clavicular and cardiac meta-[123I]iodobenzylguanidine uptake in children. Q J Nucl Med. 1995 Dec;39(4 Suppl 1):17-20. [122] Nakajo M, Shapiro B, Copp J, Kalff V, Gross MD, Sisson JC, et al. The normal and abnormal distribution of the adrenomedullary imaging agent m-[I-131]iodobenzylguanidine (I-131 MIBG) in man: evaluation by scintigraphy. J Nucl Med. 1983 Aug;24(8):672-82. [123] Squire R, Sarnacki S, Haider N. The outcome of surgical procedures at diagnosis in localised neuroblastoma, and the effect of chemotherapy on resectability: European Infant Neuroblastoma Study (abstr.). Pediatric blood & cancer. 2007;49(4):402. [124] Pranikoff T, Hirschl RB, Schnaufer L. Approach to cervicothoracic neuroblastomas via a trap-door incision. Journal of pediatric surgery. 1995 Apr;30(4):546-8. [125] Sauvat F, Brisse H, Magdeleinat P, Lopez M, Philippe-Chomette P, Orbach D, et al. The transmanubrial approach: a new operative approach to cervicothoracic neuroblastoma in children. Surgery. 2006 Jan;139(1):109-14. [126] Schwartz GJ, Brion LP, Spitzer A. The use of plasma creatinine concentration for estimating glomerular filtration rate in infants, children, and adolescents. Pediatric clinics of North America. 1987 Jun;34(3):571-90. [127] Hoover M, Bowman LC, Crawford SE, Stack C, Donaldson JS, Grayhack JJ, et al. Long-term outcome of patients with intraspinal neuroblastoma. Med Pediatr Oncol. 1999 May;32(5):353-9. [128] Katzenstein HM, Kent PM, London WB, Cohn SL. Treatment and outcome of 83 children with intraspinal neuroblastoma: the Pediatric Oncology Group experience. J Clin Oncol. 2001 Feb 15;19(4):1047-55. [129] De Bernardi B, Pianca C, Pistamiglio P, Veneselli E, Viscardi E, Pession A, et al. Neuroblastoma with symptomatic spinal cord compression at diagnosis: treatment and results with 76 cases. J Clin Oncol. 2001 Jan 1;19(1):183-90.

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