………………..……………………………………………………………………………………………………………………………………..

In Focus, RSV infection In Children

Octavio Ramilo

Outline

1. Importance of RSV disease in children

2. Current therapies for RSV bronchiolitis

3. Immune profiling and disease severity

4. New antivirals for RSV

5. Importance of RSV F protein

6. Preventive strategies for RSV disease

F

G

RSV: The Virus

Modified from Park JW, Barnett DW. South Med J 2002; 95:353–7; McLellan JS, Ray WC, Peeples ME. Curr Top Microbiol Immunol 2013; 372:83–104; Collins PL, Fearns R, Graham BS. Curr Top Microbiol Immunol 2013; 372:3–38

NS2NS1M2-1 M2-2

3´

G FSHMN P Lle

6 80 2 4 10 15 kb

tr

15,222 nt total

………………..……………………………………………………………………………………………………………………………………..………………..……………………………………………………………………………………………………………………………………..

Young infant with RSV bronchiolitis

Increase in RSV bronchiolitis hospitalizations

Garcia CG, et al. Pediatrics 2010

1,500

1,000

500

0

Bro

nch

ioli

tis h

osp

itali

sati

on

s

Study year

2002 2003 2004 2005 2006 2007

41%

59%

39%

61%

33%

67%

37%

63%

34%

66%

33%

67%

n=493 n=532

n=738n=848

n=901

n=1,077

RSV

Non-RSV

N= 4,589

*p<0.01; Chi-square test for trend

Predictors of bronchiolitis disease severityLogistic regression analyses

CCHD: complex congenital heart disease

*Odds ratios: 95% confidence intervals; p<0.0001

Garcia CG, et al. Pediatrics 2010

CLD

CCHD

Prematurity

1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5

Odds ratio*

O2 Requirement PICU Admission Mechanical Ventilation

1 2 3 4

Odds ratio*

1 2 3 4

Odds ratio*

RSV

Viral etiology of children hospitalized for bronchiolitis in the era of molecular diagnosis

Country N (%) pos Single virus

Multiple viruses

RSV single

RSV co-inf

RV single

hMPVsingle

PIV single

Israel1 490 (91%) 61% 30% 49% 22% 7% 2% 0%

France2 209 (95%) 86% 9% 46% 16% 7% 4% NA

Spain3 318 (86%) 62% 24% 43% 18% 12% 6% 1.5%

USA4 180 (93%) 70% 23% 43% 18% NA 7% 3%

France5 180 (96%) 72% 24% 53% 20% 12% 4% 3%

Netherlands6 142 (97%) 56% 41% 43% 30% 7% 3% 0.7%

1. Miron D, et al. Pediatr Infect Dis J 2010;29:e72. Marguet C, et al. PLoS One 2009;4:e45963. Calvo C, et al. Acta Paediatr 2010;99:883

4. Stempel HE, et al. Acta Paediatr 2009;98:1235. Richard N, et al. Pediatr Infect Dis J 2008;27:2136. Brand H, et al. Pediatr Pulmonol, 2011 (ahead of print)

Global RSV Disease Burden

RSV kills more children <1 year than any other pathogen except malaria

Malaria (11.8%)

RSV

Lozano, R et al. The Lancet. 2012; 380:2095-2128.

28-364 days of life

OR RSV Challenges October 2005

Treating the symptoms

▬ IV Fluids, secretions

▬ Oxygen

▬ Bronchodilators

Controlling the inflammatory response

▬ Corticosteroids

▬ Antileukotrienes

Directed against the virus

▬ Antivirals: ribavirin

Treatment for RSV Bronchiolitis

OR RSV Challenges October 2005

2 AGONISTS IN BRONCHIOLITIS

Kellner JD et al. Arch Pediatr Adolesc Med. 1996; 150:1166.

Favors Control

-2 -1 0 1

Gadomski Neb

Favors Treatment

Clinical Score

Meta-analysis ( n=15/89 )

Only randomized, placebo-

controlled trials included

Clinical scores, SaO2

and hospital outcomes

CONCLUSIONS: Modest clinical

improvement with 2 agonists

versus placebo in children with

mild to moderate bronchiolitis

Estimated cost treat 80%

children < 2yr US $ 37.5 million

Gadomski PO

Gadomski Neb

Gadomski PO

Klassen

Wang

Pooled Data set

OR RSV Challenges October 2005

STEROIDS AND BRONCHIOLITIS

Mean Difference in Days

-4 -3 -2 0-1 1 2 3

Dabbous

Springer

Roosevelt

Klassen

de Boeck

van Woensel

Pooled Analysis

Garrison MM et al. Pediatrics. 2000; 105(4):e44.

Favors PlaceboFavors Treatment

OR RSV Challenges October 2005

Dexamethasone in intubated RSV patients RSV loads in tracheal aspirates

* p<0.05, Wilcoxon rank-sum test

Study Days

2

3

4

5

6

Dexamethasone

Placebo

0 1 2 5 7

*

*

Buckingham, Jafri et al. Journal of Infectious Diseases 2002; 185: 1222-8

OR RSV Challenges October 2005

Dexamethasone in intubated RSV patients impact on disease severity

Buckingham, Jafri et al. Journal of Infectious Diseases 2002; 185: 1222-8

0

2

4

6

8

10

12

Ventilator Days ICU Days Hospital Days

Dexamethasone Placebo

RSV transmission

• Transmitted by droplets, large particles, and fomites

• RSV survives up to 6 hours on stethoscopes and up to 12 hours on hard, nonporous surfaces

• Over 50% of medical personnel infected when RSV is prevalent in community

• Nosocomial infection remains a serious problem

Blydt-Hansen T et al. Pediatr Infect Dis J. 1999;18:164-165; Hall CB et al. J Infect Dis. 1980;141:98-102; Hall CB. Clin

Infect Dis. 2000;31:590-596; Cohen B et al. Pediatr Infect Dis J. 2003;22:494-498. 2

8

MouseMAb 1129

Palivizumab

Human IgG

RSV-Specific RSV-Specific Non-RSV-Specific

-S-S- -S-S- -S-S-

Construction of palivizumab: A humanized monoclonal antibody

Randomized studies demonstrate efficacy of palivizumab in reducing RSV hospitalizations

1. The IMpact-RSV Study Group. Pediatrics 1998; 102:531–7;2. Feltes TF, et al. J Pediatr 2003; 143:532–40;3. Carbonell-Estrany X, et al. Pediatrics 2010; 125:e35–51

IMpact1 Cardiac2 Palivizumab vs motavizumab3

0

2

4

6

8

10

12

% o

f su

bje

cts

10.6%

4.8%

55%

9.7%

5.3%

45%

1.9% 1.4%

53/500 42/1012 63/648 34/639 62/3306 46/3329

Placebo Palivizumab Placebo Palivizumab Palivizumab Motavizumab

Variability in clinical presentation of RSV infections

URILRTI

Mild/Mod LRTI

Mod/SevereLRTI

Severe

98% 2%

Outpatients Inpatients

How do we explain the variability in clinical presentations?

URILRTI

Severe

Virus

Host

Complex Interplay

PBMCs

Gene Chips

Blood Reservoir / Migration

compartment

Factors released in the

bloodstream

PERIPHERAL

TISSUE

Diagnosis/

Prognosis

BLOOD IS A SOURCE OF TRANSCRIPTIONAL MARKERS

Transcriptional Profile in Children with RSV Bronchiolitis

Mann-Whitney <0.01, Benjamini MTC x1.25 fold change1

,37

0 (5

9 %

)9

47

(41

%)

14+45 Samples

2,3

17

Gen

esTraining Set

Dallas, TX (n=59)

Controls RSV

Test Set Dallas, TX (n=49)

Selected Gene Tree: RSV SIGNATURE (2,317)

Colored by: CTRL-RSV TEST MATCHED (Default Interpretation)

Gene List: QC MW P0.01 BENJX1.25(2,317) (2317)

1,3

70

(59

%)

94

7 (4

1 %

)

8+41 Samples

2,3

17

Gen

es

Ctrl RSV

Mejias et al PLoS Medicine (2013)

RSV Profile Validated in Two Independent Populations

Mejias et al PLoS Medicine 2013

Turku, Finland Columbus, Ohio

Is this profile specific for RSV?

Class Prediction Among Different Respiratory Viruses (86 Genes)

Selected Gene Tree: RSV-HRV-FLU CLASS PREDICTORS(86,K=6,P=0.05)

Colored by: RSV-HRV-FLU TRAINING(44+15+6) (Default Interpretation)

Gene List: RSV-HRV-FLU CLASS PREDICTORS(86,K=6,P=0.05) (86)

TRUE class

K-NN class

86

Pre

dic

tors

FLU RSV HRV

65 Samples

Training Set

63/65: 96%

Selected Gene Tree: 'RSV-HRV-FLU CLASS PREDICTORS(86,K=6,P=0.05)'

Colored by: RSV-HRV-FLU TEST(47+15+7) (Default Interpretation)

Gene List: RSV-HRV-FLU CLASS PREDICTORS(86,K=6,P=0.05) (86)

FLU RSV HRV

TRUE class

K-NN class

86

Pre

dic

tors

69 Samples

66/69: 95%

Test Set

Mejias et al PLoS Medicine 2013

Transcriptomics linking pathogenesis and clinical findings

Patient Genotype(DNA)

Expression Profiles(mRNA)

Clinical Disease

MICROBE

Modified from Ramilo and Mejias, Cell Host & Microbe 2009

Environment

Host

Factors

Other unknown

factors

Microbiome

Can we measure disease activity at the molecular level ?

Molecular Distance to Health (MDTH):

Score summarizes the global transcriptional perturbation in a way that can be applied in the

clinical context

*P<0.0001

Selected Gene Tree: QC KW P0.01 BENJ(1,536)

Colored by: CTRL-MILD-MODERATE-SEVERE AGE MATCHED (Default Interpretation)

Gene List: QCE KW P0.01 BENJ(1,536) (1536)

0

1000

2000

3000

4000

5000

6000

7000

8000

#12

61C

#12

82C

#12

85C

#-10

44

C

#-10

97

C

#-66

5R

SV

#-70

7R

SV

#-71

1R

SV

#-74

3R

SV

#-96

8R

SV

#12

05R

SV

#12

09R

SV

#12

35R

SV

#12

58R

SV

#12

69R

SV

#-95

6R

SV

#-99

5R

SV

#11

53R

SV

#11

94R

SV

#12

40R

SV

#12

68R

SV

#-66

7R

SV

#-68

9R

SV

#-72

1R

SV

#-65

3R

SV

#-68

7R

SV

#-97

9R

SV

#11

63R

SV

#11

78R

SV

#11

86R

SV

#11

96R

SV

#12

10R

SV

Ctrl (n=10) Mild (n=20) Moderate (n=17) Severe (n=16)

1,5

36

Gen

es

63 patients

RSV-induced immune dysregulation correlates with disease severity

Mejias et al PLoS Medicine (2013)

Is this clinically relevant?

Length of Hospitalization

r = 0.6p < 0.001

MDTH Score Correlates with RSV Disease Severity

Spearman Correlation

r = 0.4p < 0.001

Clinical Disease Severity Score*

MD

TH S

core

s

* Disease Severity Score: % Sp O2, respiratory rate, IVF, retractions, auscultation

Length of O2 Intake(days)

0 5 10 15 20 25

Mo

lec

ula

r D

isea

se

Sev

eri

ty S

co

re

0

2000

4000

6000

8000 r = 0.6p < 0.001

Days of Supplemental O2

Mejias et al PLoS Medicine (2013)

Antivirals for RSV infection

Experimental RSV infection in humans

PFU, plaque-forming units

Modified from DeVincenzo JP, et al. Am J Respir Crit Care Med 2010; 182:1305–14

Study Day

Me

an

Sym

pto

m S

co

re

Me

an

Vira

l Lo

ad

Lo

g1

0 P

FU

/m

L

Ino

cu

lation

RSV A (Memphis 37strain)

Healthy adult volunteers (N=35; aged 18–45 years)

Outcomes: a) Virologic: viral loads (culture/PCR)b) Clinical: score, mucus productionc) Host responses: cytokines

Antivirals for RSV in development

F, fusion protein; G, surface protein; L, large polymerase; N, nucleoprotein; P, phosphoprotein; SH, small hydrophobic protein

Modified from Mazur NI, et al.; Respiratory Syncytial Virus Network (ReSViNET). Lancet Respir Med 2015; 3:888–900

ALS-008176 (nucleoside analog) rapidly reduces RSV viral load and clinical disease severity in healthy adult volunteers

MD, maintenance dose

DeVincenzo JP, et al. N Engl J Med 2015; 373:2048–58

Symptom Score

Mea

n S

ymp

tom

Sco

re

8

5

4

3

2

1

0

6

7

0 21 3 4 5 6 7 8 9 10

6

5

4

3

2

1

00 21 3 4 5 6 7 8 9 10

Viral load

RSV

load

s lo

g 10

PFU

e/m

L

Placebo (n=12)375 mg (n=8)750 mg LD/150 mg MD (n=7)750 mg LD/500 mg MD (n=8)

N total=62 subjects

Start of study (12 hours after confirmation of RSV infection or 6 days after RSV inoculation, whichever occurred first)

New preventive strategies for RSV

Anti-RSV neutralizing monoclonal antibodies

Anti-RSV neutralizing antibodies

Mazur NI, et al.; Respiratory Syncytial Virus Network (ReSViNET). Lancet Respir Med 2015; 3:888–900

http://www.ablynx.com/uploads/home/d2a7b156-9ac8-44d9-8974-288a5e8e2aablynxcorporatepresentation_october2016_website.pdf

Phase I/IIa study of ALX-0171 in infants

Antiviral effect: time to undetectable virus in culture

F

G

RSV: The Virus

Modified from Park JW, Barnett DW. South Med J 2002; 95:353–7; McLellan JS, Ray WC, Peeples ME. Curr Top Microbiol Immunol 2013; 372:83–104; Collins PL, Fearns R, Graham BS. Curr Top Microbiol Immunol 2013; 372:3–38

NS2NS1M2-1 M2-2

3´

G FSHMN P Lle

6 80 2 4 10 15 kb

tr

15,222 nt total

F protein trimer-mediated fusion

Modified from McLellan JS, Ray WC, Peeples ME. Curr Top Microbiol Immunol 2013; 372:83–104

pre-HRA

HRB

Target cell

Virion

Pre-triggered(metastable)

Triggering

Target cell

Fold-back

6-helix bundle

Target cell

Virion

Post-triggered 6HB (stable)

Infection Syncytia

Membranefusion

Straightens out

Fusion peptide inserts

Target cell

Virion

Pre-hairpin intermediate

HRA

RSV F protein: Pre-fusion vs post-fusion

Modified from McLellan JS, et al. Science 2013; 340:1113–7

Pre-fusion F trimer

Pre-fusion F protomer

Post-fusion F protomer

Post-fusion F trimer

New antigenic site Ø

MEDI8897

Palivizumab(Site II)

(Site Ø)

McLellan JS, et al. Science 2013; 340:1113–7; Synagis Summary of Product Characteristics, as approved in January 2016 (EMEA/H/C/000257-N/0110); Dubovsky F. 5th Asia Pacific Global Summit and Expo on Vaccines and Vaccination, 27–29 July 2015, Brisbane, Australia

MEDI8897 blocks RSV F protein-mediated fusion

• Fully human IgG1 mAb derived from human B-cells

• Targets a unique antigenic site on pre-fusion RSV F (distinct from palivizumab)

Anti-RSV neutralizing mAbs

Wu H, et al. J Mol Biol 2007; 368:652–65; Wu H, et al. Curr Top Microbiol Immunol 2008; 317:103–23; McLellan JS, et al. Science 2013; 340:1113–7; Dubovsky F. 5th Asia Pacific Global Summit and Expo on Vaccines and Vaccination, 27–29 July 2015, Brisbane, Australia

-S-S-

Palivizumab

13 aa

Increased potency

-S-S-

Motavizumab

FCProlonged half-life

-S-S-

MEDI8897 (D25)

Binds PreF neutralizing activity

MEDI8897 mean serum concentration-time profiles

Griffin MP, et al. ESPID 2016; Brighton UK

(n=8)

(n=31)

(n=32)

87% infants in 50 mg group had serum concentrations > 6.8 μg/mL

target concentration for clinical efficacy on Day 150

RSV vaccines in clinical development

Modified from Mazur NI, et al.; Respiratory Syncytial Virus Network (ReSViNET). Lancet Respir Med 2015; 3:888–900

Maternal immunization RSV F nanoparticle vaccine(Phase 2: Maternal microneutralizing antibodies)

August A. RSV F Vaccine: Phase 2 Clinical Trial to Protect Infants via Maternal Immunization. XXI FIGO World Congress of Gynaecology and Obstetrics, 4–9 October 2015, Vancouver, Canada; oral presentation available at http://novavax.com

Active (vaccine) n=22Placebo n=28

Time from vaccination impacts placental antibody transfer (Phase 2 infants)

Modified from August A. RSV F Vaccine: Phase 2 Clinical Trial to Protect Infants via Maternal Immunization. XXI FIGO World Congress of Gynaecology and Obstetrics, 4–9 October 2015, Vancouver, Canada; oral presentation available at http://novavax.com

Assay SourceDelivery <30d

post-vacc (n=7)Delivery >30d post-

vacc (n=12)All

n=21

Anti-F IgG

Cord 7,227 8,659 8,153

Mother 12,979 6,993 8,594

Ratio C/M 0.6 1.2 0.9

Palivizumabcompetingantibodies (PCA)

Cord 177 195 189

Mother 303 178 213

Ratio C/M 0.6 1.1 0.9

• Maternal antibody peaks at 14 d after vaccination

• Period of placental transfer > 30 d maximizes Ab titers in infants

1. RSV is a major pathogen for young children worldwide

2. Current therapies for RSV are suboptimal

3. RSV induces a robust and specific RNA immune profile

4. Genomic score (MDTH) offers new objective tool to assess

disease severity in respiratory infections

5. Antivirals for RSV with different mechanisms of action are in

clinical development

6. Pipeline: new monoclonal Abs and several vaccines strategies

Summary

AcknowledgementsAsuncion Mejías

Hasan Jafri

Carla Garcia

Susana Chavez-Bueno

Ana Gomez

Dora Estripeaut

Evelyn Torres

Juanita Lozano

Alejandro Jordan

Juan P. Torres

Sara Mertz

Emilio Flaño

Mario Marcon

Gail Arthur

Mike Lawson

Dan Cohen

Blerta Dimo

Eleonora Bunsow

Grace Wentzel

Samantha Sharpe

Debby Bogaert

Wouter de Steenhuijsen

Bennett Smith

Nicolas Suarez

Romain Banchereau

Damien Chaussabel

Virginia Pascual

Jacques Banchereau

Derek Blankenship

Santu Heinonen

Tuomas Hartti

Oli Ruuskanen

Phuong Nguyen

Casey Glaser

Pablo Sanchez

Monica Ardura

Jessica Estep

Santiago Lopez

Raquel Cao

M.C. Suarez

Nate Kuppermann

Prashant Mahajan

Victoria Best

Sara Merzec

NIH: NIAID, NICHD, HRSA, OCHA

CHIRP Investigators

OR/Pneumonia/Nov2005

Thanks!