Executive Board of the Health Ministers’ Council for GCC States

Guideline on

Biosimilars

Version 1.0

Date issued

01/08/2016

Date of implementation

Document Control

Version Date Comments

1.0 08/2016

II

Contents

Subject Page

Preface II Abbreviations IV Glossary V Members of the Working Group VI

General Guidance for all Biosimilar Medicines 001

Chapter 1: Broad Outline 002

Chapter 2: Manufacturing and Quality Considerations 010

Chapter 3: Preclinical Issues 031

Chapter 4: Clinical Studies 051

Chapter 5: Other Important Issues 074

Labeling 075

Extrapolation 076

Interchangeability and Substitution 077

Stability 078

Storage conditions 085

Specific Guidance for Individual Biosimilar Medicines 088

Chapter 6: Insulins 089

Chapter 7: Interferons 095

Chapter 8: Erythropoietin 106

Chapter 9: Granulocyte Colony Stimulating Factor 114

Chapter 10: Growth Hormone 121

Chapter 11. Pharmaceutical Formulations of Biosimilars 127

Drug Master File Requirements for the Registration of Biosimilars (Follow-On-Proteins) 134

Guide for Registration Requirements 136

Explanation of some of the Requirements 143

References 183

III

Preface

Biosimilars are therapeutic biologicals that follow previously-approved innovative

biological medicinal products. They are not copies (generics), but are similar.

In this Guideline, we are using the term “biosimilar(s)” to denote proteins produced by

means of recombinant DNA technologies following the footsteps of an innovator

product after the expiration of the innovator‟s patent. They are complex and

heterogeneous in their nature; hence they are not considered generics, but as closely

similar to the innovator‟s drug as possible. They can never be “exact copies” as in other

chemical generics due to the inherent nature of protein synthesis and due to possible

difference in gene sequence, vector, cell expression system, cell line growth media,

method of gene expression, bioreactor conditions, operating conditions, binding and

elution conditions for purification, and reagents and reference standards.

The global need for biosimilars is evident. However, they represent a very complex

issue that requires an appreciation of the complicated science behind drug manufacture

and quality control, since patient safety is paramount. Biosimilar versions of proprietary

drugs, when not properly regulated, might pose safety concerns, though the issues of

safety should not be confused with issues of cost. Biosimilar drugs, in particular, should

be traceable. Adequate post-marketing surveillance should be in place for all drugs,

both the innovator‟s drugs and the copies.

Since the manufacture and regulation of biosimilars are complex issues, it has been

proposed that a step-by-step approach, building knowledge in small, easy to assimilate

steps and working towards a greater understanding of complex issues, would be most

effective.

IV

We acknowledge that the content of this document was assembled through extensive

search and research of the European Medicines Agency (EMEA) Guidelines, the

International Conference on Harmonization (ICH) Guidelines and other resources

including published, peer reviewed articles. Excerpts from their guidelines were used as

written by them. We are grateful to both EMEA and ICH for making their efforts

available to us on a public domain and for allowing others to use them and follow suit.

The cornerstone of good biosimilar products is that they must be comparable to the

reference products in terms of quality, efficacy and safety. Therefore, it is important to

emphasize that these essential aspects of biosimilars will be dealt with in all Chapters of

this document. Repetition of these issues in this document implies assertiveness, not

redundancy.

It is essential to stress that this document is ONLY a guideline. It may oversee

important issues or does not dwell on issues. New facts and technologies might emerge

as experience and science advance through time. The final approval of a submitted drug

master file and dossier is that of the Registration Committee of the

r e g u l a t o r y a u t h o r i t y . These Guidelines are to be revisited biannually for

evaluation, improvement, revision, and amendment.

V

Abbreviations

ANC Absolute Neutrophil Count

API Active Pharmaceutical Ingredient

AUC Area Under the Curve

BRP Biological Reference Product

CHO Chinese Hamster Ovary

Cmax Peak concentration of a drug in the blood

DMF Drug Master File (dossiers and applications)

EMEA European Medicines Agency

ICH International Conference on Harmonisation

(of Technical Requirements for Registration of

Pharmaceuticals for Human Use)

IM Intramuscular route of administration

INN International Non-proprietary Name

IV Intravenous route of administration

GCC Gulf Cooperation Council

MCB Master Cell Bank

PD Pharmacodynamics

PK Pharmacokinetics

PV Pharmacovigilance

rDNA Recombinant Deoxyribo-Nucleic Acid

RMP Reference Medicinal Product

SC Subcutaneous route of administration

SmPCs Summary of Product Characteristics

T1/2 Half-life

USFDA United States Food and Drug Administration

WCB Working Cell Bank

WHO World Health Organization

VI

Glossary

Applicant = The company or manufacturer who submits a drug master

file for marketing authorization (approval) of a biosimilar.

Biosimilars* = Similar biological medicinal products

= Follow-on-proteins

Drug master file = The file or files that contains all relevant information

required for the registration of a biosimilar. A drug master

file is composed of dossiers containing all relevant

information and an application. It is also referred to as

Common Technical Document (CTD)

Generic drug = A drug that is a product shown to be exactly the same as

an innovative drug, and is acceptable to be

interchangeable with the innovative drug.

Innovator = The company that first made the medicine available and

originally manufactured it. It is, sometimes, referred to as

Originator.

* We have used the term “biosimilars” to denote those biomedicines that are

proteins produced through recombinant DNA technology, and are similar (not a

generic) to an existing therapeutic protein produced through the same

recombinant DNA technology and not covered by a patent law or other

intellectual property rights. As such, some of the so-called “biosimilars” but are

not proteins produced through recombinant DNA technology (such as Heparin)

are not included in this Guideline.

VII

Working Group Members

Mohammed N. Al-Ahdal, BPharm, PhD - Chairman

Amal J. Fatani, BPharm, PhD - Member

Murad M. Al-Saggaf, BPharm, MSc - Member

Abdulmohsen H. Al-Rohaimi, DDS, PhD - Member

Hadeel F. Daghash, BPharm, PharmD - Member

Naser O. Aldosri, BPharm, MSc - Coordinator

Past Members

Fatimah Meraiki, BPharm, PharmD

Ali Al-Homaidan, BPharm, MSc

Khalid Al-Moaikel, BPharm

Apologies

Yousuf Al-Omi, BPharm, MSc (One meeting attended)

VIII

1

General Guidance for Biosimilar

Medicines

2

CHAPTER 1.0

Broad Outline

3

1.1 Introduction

Scientific advances and modern technologies have initiated new avenues for the

effective treatment of human diseases that were previously beyond the scope of the

classical pharmaceuticals containing chemically synthesized compounds as active

ingredients. The process for development of further innovative drugs using

biotechnological technologies and procedures is still growing. As of the writing of these

guidelines, it can be roughly estimated that at present more than 600 biotech

pharmaceuticals and vaccines are undergoing preclinical and clinical tests with the view

to offer new treatment options especially for neurodegenerative disorders and cancer.

Despite the advances in the methods and techniques available today for the full

characterization of biosimilars, limitations of some of these methods and techniques

prompt the initiation of a number of specific guidelines relevant to comparability of

safety and efficacy. Therefore, preclinical and clinical issues to be addressed within the

development programs of these biosimilars.

A company may choose to develop a new biosimilar claimed to be “similar” to the

“innovator,” which has been approved by EMEA and/or t h e l o c a l

r e g u l a t o r y a u t h o r i t y , on the basis of a complete Drug Master File (DMF)

in accordance with the requirements for the registration applications, based on the

demonstration of the similar nature of the two rDNA-produced therapeutic products.

Comparability studies are needed to generate evidence substantiating the similar

nature, in terms of quality, safety and efficacy, of the new biosimilar product and the

reference medicinal product (RMP), which should be approved and registered at

EMEA and/or the local regulatory authority.

The regulatory authority has the role of issuing specific guidelines concerning the

scientific data (manufacturing, preclinical and clinical) to be provided to

substantiate the claim of similarity in the registration file for any biological

medicinal product especially those containing highly purified, biotechnology-derived

proteins as active substances.

4

The scarcity of information and the scattered regulations and guidance have prompted

the GCC to develop these guidelines. These guidelines are compiled with the vision

that they will assist drug companies in the development of biosimilars, facilitate their

registration in the KSA, assist official DMF reviewers, and support the availability of

safe, efficacious and cheaper alternatives of rDNA-produced drugs to the public.

1.2 Scope

The purposes of these guidelines are:

(1) To introduce and outline the principles for biosimilars approval.

(2) To provide applicants with a “user guide,” showing regulations and relevant

scientific information in the various current international guidelines, in order to

substantiate the claim of similarity.

Companies developing biosimilars are invited to contact the local regulatory

authority to obtain further advice or prior approval on their development, particularly

during the clinical studies phase of their drug development.

1.3 Basic Principles

The success of a biosimilar development approach will depend on the ability to

characterize the product and, therefore, to demonstrate the similar nature of the

concerned product to its innovative predecessor. The following points must be

considered:

(1) By definition, biosimilars are not generic medicinal products. It is expected that

there may be subtle differences between biosimilars from different

manufacturers compared with innovator products, which may not be fully

apparent until greater experience in their use has been established. Therefore, in

order to support pharmacovigilance monitoring, the specific medicinal product

given to the patient should be clearly identified.

5

(2) Biosimilars exhibit a spectrum of molecular complexity. Therefore, they are

usually more strenuous to characterize than chemically-derived medicinal

products.

(3) Parameters such as the three-dimensional structure, the amount of acido-basic

variants or post-translational modifications such as the glycosylation profile can

be significantly altered by changes, which may initially be considered to be

“minor” in the manufacturing process.

(4) Due to the complexity of biosimilars, the comparability exercise will have to

be followed, rather than a demonstration of bioequivalence, which is

scientifically not appropriate for these types of products.

(5) The suitability of biosimilars depends on the state-of-the-art of analytical

procedures, the manufacturing processes employed, robustness and the

monitoring of quality aspects, as well as clinical and regulatory experiences.

(6) With regard to safety, efficacy and quality, biosimilars must fulfill and satisfy

the technical and product-class specific provisions guidelines and requirements

of the monographs of official Pharmacopoeias, in addition to any supplementary

requirements mentioned in international guidelines; in particular, GCC, ICH

and EMEA.

(7) When developing a biosimilar and carrying out the comparability exercise to

demonstrate that this product is similar to the innovator that is already

authorized, several relevant guidelines from EMEA and ICH should also be

taken into account. Most of these are referenced at the end of this document.

(8) It is acknowledged that a manufacturer developing a biosimilar may not have

full access to all necessary information of the RMP that could allow an

exhaustive comparison. Nevertheless, the level of detail must be such that firm

conclusions can be made.

6

1.4 Biosimilars reference medicinal product

The reference medicinal product (RMP) to be used for comparability purposes

throughout the process is required to be that of the innovator, which should have been

approved by EMEA, and preferably registered in the KSA. It is of particular importance

to state that the same RMP should be used for all parts of the DMF to be submitted to

the regulatory authority. The following points have to be met:

(1) The registered RMP should be used throughout the comparability program for

quality, safety and efficacy studies during the manufacturing, preclinical and

clinical phases of the biosimilar development, in order to allow the generation of

coherent data and conclusions.

(2) The active substance of a biosimilar must be similar, in molecular and biological

terms, to the active substance of the RMP.

(3) The final product (pharmaceutical form, strength and route of administration) of

the biosimilar should be the same as that of the RMP.

4) Comparability of the biosimilar product with the chosen RMP should be

addressed for both the final medicinal product and the active substance in the

medicinal product.

5) A clear scientific justification of the criteria followed to select the RMP should

be provided, with specific attention to its critical parameters and quality

attributes. The same RMP must be used during the undertaking of quality, safety

and efficacy studies.

6) It is necessary to conduct appropriate comparative tests at the level of the active

substance, in order to provide assurance that the molecular structure of the

active substance present in the biosimilar product can be considered comparable

to that in the RMP.

7

7) Quality aspects of a biosimilar are a fundamental element in the comparability

exercise versus the RMP, and should always be considered with regard to any

implications for safety and efficacy.

8) A stepwise approach should be undertaken to justify any differences in the

quality attributes of the biosimilar versus the RMP, in order to make a

satisfactory justification of the potential implications with regard to the safety

and efficacy of the biosimilar product.

9) It is not expected that the quality attributes in the biosimilar product and RMP

will be identical. For example, minor structural differences in the active

substance, such as variability in post-translational modifications may be

acceptable; however, it must be justified.

10) The comparability studies should be facilitated when the pharmaceutical form,

formulation, strength, etc. of the biosimilar product are the same as the RMP.

11) The brand name, pharmaceutical form, formulation and strength of the RMP

used in the comparability studies should be clearly identified.

12) The shelf life of the RMP should be considered when performing a

comparability studies, and its effect on the quality profile should be discussed.

1.5 Important issues for demonstrating biosimilarity

1.5.1 Comparability: Quality Safety and Efficacy

Just as for conventional chemical products, the prerequisites for marketing

authorization of a biosimilar are proof of quality, safety, and efficacy. Three issues

must be clearly addressed when assessing comparability between a biosimilar and the

RMP:

8

(1) Impact of the difference in manufacturing process (cell line, media used in

cell culture and purification processes.

Clearly the quality of a protein therapeutic depends highly on its manufacturing

process, formulation, and storage conditions. Validated in-process controls and

analytical methods, well-established reference standards, definitive drug

substance and product release specifications, and definitive stability

specifications are important issues that define safety and efficacy.

(2) Analytical characterization including potency determination by bioassays

and immunogenicity effect .

Sometimes analytical methods can elucidate differences between one

biopharmaceutical molecule and a copy of it, but such tests cannot provide

information about how it will affect patients. Some differences might be

irrelevant and harmless; others could provoke an immune reaction. Quite often

we do not know enough at the development stage to predict the behavior of a

biopharmaceutical molecule. So manufacturers of biosimilars will have to

perform tests to prove that their copies are just as safe and efficacious as the

RMPs. That is the only way to maintain patient safety.

(3) Similarity of preclinical and clinical study results.

1.5.2 Immunogenecity

Most therapeutic proteins are immunogenic despite the fact that their amino acid

sequences are identical (or nearly identical) to endogenous proteins. Formation of

antibodies often appears to have no clinical effect. In some cases, however, the clinical

effects are significant and cause more severe disease. Immunogenicity may result in a

loss of efficacy or enhanced function, altered biodistribution and pharmacokinetics,

increased active dose and toxicity, and interference with other diagnostics and

therapeutics. It may also cause hypersensitivity reactions, cross-neutralization of

endogenous substances, or changes in physiological functions. General factors that

influence the occurrence of an immune response include specific properties of an

immunogen, its molecular size and solubility, the route of administration, storage

9

methods, dose levels, type of packaging, and length of treatment. Furthermore,

immunogenicity depends on host factors such as genotype, age, concomitant diseases

associated with immune disregulation, or previous exposure to other therapeutic

proteins that might cause cross-reactivity. Most current physicochemical

characterization assays are inadequate for predicting protein immunogenicity. The only

satisfactory means so far of assessing relative immunogenicity of a biosimilar and its

chosen RMP counterpart is to compare them in a trial using the same assay, then

validate the results to show differences between products. In addition, risk assessment,

pharmacovigilance, and post-marketing studies are essential.

1.6 Case by case situations

Some biosimilar molecules are relatively simple and easy to identify and isolate. An

example is insulin, a peptide hormone with a relatively simple molecular structure that

is comparatively straightforward to identify and copy. Molecules such as interferon and

erythropoietin, which are glycosylated and/or contain different isoforms, are more

complex and rather difficult to identify. Since the process makes the product,

differences in a manufacturing process may lead to structural variations and different

pharmacodynamic and pharmacokinetic properties of a product, which has a different

safety and efficacy profile. For patient safety and for medicine availability, it is

important that appropriate characterization as well as preclinical and clinical evaluation

of a biosimilar product is carried out on a case by case basis in case when negligible

deviation from the RMP is evident, as results with current and evolving technology are

unpredictable. In addition, versions of a biosimilar made by different manufacturers

must be evaluated on a case by case basis.

.

10

CHAPTER 2.0

Manufacturing and Quality

Considerations

11

2.1 Introduction

Biosimilars development process, followed by validated manufacturing process, is the

start of the long pathway of a beneficial product. Expression system, fermentation or

culture process, purification process, drug substance (e.g., batch definition, pooling

strategy), formulation and filling, and general parameters affecting all manufacturing

steps (e.g., water quality, temperature, personnel) are all important elements of the

process. Any manufacturing change, even among batches, can produce process-related

impurities, culture/fermentation-derived impurities, purification-derived impurities, and

final product-related impurities. Thus, any deviation from the RMP manufacturing (the

innovator‟s) process may have a minor or major impact on product quality, safety,

and/or efficacy. Comparing results of in-process controls of intermediates can give a

first hint of such product changes. However, such comparisons would be possible only

for innovators because follow-on manufacturers will not have access to the innovator‟s

process intermediates. Deviant conformations, altered posttranslational modifications,

and different selections of subtype isoforms are potential consequences of process

deviations that could result in altered microheterogeneity. Substitution of a single amino

acid can alter biological activity. Patterns of absorption may be influenced by

formulation. Finally, the batch-to-batch variability is inevitable with biologic products

and contributes to comparability difficulties. In this Chapter, regulatory processes for

manufacturing will be addressed. However, full information will be referenced in

proper locations.

2.2 Technical issues

2.2.1 Manufacturing process

The biosimilar product is in part defined by its own specific manufacturing

process for both the drug substance and the final drug product. For a biosimilar

registration in KSA, it would be expected that the relevant international

guidelines of ICH, EMEA and USFDA have been considered by the

manufacturer through each stage of the drug development and production. The

12

ICH guidelines mentioned below are examples for international guidelines that

shall be followed for different development and manufacturing activities:

a) ICH Q5D shall be followed for cell line derivation, origin, source and

history of cells used, primary cell substrates, generation of the cell

substrate, cell banking, cell banking system, cell banking procedures,

general principles of characterization and testing of cell banks, tests of

identity of the cell banks, metazoan cells, microbial cells, tests of purity

of the cell banks, cell substrate stability, and all other issues in this

guidance.

b) ICH Q5B shall be followed for analysis for expression constructs, the

characterization of the expression system, and all other issues in this

guidance.

C) ICH Q5A (R1) shall be followed for cell line qualification (MCB and

WCB), cells at the limit of in vitro cell age used for production,

recommended viral detection and identification assays, virus testing in

unprocessed bulk, virus clearance and virus testing on purified bulk,

evaluation and characterization of virus clearance procedures, and all

other issues in this guidance.

d) ICH Q6B shall be followed for test procedures and acceptance criteria,

including tests and release specifications of the intermediate product

(drug substance specifications), tests and release of the finished product,

and all other issues in this guidance.

e) ICH Q5C shall be followed for stability testing of biotechnological

products and all other issues in this guidance.

2.2.2 Comparability Consideration

Biosimilars exhibit a spectrum of molecular complexity especially among the

various products of rDNA technology. They are usually more strenuous to

characterize than chemically-synthesized medicinal products.

13

2.2.2.1 Comparability studies should be performed during the

manufacturing stage comparing the biosimilar under

development to the RMP in all aspects including, but not limited

to, qualitative and quantitative composition of the final

preparation, strength and concentration, and formulation.

2.2.2.2 The manufacturing processes should be developed and optimized

taking into account information on all manufacturing processes

(expression system, cell substrate, culture, purification, viral

safety, excipients, formulation, primary packaging interactions,

etc.) and consequences on active substance characteristics.

2.2.2.3 A biosimilar product should be defined by the molecular

composition of the active substance resulting from its

manufacturing process, which may introduce its own process-

related impurities. It is the duty of the applicant to demonstrate

the consistency and robustness of his own process according to

existing ICH guidelines.

2.2.2.4 Formulation studies should be considered in the course of the

development of a suitable dosage form, even if excipients are

qualitatively and quantitatively the same as the RMP, and should

demonstrate the suitability of the proposed formulation with

regards to stability, compatibility (i.e. with excipients, diluents

and packaging materials), and integrity of the active substance

(both biologically and physico-chemically) for its intended

medicinal use.

2.2.2.5 Although it is acknowledged that the manufacturing process will

be optimized during development, it is advisable to generate the

required clinical data for the comparability study of product

14

manufactured with the final manufacturing process representing

the quality profile of the batches to be commercialized.

2.2.2.6 Viral safety should be ensured as directed in ICH-Q5A(R1).

2.3 Quality Aspects

2.3.1 Specifications

Specifications are critical quality standards that are proposed and justified by the

manufacturer and approved by regulatory authorities as conditions of approval

to ensure product quality and consistency. They should focus on those molecular

and biological characteristics found to be useful in ensuring the safety and

efficacy of the product.

2.3.1.1 The selection of tests to be included in the specifications is

product specific and should be defined according to the GCC

and ICH Q6B.

2.3.1.2 The rationale used to establish the proposed range of acceptance

criteria should be described.

2.3.1.3 Each acceptance criterion should be established and justified

based on data obtained from lots used in preclinical and/or

clinical studies, and by data from lots used for the manufacturing

process validation, data from stability studies, relevant

development data and data obtained from the quality, safety and

efficacy studies.

2.3.1.4 The setting of specifications should be supported by global

reasoning based on the applicant's experience of the biosimilar

product (quality, safety and efficacy) and own experimental

15

results obtained by testing the RMP. These data should

demonstrate that the limits set for a given test are not wider than

the range of variability of the representative RMP, unless

justified.

2.3.1.5 Accelerated stability studies of the RMP and of the biosimilar

product can be used to further define and compare stability

profiles.

2.3.2 Analytical characterization

Extensive state-of-the-art characterization studies should be applied to the

biosimilar and RMP, in parallel at both the active substance and the final

medicinal product levels, to demonstrate with a high level of assurance that the

quality of the biosimilar is comparable to the RMP. The direct comparison of

the active substance in the biosimilar product to a publicly available standard as

a reference is not appropriate to demonstrate comparability of the active

substance. This is because the manufacturer generally does not have access to

the active substance of the RMP and since this material may not have known

and defined safety and efficacy profiles. However, the use of these standards

plays an important role during development. In cases where the required

analyses of quality attributes of the active substance of the RMP can be made at

the finished product stage, testing of the isolated active ingredient may not be

needed.

2.3.3 Suitability and validation of analytical methods

Given the complexity of the molecule and its inherent heterogeneity, the set of

analytical techniques should represent the state-of-the-art methods in the

comparability exercise capable of detecting slight differences in all aspects

pertinent to the evaluation of quality. Methods used in the characterization

studies form an integral part of the quality data package and should be

appropriately qualified for the purpose of comparability. Before entering the

16

clinical trial(s) needed for comparability purposes, release tests should be

validated in accordance with international guidelines and accepted standards and

reference materials should be used for method qualification and validation.

2.3.4 Physicochemical properties

2.3.4.1 The physicochemical comparison comprises the evaluation of

physicochemical parameters and the structural identification of

product-related substances and impurities, including the

determination of degradation by performing stress and

accelerated stability studies.

2.3.4.2 A physicochemical characterization program should include

determination of the composition, physical properties, primary

and higher order structures of the active substance of the

biosimilar product.

2.3.4.3 An inherent degree of structural heterogeneity occurs in proteins

due to the biosynthetic process, therefore, the biosimilar product

can contain a mixture of post-translationally modified forms.

Appropriate efforts should be made to investigate and identify

these forms. The manufacturer should consider the concept of the

desired product (and its variants) as defined in ICH Q6B when

designing and conducting a comparability exercise. The

complexity of the molecular entity with respect to the degree of

molecular heterogeneity should also be considered

2.3.4.4 Depending on the physico-chemical properties of the molecule

(e.g. from primary to quaternary structure, length of the

sequence, post-translational modifications such as extent and

nature of glycosylation, N/C terminal modifications), it can

sometimes be difficult to define precisely the product and there is

a need to use an extensive series of analytical techniques

17

exploiting the various physicochemical properties (size, charge,

hydrophobicity, etc.) and biological activity of the molecule.

2.3.5 Immunochemical properties

For some drug substances or drug products, the protein molecule may need to be

examined using immunochemical procedures (e.g., ELISA, Western-blot)

utilizing antibodies which recognize different epitopes of the protein molecule.

Immunochemical properties of a protein may serve to establish its identity,

homogeneity or purity, or serve to quantify it.

2.3.6 Biological activity and properties

An important property is the biological activity that describes the specific ability

or capacity of a product to achieve a defined biological effect. A valid biological

assay (animals, cell culture, and/or ligand binding) to measure this activity shall

be used by the manufacturer.

2.3.6.1 The comparability studies should include an assessment of the

biological properties of the biosimilar product and the RMP.

Biological assays using different approaches to measure the

biological activity should be considered as appropriate (i.e.

depending on the biological properties of the product).

2.3.6.2 The results of relevant biological assay(s) should be provided and

expressed in units of activity calibrated against an international

or national reference standard. These assays should comply with

appropriate International Pharmacopoeia requirements for

biological assays.

2.3.6.3 Biological assay results can serve multiple purposes in the

confirmation of product quality attributes that are useful for

18

characterization and batch analysis and, in some cases, could

serve a link to clinical activity. The manufacturer should

consider the limitations of biological assays, such a high

variability, that might prevent detection of differences that occur

as a result of manufacturing process change.

2.3.6.4 In cases where the biological assay also serves as a complement

to physicochemical analysis, where physicochemical or

biological assays are not considered adequate to confirm that the

higher order structure is maintained, it might be appropriate to

conduct a preclinical or clinical study.

2.3.6.5 When changes are made to a product with multiple biological

activities, manufacturers should consider performing a set of

relevant functional assays designed to evaluate the range of

activities.

2.3.6.6 Importantly, a biological assay to measure the biological activity

of the product may be replaced by physicochemical tests only in

those instances where:

(1) Sufficient physicochemical information about the drug,

including higher-order structure, can be thoroughly

established by such physicochemical methods, and

relevant correlation to the demonstrated biological

activity.

(2) There exists a well-established manufacturing history.

2.3.7 Heterogeneity

2.3.7.1 An inherent degree of structural heterogeneity occurs in proteins

due to the biosynthetic processes used by living organisms to

produce them; therefore, the desired product can be a mixture of

anticipated post-translationally modified forms (e.g.,

glycoforms). These forms may be active and their presence may

19

have no deleterious effect on the safety and efficacy of the

product. The manufacturer should define the pattern of

heterogeneity of the desired product and demonstrate consistency

with that of the lots used in preclinical and clinical studies. If a

consistent pattern of product heterogeneity is demonstrated, an

evaluation of the activity, efficacy and safety (including

immunogenicity) of individual forms may not be necessary.

2.3.7.2 This can also be produced during manufacture and/or storage of

the drug substance or drug product. Since the heterogeneity of

these products defines their quality, the degree and profile of this

heterogeneity should be characterized, to assure lot-to-lot

consistency. When these variants of the desired product have

properties comparable to those of the desired product with

respect to activity, efficacy and safety, they are considered

product-related substances. When process changes and

degradation products result in heterogeneity patterns which differ

from those observed in the material used during preclinical and

clinical development, the significance of these alterations should

be evaluated.

2.3.8 Purity, impurities, and contaminants

The purity and impurity profiles of the active substance and the final medicinal

product should be assessed both qualitatively and quantitatively by a

combination of state of the art analytical procedures for both the RMP and the

biosimilar product.

2.3.8.1 The combination of analytical procedures selected should

provide data to evaluate whether a change in purity profile has

occurred in terms of the desired product. Where the change

results in the appearance of new impurities, the new impurities

should be identified and characterized when possible. Depending

on the impurity type and amount, it might be appropriate to

20

conduct preclinical studies to confirm that there is no adverse

impact on safety or efficacy of the drug product.

2.3.8.2 The determination of absolute, as well as relative purity, presents

considerable analytical challenges, and the results are highly

method-dependent. The purity of the drug substance and drug

product is assessed by a combination of analytical procedures.

2.3.8.3 Due to the unique biosynthetic production process and molecular

characteristics of biotechnological and biological products, the

drug substance can include several molecular entities or variants.

When these molecular entities are derived from anticipated post-

translational modification, they are part of the desired product.

When variants of the desired product are formed during the

manufacturing process and/or storage and have properties

comparable to the desired product, they are considered product-

related substances and not impurities. Individual and/or

collective acceptance criteria for product-related substances

should be set, as appropriate.

2.3.8.4 The manufacturer should also assess impurities which may be

present. Impurities may be either process or product-related.

They can be of known structure, partially characterized, or

unidentified. When adequate quantities of impurities can be

generated, these materials should be characterized to the extent

possible and, where possible, their biological activities should be

evaluated.

2.3.8.5 Process-related impurities encompass those that are derived from

the manufacturing process (cell substrates, host cell proteins, host

cell DNA, cell culture, inducers, antibiotics, or media

components) and from downstream processing. These are

expected to differ qualitatively from one process to another, and

21

therefore, the qualitative comparison of these parameters may not

be relevant in the comparability exercise. However, the impact of

these process-related impurities should be confirmed by

appropriate studies.

2.3.8.6 Product-related impurities (precursors and certain degradation

products) are molecular variants arising during manufacture

and/or storage, which do not have properties comparable to those

of the desired product with respect to activity, efficacy, and

safety. Further, the acceptance criteria for impurities should be

based on specific degradation pathways and potential post-

translational modifications of the individual proteins. They

should also be based on data obtained from lots used in

preclinical and clinical studies and manufacturing consistency

lots. New analytical technology and modifications to existing

technology are continually being developed and should be

utilized when appropriate. Acceptance criteria of impurities

should be clearly stated and according to international guidelines.

2.3.8.7 Contaminants in a product include all adventitiously introduced

materials not intended to be part of the manufacturing process,

such as chemical and biochemical materials (e.g., microbial

proteases), and/or microbial species. Contaminants should be

strictly avoided and/or suitably controlled with appropriate in-

process acceptance criteria or action limits for drug substance or

drug product specifications. New contaminants should be

evaluated to assess their potential impact on the quality, safety

and efficacy of the product.

2.4 Changes introduced during development and post registration

Manufacturers of biosimilar products frequently make changes to manufacturing

processes of products both during development and after approval. Reasons for such

22

changes include improving the manufacturing process, increasing scale, improving

product stability, and complying with changes in regulatory requirements. When

changes are made to the manufacturing process, the manufacturer generally evaluates

the relevant quality attributes of the product to demonstrate that modifications did not

occur that would adversely impact the safety and efficacy of the drug product. Such an

evaluation should indicate whether or not confirmatory preclinical or clinical studies are

appropriate. The objective of this section is to provide principles for assessing the

comparability of biosimilar products before and after changes are made in the

manufacturing process for the drug substance or drug product. The main emphasis is

on quality aspects. The principles adopted and explained in this section apply to:

(1) Products where manufacturing process changes are made by a single

manufacturer, including those made by a contract manufacturer, who can

directly compare results from analysis of pre-change and post-change product.

(2) Products where manufacturing process changes are made in development or for

which registration has been granted.

2.4.1 Consideration for manufacturing process changes

Any change or modification made to a production process may impact

on the quality, safety and efficacy of the finished product. Many

different types of changes can be introduced in a manufacturing process.

A non-exhaustive list is detailed below:

(1) Formulation and filling (excipients, equipment, change in the

manufacturing protocol, and shipping conditions).

(2) Finished product (batch definition, shelf-life, container and

closure system, shipping conditions, and storage conditions).

(3) Expression system: Master cell bank (new bank derived from an

existing cell line or an initial clone).

(4) Expression system: Working cell bank (a manufacturing change

in raw material fermentation or a new method of production and

storage conditions).

23

(5) Raw materials (new supplier, specifications, addition,

substitution, or elimination of raw materials, and medical

composition).

(6) Fermentation and culture process (cell culture conditions [pH,

oxygen, temperature, time, and mode], scale of fermentation and

cell culture, equipment, and change or additional fermentation

site and facility).

(7) Purification process: Column or resin change (size of the column,

supplier, and cleaning and storage conditions).

(8) Purification process: Reagents (new supplier, specifications,

replacement of raw materials).

(9) Purification process: Protocol (addition, substitution, or

elimination of a specific step).

(10) Purification process (scale of downstream process, change or

additional purification site/facility, equipment).

A well-defined manufacturing process with its associated process

controls assures that acceptable product is produced on a consistent

basis. Approaches to determining the impact of any process change will

vary with respect to the specific process, the product, the extent of the

manufacturer‟s knowledge of and experience with the process, and

development data generated. The manufacturer should confirm that the

process controls in the modified process provide at least similar or more

effective control of the product quality, compared to those of the RMP

original processes.

2.4.1.1 A careful consideration of potential effects of the

planned change on steps downstream and quality

parameters related to these steps is extremely

important (e.g., for acceptance criteria, in-process

specification, in-process tests, in-process hold

times, operating limits, and validation/evaluation,

if appropriate). This analysis will help identify

which tests should be performed during the

24

comparability exercise, which in-process or batch

release acceptance criteria or analytical

procedures should be re-evaluated and which steps

should not be impacted by the proposed change.

For example, analysis of intermediates might

suggest potential differences that should be

evaluated to determine the suitability of existing

tests to detect these differences in the product.

The rationale for excluding parts of the process

from this consideration should be justified.

2.4.1.2 While the process will change and the associated

controls might be redefined, the manufacturer

should confirm that pre-change and post-change

product are comparable. To support the

comparison it is often useful to demonstrate, for

example, that specific intermediates are

comparable or that the modified process has the

capability to provide appropriate levels of removal

for process- and product-related impurities,

including those newly introduced by the process

change. To support process changes for approved

products, data from commercial-scale batches are

generally indicated.

2.4.1.3 The process assessment should consider such

factors as the criticality of the process step and

proposed change, the location of the change and

potential for effects on other process steps, and the

type and extent of change. Information that can

aid this assessment is generally available from

several sources. The sources can include

knowledge from process development studies,

25

small scale evaluation/validation studies,

experience with earlier process changes,

experience with equipment in similar operations,

changes in similar manufacturing processes with

similar products, and literature.

2.4.1.4 The in-process controls, including critical control

points and in-process testing, should ensure that

the post-change process is well controlled and

maintains the quality of the product. Typically,

re-evaluation/re-validation activities for a simple

change might be limited to the affected process

step, if there is no evidence to indicate that there is

impact on the performance of subsequent

(downstream) process steps, or on the quality of

the intermediates resulting from the subsequent

steps. When the change considered affects more

than a single step, more extensive analysis of the

change and resultant validation might be

appropriate.

2.4.1.5 Demonstration of state of control with the

modified/changed manufacturing process might

include, but is not limited to, such items as:

(1) Establishment of modified specifications

for raw, source and starting materials, and

reagents.

(2) Appropriate bioburden and/or viral safety

testing of the post-change cell banks and

cells at the limit of in vitro cell age for

production.

(3) Adventitious agent clearance.

26

(4) Removal of product- or process-related

impurities, such as residual host cell DNA

and proteins.

(5) Maintenance of the purity level.

2.4.1.6 For approved products, an appropriate number of

post-change batches should be analyzed to

demonstrate consistent performance of the

process.

2.4.1.7 To support the analysis of the changes and the

control strategy, the manufacturer should prepare

a description of the change that summarizes the

pre-change and the post-change manufacturing

process and that clearly highlights modifications

of the process and changes in controls in a side-

by-side format.

2.4.2 Considerations for the comparability exercise

A determination of comparability can be based on a combination of

analytical testing, biological assays, and, in some cases, preclinical and

clinical data. If a manufacturer can provide assurance of comparability

through analytical studies alone, preclinical or clinical studies with the

post-change product are not warranted. However, where the relationship

between specific quality attributes and safety and efficacy has not been

established, and differences between quality attributes of the pre- and

post-change product are observed, it might be appropriate to include a

combination of quality, preclinical, and/or clinical studies in the

comparability exercise. To identify the impact of a manufacturing

process change, a careful evaluation of all foreseeable consequences for

the product should be performed. Generally, quality data on the pre- and

post-change product are generated, and a comparison is performed that

integrates and evaluates all data collected (such as routine batch

analyses, in-process control, process validation and evaluation data,

27

characterization and stability, if appropriate). The comparison of the

results to the predefined criteria should allow an objective assessment of

whether or not the pre- and post-change product are comparable.

Following the evaluation of the quality attributes, the manufacturer could

be faced with one or more of several outcomes.

2.4.2.1 Based on appropriate comparison of relevant

quality attributes, pre- and post-change product

are highly similar and considered comparable, i.e.,

no adverse impact on safety or efficacy profiles is

foreseen.

2.4.2.2 Although the pre- and post change product appear

highly similar:

(1) The analytical procedures used are not

sufficient to discern relevant differences

that can impact the safety and efficacy of

the product. The manufacturer should

consider employing additional testing

(further characterization) or preclinical

and/or clinical studies to reach a definitive

conclusion.

(2) Some differences have been observed in

the quality attributes of the pre-change and

post-change product, but it can be justified

that no adverse impact on safety or

efficacy profiles is expected, based on the

manufacturer‟s accumulated experience,

relevant information, and data. In these

circumstances, pre- and post-change

product can be considered comparable.

(3) Some differences have been identified in

the comparison of quality attributes and a

28

possible adverse impact on safety and

efficacy profiles cannot be excluded. In

such situations, the generation and analysis

of additional data on quality attributes are

unlikely to assist in determining whether

pre- and post-change product are

comparable. The manufacturer should

consider performing preclinical and/or

clinical studies.

2.4.2.3 Differences in the quality attributes are so

significant that it is determined that the products

are not highly similar and are therefore not

comparable. This outcome is not within the scope

of this document and is not discussed further. The

goal of the comparability exercise is to ascertain

that pre- and post-change drug product is

comparable in terms of quality, safety, and

efficacy. The extent of the studies necessary to

demonstrate comparability will depend on:

(1) The production step where the changes are

introduced.

(2) The potential impact of the changes on the

purity as well as on the physicochemical

and biological properties of the product,

particularly considering the complexity

and degree of knowledge of the product

(e.g., impurities, product- related

substances). For more details on the

impact of the change on each of those

properties, refer to ICH guideline Q5E.

(3) The availability of suitable analytical

techniques to detect potential product

29

modifications and the results of these

studies. More details can be found ICH

Q5E.

(4) The relationship between quality attributes

and safety and efficacy, based on overall

preclinical and clinical experience. More

details can be found in ICH Q5E.

2.4.3 Demonstration of comparability during development

2.4.3.1 During product development, it is expected that multiple

changes in the manufacturing process will occur that

could impact drug product quality, safety, and efficacy.

Comparability exercises are generally performed to

demonstrate that preclinical and clinical data generated

with pre-change product are applicable to post-change

product in order to facilitate further development and,

ultimately, to support the marketing authorization.

Comparability studies conducted for products in

development are influenced by factors such as the stage

of product development, the availability of validated

analytical procedures, and the extent of product and

process knowledge, which are limited at times due to the

available experience that the manufacturer has with the

process.

2.4.3.2 Where changes are introduced in development before

preclinical studies, the issue of assessing comparability is

not generally raised because the manufacturer

subsequently conducts preclinical and clinical studies

using the post-change product as part of the development

process. During early phases of preclinical and clinical

studies, comparability testing is generally not as extensive

as for an approved product. As knowledge and

30

information accumulate, and the analytical tools develop,

the comparability exercise should utilize available

information and will generally become more

comprehensive. Where process changes are introduced in

late stages of development and no additional clinical

studies are planned to support registration, the

comparability exercise should be as comprehensive and

thorough as the one conducted for an approved product.

Some outcomes of the comparability studies on quality

attributes can lead to additional preclinical or clinical

studies.

2.4.3.3 For a comparability exercise to occur during

development, appropriate assessment tools should be

used. Analytical procedures used during development

might not be validated, but should always be scientifically

sound and provide results that are reliable and

reproducible. Physicochemical and biological tests alone

might be considered inadequate to determine

comparability, and therefore, bridging preclinical and/or

clinical studies, as appropriate, might be needed. The

Comparability Bridging Study is a study performed to

provide preclinical or clinical data that allows

extrapolation of the existing data from the drug product

produced by the current process to the drug product from

the changed process.

31

CHAPTER 3.0

Preclinical Issues

32

3.1 Introduction

This Chapter addresses the general principles for the Preclinical (non-clinical)

development and assessment of registration applications of biosimilar products

containing recombinant proteins as active substance(s). It describes the preclinical

safety evaluation of biotechnology-derived pharmaceuticals in general and how they

relate to biosimilars. It also discusses issues regarding changes introduced by the

manufacturer of a given product.

The studies to be carried out should be comparative in nature and designed to detect

differences in response between the biosimilar product and the RMP. The

concentration will be on issues regarding biological activity, pharmacokinetics,

comparability, efficacy, safety, and immunogenicity.

Data from preclinical studies can provide useful pointers to potential therapeutic

differences in the biological properties of the biosimilar product compared with the

RMP.

In some few cases it may be appropriate to undertake few or even no preclinical

studies, but in most other situations a more detailed evaluation may be helpful. The

following points must be taken into consideration:

It is important to note that design of an appropriate preclinical study program

requires a clear understanding of the specific characteristics for each product.

(2) Results from the physicochemical and biological characterization studies should

be reviewed from the point-of-view of potential impact on efficacy and safety.

(3) Ongoing consideration should be given to the use of emerging technologies such

as in vitro tests involving for example real-time binding assays and to in vivo

techniques dealing with the developing genomic/proteomic microarray sciences.

These may present opportunities to detect minor changes in biological response

to pharmacologically active substances.

33

Preclinical studies may be used to highlight differences between the biosimilar

product and the RMP. Such studies may have a useful role in the preliminary

assessment of safety at one or more points in the development process, thus enabling

clinical studies to be undertaken with greater confidence. The following approach

may be considered and should be tailored to the specific product concerned on a case-

by-case basis:

(1) Quality findings

(a) Drug product – The type, nature, and extend of differences between the

biosimilar product and the RMP (as well as post-change product and the

pre-change product of the biosimilar, if any) with respect to quality

attributes including product-related substances, the impurity profile,

stability and excipients. For example, new impurities could warrant

toxicological studies for qualification.

(b) Results of the evaluation/validation studies on the new process including

the results of relevant in-process tests.

(c) Availability, capabilities and limitations of tests used for any

comparability studies.

(2) The nature and the level of knowledge of the product

(a) Product complexity, including heterogeneity and higher order structure –

Physicochemical and in vitro biological assays might not be able to

detect all differences in structure and/or function.

(b) Structure-activity relationship and strength of the association of quality

attributes with safety and efficacy.

(c) Relationship between the therapeutic protein and endogenous proteins

and the consequences for immunogenicity.

(d) Mode(s) of action (unknown vs. known, single vs. multiple active sites).

(e) Therapeutic indications/target patient groups - The impact of possible

differences can vary between the target populations covered by the

different indications.

(f) Posology, e.g., dosing regimen and route of administration, for instance,

repeated administration via subcutaneous route is more likely to be

associated immunogenicity than intravenous administration of a single

dose.

34

(g) The therapeutic window/dose-response curve.

(h) Previous experience, e.g., immunogenicity, safety. Experience with the

original product or with other products in the same class can be relevant.

Adherence to the principles presented in this Chapter is intended to improve the quality

and consistency of the preclinical safety data supporting the development o f

biopharmaceuticals.

The company should justify, in the DMF, its approach chosen during the development

of the biosimilar product. When an application for a biosimilar product, which refers

to a RMP, is submitted for a marketing authorization by an independent applicant

after the expiry of the data protection period, the following approach shall be applied:

(1) The company pursues to demonstrate that medicinal product is similar in terms

of quality, safety and efficacy the RMP. It may not be necessary to repeat all

safety and efficacy studies if the applicant can demonstrate that:

(a) It is possible to characterize the product in detail with respect to

physico-chemical properties and biological in vitro activity.

(b) Comparability can be shown from a chemical-pharmaceutical

perspective. During the whole comparability exercise, the same RMP

should be used.

(2) In case the RMP has more than one indication, the efficacy and safety of the

medicinal product claimed to be similar has to be justified or, if necessary,

demonstrated separately for each of the claimed indications. Justification will

depend on the clinical experience, the available literature data for the RMP,

whether or not the same receptor(s) are involved in all indications, the pre-

clinical data, and the immunogenicity profile.

(3) Safety data will be needed prior to marketing authorization, but also post-

marketing as possible differences might become evident later, even though

comparability with regard to efficacy has been shown.

(4) Preclinical safety studies to predispose for human clinical trials are important to

conduct and address, as explained in ICH topic M3(R1).

35

3.2 Issues regarding biological activity (in vitro and in vivo studies)

Considerations should be given to the use of emerging technologies. In vitro

techniques such as `real-time' binding or antigenicity assays may prove useful. The

development of microarray and other technologies may, in the future, present

opportunities for comparing minor changes in the in vivo biological response to

pharmacologically active substances by monitoring qualitative and quantitative

changes in the profile of biological samples. Interpretation of such studies is an

evolving science and the clinical relevance of these techniques remains to be

determined. However, useful information may be obtained, particularly since studies

would be designed to detect subtle differences in response to two similar products and

not just the response per se.

3.2.1 In vitro studies

Biological activity may be evaluated using in vitro assays to determine which

effects of the product may be related to clinical activity. The use of cell lines

and/or primary cell cultures can be useful to examine the direct effects on

cellular phenotype and proliferation. In vitro cell lines derived from mammalian

cells can be used to predict specific aspects of in vivo activity and to assess

quantitatively the relative sensitivity of various species (including human) to the

biopharmaceutical. Such studies may be designed to determine, for example,

receptor occupancy, receptor affinity, and/or pharmacological effects, and to

assist in the selection of an appropriate animal species for further in vivo

pharmacology and toxicology studies. Thus a battery of receptor-binding

studies, many of which may already be available from quality-related

bioassays, should normally be undertaken in order to assess if any alterations

in reactivity have occurred and to determine the likely causative factor(s).

Sufficient number of dilutions per curve is suggested to fully characterize the

concentration-response relationship. It is important that assays used for

comparability will have appropriate sensitivity to detect minute differences.

36

3.2.2 In vivo studies

In vivo studies to assess pharmacological activity, including defining

mechanism(s) of action, are often used to support the rationale of the proposed

use of the product in clinical studies.

Animal studies should be designed to maximize the information obtained and to

compare biosimilar product and RMP intended to be used in the clinical trials.

Such studies should be performed in a species known to be relevant and employ

state of the art technology. Due to the species specificity of many biotechnology-

derived pharmaceuticals, it is important to select relevant animal species for

toxicity testing.

If there are specific uncertainties or concerns regarding safety in vivo studies,

one or more suitable animal models may be considered. Greater reliance would

be placed on results from studies in a species shown for the innovator

(original/reference) product to be a good model for man. Animal studies should

be designed to maximize the information obtained and to compare the

biosimilar product and the RMP in the final formulation. In the general case and

where the model allows, consideration should be given to monitoring a number

of endpoints such as:

(a) Changes in pharmacodynamic parameters relevant to the clinical

application.

(b) Changes in pharmacokinetic parameters, e.g. clearance.

(C) Specifically designed toxicological observations (in-life and post-

mortem).

(d) The immune response, e.g. antibody titres, neutralizing capacity, cross-

reactivity.

(e) Areas of specific concern, e.g. respiratory, renal or cardiovascular

parameters.

37

It is worth noting that in vivo studies should be designed to detect differences

in response and not just the response per se. This would apply particularly in

specific areas such as immunogenicity.

In vivo toxicology studies should be performed in such a way that the

biosimilar product and the RMP are compared at several dose levels, to allow a

comparison of dose-response curves.

The duration of the studies should be sufficiently long to allow detection of any

differences in toxicity and/or immunogenicity between the biosimilar product and

the RMP, taking into account the intended duration of use.

Preclinical toxicity as determined in at least one repeat dose toxicity study,

including toxicokinetic measurements. If there are specific safety concerns, these

might be addressed by including relevant observations (i.e. local tolerance) in the

same repeat dose toxicity study.

Normally other routine toxicological studies such as safety pharmacology,

reproduction toxicology, mutagenicity and carcinogenicity studies are not

required for biosimilars, unless indicated by results of repeat dose studies.

In summary, the combined results from in vitro and in vivo studies assist in the

extrapolation of the findings to humans. A brief description of the above-

mentioned methodologies will be mentioned at the end of this Chapter expanding

on preclinical testing techniques of biotechnology-derived pharmaceuticals

(according to ICH topic S6)

3.3 Issues regarding pharmacokinetics and metabolism

3.3.1 Pharmacokinetics

It is difficult to establish uniform guidelines for pharmacokinetic studies

for biotechnology derived pharmaceuticals. Single and multiple dose

38

pharmacokinetics, toxicokinetics, and tissue distribution studies in

relevant species are useful; however, routine studies that attempt to

assess mass balance are not useful.

Differences in pharmacokinetics among animal species may have a

significant impact on the prediction of animal studies or on the

assessment of dose response relationships in toxicity studies.

Alterations in the pharmacokinetic profile due to immune-mediated

clearance mechanisms may affect the kinetic profiles and the

interpretation of the toxicity data. For some products there may also be

inherent, significant delays in the expression of pharmacodynamic

effects relative to the pharmacokinetic profile (e.g., cytokines) or there

may be prolonged expression of pharmacodynamic effects relative to

plasma levels.

3.3.2 Metabolism

The expected consequence of metabolism of biotechnology-derived

pharmaceuticals is the degradation to small peptides and individual

amino acids. Therefore, the metabolic pathways are generally

understood. Classical biotransformation studies as performed for

pharmaceuticals are not needed. Understanding the behavior of the

biopharmaceutical in the biologic matrix, (e.g., plasma, serum, cerebral

spinal fluid) and the possible influence of binding proteins is important

for understanding the pharmacodynamic effect.

3.4 Issues regarding efficacy and safety

In this section, issues are mentioned that should be considered when drafting

and justifying a development plan to address the efficacy and safety of the both

the biosimilar product and any possible change thereafter. Depending on the

39

product and the (anticipated/recorded) change, the information package may

consist of preclinical and/or clinical data. Applications should be accompanied

by an assessment of the potential impact of the variation on efficacy and safety.

The rationale behind the development plan should be outlined and justified.

It is important to note that safety issues require a clear understanding of the

product characteristics in order to design suitable study protocols. Results from

the physicochemical characterization studies should be reviewed from the point

of view of potential impact on efficacy and safety (in vitro and in vivo

biological activity, metabolism, kinetics, immunogenicity, and other necessary

parameters). The methods chosen to detect heterogeneity between the biosimilar

product and the RMP should be described. Sufficient information, and cross-

referencing to other sections, should be supplied in the preclinical section to

justify the approach taken in subsequent studies.

The primary goals of preclinical safety evaluation are:

(1) To identify an initial safe dose and subsequent dose escalation schemes

in humans.

(2) To identify potential target organs for toxicity and for the study of

whether such toxicity is reversible.

(3) To identify safety parameters for clinical monitoring.

Safety concerns may arise from the presence of impurities or contaminants. It is

preferable to rely on purification processes to remove impurities and

contaminants rather than to establish a preclinical testing program for their

qualification. In all cases, the product should be sufficiently characterized to

allow an appropriate design of preclinical safety studies.

There are potential risks associated with host cell contaminants derived from

bacteria, yeast, insect, plants, and mammalian cells. The presence of cellular host

contaminants can result in allergic reactions and other immunopathological

effects. The adverse effects associated with nucleic acid contaminants are

theoretical but include potential integration into the host genome. For products

40

derived from insect, plant and mammalian cells, or transgenic plants and animals,

there may be an additional risk of viral infections. Preclinical safety testing

should consider:

(1) Selection of the relevant animal species.

(2) Age.

(3) Physiological state.

(4) The manner of delivery, including dose, route of administration, and

treatment regimen.

(5) Stability of the test material under the conditions of use.

3.5 Issues regarding Immunogenicity

Many biotechnology-derived pharmaceuticals intended for human are

immunogenic in animals. Therefore, measurement of antibodies associated with

administration of these types of products should be performed when conducting

repeated dose toxicity studies in order to aid in the interpretation of these studies.

Antibody responses should be characterized (e.g., titer, number of responding

animals, neutralizing or non-neutralizing), and their appearance should be

correlated with any pharmacological and/or toxicological changes.

Specifically, the effects of antibody formation on

pharmacokinetic/pharmacodynamic parameters, incidence and/or severity of

adverse effects, complement activation, or the emergence of new toxic effects

should be considered when interpreting the data. Attention should also be paid to

the evaluation of possible pathological changes related to immune complex

formation and deposition.

The detection of antibodies should not be the sole criterion for the early

termination of a preclinical safety study or modification in the duration of the

study design, unless the immune response neutralizes the pharmacological and/or

toxicological effects of the biosimilar in a large proportion of the animals. In

41

most cases, the immune response to biosimialrs (and all biopharmaceuticals) is

variable, like that observed in humans. If the interpretation of the data from the

safety study is not compromised by these issues, then no special significance

should be ascribed to the antibody response.

The induction of antibody formation in animals is not predictive of a potential for

antibody formation in humans. Humans may develop serum antibodies against

humanized proteins, and frequently the therapeutic response persists in their

presence. The occurrence of severe anaphylactic responses to recombinant

proteins is rare in humans. In this regard, the results of guinea pig anaphylaxis

tests, which are generally positive for protein products, are not predictive for

reactions in humans; therefore, such studies are considered of little value for the

routine evaluation of these types of products.

3.6 Issues regarding comparability

Two situations are indicated in which comparability becomes an issue:

(1) When a product is claimed to be similar to the RMP after the

expiry of the data protection period (new application procedure).

(2) When a change is introduced in the manufacturing process of the

biosimilar product (either before or after the granting of a

marketing authorization [variation procedure]).

In either case the company will have to demonstrate or justify that th e

biosimilar product and RMP have similar profiles in terms of quality,

safety and efficacy. This might be a sequential process, beginning with

quality studies (partial or comprehensive) and supporte d, as necessary, by

preclinical and/or clinical bridging studies to provide useful signals of

potential therapeutic differences.

The information obtained from preclinical data and timing of submission of

these data will have to be judged on a case by case basis and will be guided

by:

42

(1) The extent to which the product may be characterized.

(2) The nature of the changes in the `new' product compared to the

RMP.

(3) The observed/potential differences between the two products.

3.7 Issues regarding manufacturing changes

Manufacturers frequently make changes to manufacturing processes of their

biotechnological/biological (biosimilar) products, both pre- and post-approval.

The marketing authorization holder will have to demonstrate or justify that the

product have comparable quality, safety and efficacy to the innovator‟s product.

In general, the product that is used in the definitive pharmacology and toxicology

studies should be comparable to the product proposed for the initial clinical

studies. However, it is appreciated that during the course of development

programs, changes normally occur in the manufacturing process in order to

improve product quality and yields. The potential impact of such changes for

extrapolation of the animal findings to humans should be considered. The

comparability of the test material during a development program should be

demonstrated when a new or modified manufacturing process or other significant

changes in the product or formulation are made in an ongoing development

program. Comparability can be evaluated on the basis of biochemical and

biological characterization (identity, purity, stability, and potency).

The use of one or more assay methods should be addressed on a case-by-case

basis and the scientific rationale should be provided. One validated method is

usually considered sufficient. For example, quantitation of TCA-precipitable

radioactivity following administration of a radiolabeled protein may provide

adequate information, but a specific assay for the analyte is preferred. Ideally the

assay methods should be the same for animals and humans. The possible

influence of plasma binding proteins and/or antibodies in plasma/serum on the

assay performance should be determined.

43

In some cases additional studies may be needed (pharmacokinetics,

pharmacodynamics and/or safety). The scientific rationale for the approach taken

should be provided.

3.8 Preclinical testing techniques of biosimilars

The following is a brief description of the above-mentioned methodologies, in the

event that more information is required. It is based on ICH topic S6 guidelines,

which addresses issues related to biotechnology-derived pharmaceuticals.

3.8.1 Animal species/model selection

The biological activity together with species and/or tissue specificity of

many biosimilars often preclude standard toxicity testing designs in

commonly used species (e.g., rats and dogs). Safety evaluation programs

should include the use of relevant species.

A relevant species is one in which the test material is pharmacologically

active due to the expression of the receptor or an epitope. A variety of

techniques (e.g., immunochemical or functional tests) can be used to

identify a relevant species. Knowledge of receptor/epitope distribution

can provide greater understanding of potential in vivo toxicity.

Safety evaluation programs should normally include two relevant

species. However, in certain justified cases one relevant species may

suffice (e.g., when only one relevant species can be identified or where

the biological activity of the biosimilar is well understood).

In addition, even where two species may be necessary to characterize

toxicity in short term studies, it may be possible to justify the use of only

one species for subsequent long term toxicity studies (e.g., if the toxicity

profile in the two species is comparable in the short term).

44

Toxicity studies in non-relevant species may be misleading and are

discouraged. When no relevant species exists, the use of relevant

transgenic animals expressing the human receptor or the use of

homologous proteins should be considered. The information gained from

use of a transgenic animal model expressing the human receptor is

optimized when the interaction of the product and the humanized

receptor has similar physiological consequences to those expected in

humans.

While useful information may also be gained from the use of

homologous proteins, it should be noted that the production process,

range of impurities/contaminants, pharmacokinetics, and exact

pharmacological mechanism(s) may differ between the homologous

form and the product intended for clinical use. Where it is not possible to

use transgenic animal models or homologous proteins, it may still be

prudent to assess some aspects of potential toxicity in a limited toxicity

evaluation in a single species, e.g., a repeated dose toxicity study of ≤ 14

days duration that includes an evaluation of important functional

endpoints (e.g., cardiovascular and respiratory).

In recent years, there has been much progress in the development of

animal models that are thought to be similar to the human disease. These

animal models include induced and spontaneous models of disease, gene

knockout(s), and transgenic animals. These models may provide further

insight, not only in determining the pharmacological action of the

product, pharmacokinetics, and dosimetry, but may also be useful in the

determination of safety (e.g., evaluation of undesirable promotion of

disease progression). In certain cases, studies performed in animal

models of disease may be used as an acceptable alternative to toxicity

studies in normal animals. The scientific justification for the use of these

animal models of disease to support safety should be provided.

45

Animal models of disease may be useful in defining toxicity endpoints,

selection of clinical indications, and determination of appropriate

formulations, route of administration, and treatment regimen. It should

be noted that with these models of disease there is often a paucity of

historical data for use as a reference when evaluating study results.

Therefore, the collection of concurrent control and baseline data is

critical to optimize study design.

3.8.2 Number/gender of animals

The number of animals used per dose has a direct bearing on the ability

to detect toxicity. A small sample size may lead to failure to observe

toxic events due to observed frequency alone regardless of severity. The

limitations that are imposed by sample size, as often is the case for non-

human primate studies, may be in part compensated by increasing the

frequency and duration of monitoring. Both genders should generally be

used or justification given for specific omissions.

3.8.3. Administration/dose selection

The route and frequency of administration should be as close as possible

to that proposed for clinical use. Consideration should be given to

pharmacokinetics and bioavailability of the product in the species being

used, and the volume which can be safely and humanely administered to

the test animals. For example, the frequency of administration in

laboratory animals may be increased compared to the proposed schedule

for the human clinical studies in order to compensate for faster clearance

rates or low solubility of the active ingredient. In these cases, the level of

exposure of the test animal relative to the clinical exposure should be

defined.

Consideration should also be given to the effects of volume,

concentration, formulation, and site of administration. The use of routes

of administration other than those used clinically may be acceptable if

46

the route must be modified due to limited bioavailability or to

size/physiology of the animal species.

Dosage levels should be selected to provide information on a dose-

response relationship, including a toxic dose and a no observed adverse

effect level. For some classes of products with little to no toxicity, it may

not be possible to define a specific maximum dose. In these cases, a

scientific justification of the rationale for the dose selection and

projected multiples of human exposure should be provided. To justify

high dose selection, consideration should be given to the expected

pharmacological/physiological effects, availability of suitable test

material, and the intended clinical use.

Where a product has a lower affinity to or potency in the cells of the

selected species than in human cells, testing of higher doses may be

important. The multiples of the human dose that are needed to determine

adequate safety margins may vary with each class of biotechnology-

derived pharmaceutical and its clinical indication(s).

3.8.4 Toxicity studies

As mentioned earlier, preclinical toxicity should be determined in at

least one repeat dose toxicity study, including toxicokinetic

measurements. Toxicokinetic measurements should include

determination of antibody titres, cross reactivity and neutralizing

capacity. If there are specific safety concerns, these might be addressed

by including relevant observations (i.e. local tolerance) in the same

repeat dose toxicity study.

3.8.4.1 Local tolerance studies

Local tolerance should be evaluated. The formulation

intended for marketing should be tested; however, in

certain justified cases, the testing of representative

47

formulations may be acceptable. In some cases, the

potential adverse effects of the product can be evaluated

in single or repeated dose toxicity studies thus obviating

the need for separate local tolerance studies.

3.8.4.2 Single dose toxicity studies

Single dose studies may generate useful data to describe

the relationship of dose to systemic and/or local toxicity.

These data can be used to select doses for repeated dose

toxicity studies. Information on dose-response

relationships may be gathered through the conduct of a

single dose toxicity study, as a component of

pharmacology or animal model efficacy studies. The

incorporation of safety pharmacology parameters in the

design of these studies should be considered.

3.8.4.3 Repeated dose toxicity studies

When feasible, these studies should include

toxicokinetics. A recovery period should generally be

included in study designs to determine the reversal or

potential worsening of pharmacological/toxicological

effects, and/or potential delayed toxic effects.

For biosimilars that induce prolonged

pharmacological/toxicological effects, recovery group

animals should be monitored until reversibility is

demonstrated. The duration of repeated dose studies

should be based on the intended duration of clinical

exposure and disease indication. This duration of animal

dosing has generally been 1-3 months for most

biotechnology-derived pharmaceuticals. For biosimilars

intended for short-term use (e.g., ≤ to 7 days) and for

48

acute life-threatening diseases, repeated dose studies up to

two weeks duration have been considered adequate to

support clinical studies as well as marketing

authorization.

For those biosimilars intended for chronic indications,

studies of 6 months duration have generally been

appropriate although in some cases shorter or longer

durations have supported marketing authorizations. For

biosimilars intended for chronic use, the duration of long

term toxicity studies should be scientifically justified.

3.8.4.4 Reproductive performance and developmental toxicity

studies

Reproductive/developmental toxicity studies is not

needed with biosimilars, dependent upon the product,

clinical indication, and intended patient population.

There may be extensive public information available

regarding potential reproductive and/or developmental

effects of a particular class of compounds (e.g.,

interferons) where the only relevant species is the non-

human primate. In such cases, mechanistic studies

indicating that similar effects are likely to be caused by a

new but related molecule, may obviate the need for

formal reproductive/developmental toxicity studies. In

each case, the scientific basis for assessing the potential

for possible effects on reproduction/development should

be provided.

49

3.8.4.5 Genotoxicity studies

The range and type of genotoxicity studies routinely

conducted for pharmaceuticals are not applicable to

biosimilars and, therefore, are not needed. Moreover, the

administration of large quantities of peptides/proteins

may yield results that cannot be interpreted. It is not

expected that these substances would interact directly

with DNA or other chromosomal material.

With some biopharmaceuticals there is a potential

concern about accumulation of spontaneously mutated

cells (e.g., via facilitating a selective advantage of

proliferation) leading to carcinogenicity. The standard

battery of genotoxicity tests is not designed to detect

these conditions. Alternative in vitro or in vivo models to

address such concerns may have to be developed and

evaluated.

Studies in available and relevant systems, including

newly developed systems, should be performed in those

cases where there is cause for concern about the product

(e.g., because of the presence of an organic linker

molecule in a conjugated protein product). The use of

standard genotoxicity studies for assessing the genotoxic

potential of process contaminants is not considered

appropriate. If performed for this purpose, however, the

rationale should be provided.

3.8.4.6. Carcinogenicity studies

Standard carcinogenicity bioassays are generally

inappropriate for biosimilars. However, product-specific

50

assessment of carcinogenic potential may still be needed

depending upon duration of clinical dosing, patient

population and/or biological activity of the product (e.g.,

growth factors, immunosuppressive agents, etc..) When

there is a concern about carcinogenic potential a variety

of approaches may be considered to evaluate risk.

Products that may have the potential to support or induce

proliferation of transformed cells and clonal expansion

possibly leading to neoplasia should be evaluated with

respect to receptor expression in various malignant and

normal human cells that are potentially relevant to the

patient population under study. The ability of the product

to stimulate growth of normal or malignant cells

expressing the receptor should be determined. When in

vitro data give cause for concern about carcinogenic

potential, further studies in relevant animal models may

be needed. Incorporation of sensitive indices of cellular

proliferation in long term repeated dose toxicity studies

may provide useful information.

It is known that conventional approaches to toxicity testing of pharmaceuticals

may not be appropriate for biopharmaceuticals due to the unique and diverse

structural and biological properties of the latter that may include species

specificity, immunogenicity, and unpredicted pleiotropic activities. Normally

other routine toxicological studies such as safety pharmacology,

reproduction toxicology, mutagenicity and carcinogenicity studies are not

required for similar biological medicinal products, unless indicated by results of

repeat dose studies.

51

CHAPTER 4.0

Clinical Studies

52

4.1 Introduction

This Chapter lays down the general principles for the clinical studies necessary for

biosimilars as well as for changes introduced in the manufacturing process of a given

biosimilar during development and post-authorization. The general principles for the

clinical development and assessment of the application for registering a biosimilar are

addressed. Immunogenicity and risk management are discussed. The requirements of

the clinical studies depend on the existing knowledge about the RMP and the claimed

therapeutic indication(s). Comparability exercise must be conducted.

It is recommended to generate the required clinical data for the comparability study

with the test product as produced with the final manufacturing process and, therefore,

representing the quality profile of the batches to become commercialized. Any

deviation from this recommendation should be justified and supported by adequate

additional data.

4.2 Demonstration of clinical comparability

The clinical comparability exercise is a stepwise procedure that should begin

with pharmacokinetic (PK) and pharmacodynamic (PD) studies followed by

clinical efficacy and safety trial(s) or, in certain cases,

pharmacokinetic/pharmacodynamic (PK/PD) studies for demonstrating clinical

comparability.

4.2.1 Pharmacokinetic Studies

Pharmacokinetic studies are an essential part of the clinical

comparability exercise. Since the aim is to demonstrate comparability

and not the characterization of clinical pharmacology of the product per

se, such studies should be comparative.

53

The route of administration should be in accordance with the intended

clinical use. If the product is planned to be administered by more than

one route (e.g. s.c. and i.v.), it may become necessary to test all routes.

The selected dose should be in the steep part of the dose-response curve,

in order to detect relevant differences.

The choice of the population (healthy volunteers vs. patients) is

primarily driven by the mode of action of the product. Since PK and PD

studies are preferably combined, the choice of the study population

should be selected based on the PD effects to be shown, i.e. whether the

PD effects are detectable in a relevant manner in the population of

choice.

The design of comparative PK studies should not necessarily

mimic that of the standard “clinical comparability” design, since

similarity in terms of absorption/bioavailability is not the only

parameter of interest for biosimilars. In fact, differences in elimination

characteristics between products e.g. clearance and elimination half-life

should be explored.

The choice of the design for single dose studies, steady-state

studies, or repeated determination of PK parameters should be

justified by the applicant. The ordinary crossover design is not

appropriate for therapeutic proteins with a long half-life, e.g.

therapeutic antibodies and pegylated proteins, or for proteins for which

formation of anti-drug antibodies is likely.

The acceptance range to conclude clinical comparability with respect

to any pharmacokinetic parameter should be based on clinical

judgment, taking into consideration all available efficacy and safety

information on the RMP and the biosimilar product. Hence, the criteria

used in standard clinical comparability studies, initially developed

for chemically derived, orally administered products may not be

54

appropriate and the clinical comparability limits should be defined

and justified prior to conducting the study. Generally, the requirements

for therapeutic proteins with respect to evaluating the pharmacokinetics

of the product are the same as for conventional products, but specific

considerations are needed related to the inherent characteristics of

proteins.

The pharmacokinetics (absorption, distribution and elimination) should

be characterized during single-dose and steady-state conditions in

relevant populations. However, the pharmacokinetic requirements may

differ depending on the type of protein and its intended use.

4.2.2 Pharmacodynamic Studies

The pharmacodynamic effect should be compared in a population where

the possible differences can best be observed. The design and duration of

the studies must be justified. Pharmacodynamics should preferably be

evaluated as part of the comparative pharmacokinetic study, since

alterations in pharmacodynamics can sometimes be explained by altered

kinetics and such design may provide useful information on the

relationship between exposure and effect.

The selected dose should be in the steep part of the dose-response curve.

Studies at more than one dose level may be useful. Again, studies

should be comparative in nature and not merely show the

pharmacodynamics of the product per se.

4.2.2.1 Markers for primary and secondary

pharmacodynamics

As a principle, an endpoint should be selected that fulfils

the following requirements: (1) sensitive enough to detect

small differences, (2) measurable with sufficient

55

precision, and (3) clinically relevant for the target

population. Please refer to ICH topic E10 for further

details. Care should be taken in these cases to investigate

a reasonable dose range to demonstrate assay sensitivity.

Studies at more than one dose level may be useful.

The choice of marker(s) should be justified and the

margin defining equivalence should be pre-specified and

justified.

In this respect, the choice of the population should be

justified. Demonstration of certain primary or secondary

PD markers might only be apparent in the diseased

population as opposed to healthy volunteers. For

example, immunomodulators aiming at modulating

pathologically altered immune effector cells would not

necessarily exert similar effects in healthy volunteers.

4.2.2.2 Pharmacodynamics markers as substitutes for efficacy

The PD markers should be selected on the basis of their

relevance to demonstrate therapeutic efficacy of the

product. Usually in clinical trials, efficacy is defined by

one or more clinical endpoint(s). A PD marker is a

relevant marker for efficacy, if therapy-induced changes

in that marker to a large extent can explain changes in

clinical outcome.

PD markers are usually more sensitive to changes in

activity of the product and can be assessed earlier than

clinical endpoints and, therefore, they might in some

cases represent the most appropriate endpoint. However,

as the goal of the comparative exercise is showing

56

equivalence of the products, usually data are needed

concerning the quantitative relationship between the PD

marker and the clinical endpoint to enable defining and

justifying the equivalence margin in terms of efficacy.

Sometimes it may be useful to use more than one PD

marker.

Research in surrogate endpoints by the

applicants/marketing authorization holders is encouraged,

since a surrogate marker will be useful in the course of

product development.

4.2.3 Confirmatory pharmacokinetic/pharmacodynamic studies

Normally comparative clinical trials are required for the

demonstration of clinical comparability. In certain cases, however

comparative PK/PD studies between the biosimilar product and the RMP

may be sufficient to demonstrate clinical comparability, provided that all

the following conditions are met:

(a) The PK of the RMP are well characterized. There is sufficient

knowledge of the PD properties of the RMP, including binding to

its target receptor(s) and intrinsic activity. Sometimes, the

mechanism of action of the biological product will be disease-

specific.

(b) The relationship between dose/exposure and response/efficacy of

the RMP (the therapeutic “concentration-response” curve) is

sufficiently characterized.

(c) At least one PD marker is accepted as a surrogate marker for

efficacy, and the relationship between dose/exposure to the

product and this surrogate marker is well known. A PD

marker may be considered a surrogate marker for efficacy if

therapy-induced changes of that marker can explain changes

in clinical outcome to a large extent. Examples include absolute

57

neutrophil count to assess the effect of granulocyte-colony

stimulating factor (G-CSF), and early viral load reduction in

chronic hepatitis C to assess the effect of alpha interferons.

4.3 Demonstration of clinical efficacy

Efficacy of a biosimilar must be documented through clinical trials that should

demonstrate clinical comparability between the biosimilar product and the

RMP. Clinical comparability margins should be pre-specified and justified,

primarily on clinical grounds. If a clinical comparability trial design is not

feasible, other designs should be explored and their use discussed with the

competent authorities.

4.3.1 Study design

Equivalent therapeutic efficacy should be demonstrated. Frequently,

clinical studies should be randomized and double blind to avoid bias.

Possible differences in efficacy should normally be investigated in

studies with the highest probability of showing a difference [see ICH

topic E10].

The acceptable equivalence margin should be set taking into account the

product release specifications, clinical relevance and statistical

considerations, and be pre-specified. The sample size required should

not solely be based on considerations on clinical efficacy, but also on

detection of differences in safety (see below).

58

If an equivalence trial design is not feasible, other designs should be

explored and their use discussed with the competent authorities.

4.3.2 Selection of the most relevant patient population/therapeutic

indication

Since therapeutic proteins can be used for different indications and/or

different patient populations, differential impact on efficacy and/or

safety needs to be considered. Usually, a patient population or indication

should be chosen where differences are best distinguishable, i.e. the most

sensitive model for efficacy. The choice, however, will also depend on

the susceptibility and vulnerability of this population to potential safety

problems, and will have to be justified by the applicant. The Applicant

needs to thoroughly discuss and justify if efficacy and safety results of

the comparative study in one indication or population can reasonably be

assumed to be applicable to other populations or indications. Concerns

regarding human gender, women with child-bearing age potential,

pregnant women, and children must be taken into consideration, as

explained in ICH topic M3(R1).

4.3.3 Determination of study size

The size of a trial is influenced by the disease to be investigated, the

objective of the study and the study endpoints. Statistical assessments of

sample size should be based on the expected magnitude of the treatment

effect, the variability of the data, the specified (small) probability of

error, and the desire for information or subsets of the population or

secondary endpoints. In some circumstances a larger database may be

needed to establish the safety of a drug. The number of subjects in a

clinical trial should always be large enough to provide a reliable answer

to the questions addressed. This number is usually determined by the

59

primary objective of the trial. If the sample size is determined on some

other basis, then this should be made clear and justified. For example, a

trial sized on the basis of safety questions or requirements or important

secondary objectives may need larger numbers of subjects than a trial

sized on the basis of the primary efficacy question. ICH topics E1, E7

and E9 offer the necessary details.

4.3.4 Selection of appropriate endpoints

As a principle, endpoints should be selected that show differences with

the highest accuracy. The clinical requirements for comparative studies

being part of a comparability exercise can be different from those for

conventional confirmatory studies. For marketed products, the endpoints

might not necessarily be those, which had been selected for the

confirmatory trials if they are not sufficiently suitable for detection of

differences. As noted above, pharmacodynamic or other markers like

imaging techniques can be more suitable than genuine clinical endpoints.

The choice of endpoints needs to be fully justified.

4.3.5 Study location

Clinical studies can be carried out at more than a single medical center,

providing that all are approved for performing clinical studies. The

numbers of enrolled subjects projected for each trial site should be

specified. Reason(s) for choice of sample size, including reflections on

(or calculations of) the power of the trial and clinical justification must

be provided. Guidance at ICH topics E6, E8, E9 and E10 is to be taken

in consideration.

60

4.3.6 Study duration

The study duration is essentially driven by the choice of the clinical

endpoint. The duration should be sufficient to detect also minor

differences with sufficient accuracy. Available data from literature

should be included in the justification and discussion of study duration.

Since safety data evaluation is an essential part of the clinical

comparability exercise, the study duration should also be determined

with the aim of detecting relevant differences of safety findings

adequately.

4.4 Demonstration of clinical safety

Even if the efficacy is shown to be comparable, biosimilars may exhibit a

difference in the safety profile (in terms of nature, seriousness, or incidence

of adverse reactions). Pre-licensing safety data should be obtained in a number

of patients sufficient to address the adverse effect profiles of the test and the

RMP. Care should be given to compare the type, severity and frequency of the

adverse reactions between the biosimilar product and the RMP. Data from

pre-approval clinical studies are normally insufficient to identify all potential

differences. Therefore, clinical safety of biosimilars must be monitored

closely on an ongoing basis during the post-approval phase including

continued benefit-risk assessment.

In their discussion of adverse events, applicants should not only include the

incidence, but also possible differences in clinical presentation (duration,

severity and seriousness, reversibility, response to treatment etc..). Further

studies post-licensing may occasionally be needed, e.g. pharmaco-

epidemiological studies. ICH topic E10 explains more.

61

4.4.1 Extent of the safety database

In general, safety data can be gathered as part of the clinical study

aiming at establishing equivalent efficacy. Study duration and sample

size calculation should consider frequency, severity and seriousness of

expected adverse events as well as the clinical setting of the use of the

drug, such as for acute and/or chronic use. Again the general rule applies

that the aim of such trial is not the detection of adverse events per se, but

the evaluation of differences in occurrence. ICH topic E1 explains this

issue in details.

4.4.2 Safety endpoints

Specific safety endpoints should be selected, taking into account both the

typical safety findings known for this biosimilar and/or this biosimilar

class, as well as potential other safety findings which can be deduced

from the mechanism of action. Since unexpected safety findings might

occur, applicants are discouraged from setting up methods in the study

protocols solely aiming at the detection of known safety issues. The

evaluation of comparative immunogenicity should be integral part of

safety evaluation (see the part on Immunogenicity below).

Within the authorization procedure the applicant should present a risk

management plan or after licensing of the product an update of the

existing one in accordance with current GCC regulations and

pharmacovigilance guidelines. This should take into account risks

identified during product development, as well as potential risks.

In the periodic safety update reports (PSURs) which must be submitted

to the regulatory authority, the marketing authorization holder

should address reports and any other information on tolerability that

might be related to the original biosimilar or to a process change.

The cycle of submission of the PSURs might be decided upon by the

regulatory authority on a case-by-case basis.

62

4.5 Immunogenecity

This is the most important aspect of safety of biosimilars. For many proteins and

peptides, a number of patients develop clinically relevant anti-drug antibodies.

The immune response against therapeutic proteins differs between products

since the immunogenic potential is influenced by many factors.

Considerable heterogeneity in antibody response may be observed since an

individual may form multiple antibodies with different affinities, epitopes and

binding capacities. Thus, data should be collected from a sufficient number of

patients to characterize the variability in antibody response. Since anti-drug

antibodies may alter the pharmacokinetics and pharmacodynamics of a protein,

testing for antibody response is always necessary when developing a new

protein. This is especially important for new drugs intended for multiple-dose or

long-term treatment.

The timing of sampling for antibody response should be carefully evaluated and

justified. For example, a sufficient interval between the last dose and the time-

point for antibody detection is crucial, since the drug molecule needs to be

eliminated from the circulation, or otherwise interference with the antibody-

assay is likely. Thus, to minimize interference with the analysis, it is

recommended that samples be collected when drug concentration is low, i.e.

preferably after 6-7 half-lives, and when anti-drug antibodies have developed.

When measuring antibodies during drug treatment, any possible analytical

interference should be investigated and discussed. Information on antibody

formation should preferably be gathered previously in Phase I/II (Phase II likely

to have longer exposure time) to guide planning of Phase III.

The presence of anti-therapeutic protein antibodies should be determined using

both an immunoassay for the presence of binding antibodies and a biological

assay for the presence of neutralizing antibodies. The assays should be fully

63

validated, sufficiently sensitive to detect clinically relevant antibodies, and able

to detect the presence of rapidly dissociating (low affinity) antibodies.

Although the pharmacodynamic effect is directly altered only by neutralizing

antibodies, the pharmacokinetics may be affected irrespective of the neutralizing

capacity. Antibody formation can cause increased or decreased clearance of the

therapeutic protein, although the former effect is the most common. Therefore,

alterations in clinical effect due to anti-drug antibody formation might be a

composite of both pharmacokinetic and pharmacological changes. Whenever

there is a relevant antibody response to the drug, the effect of anti-drug

antibodies on the pharmacokinetics of a protein should be studied unless

justified by the applicant.

Due to variability between individuals, it is important that samples are collected

within the same subjects pre- and post dosing. Pharmacokinetic sampling in

Phase III studies is important in the assessment of anti-drug antibody effects due

to the generally prolonged exposure of the drug and the increased number of

patients in the study. Effects of antibody formation may be studied using

population pharmacokinetic analysis, treating presence of anti-drug antibodies

as a covariate. As a minimum, plasma samples for pharmacokinetic analysis

should be collected after the first and last dose to compare the plasma

concentrations and degree of accumulation in antibody positive and negative

subjects. Special consideration should be given to patients withdrawing from a

trial. Correlations between the onset and degree of the antibody response and the

drug exposure or relevant pharmacokinetic parameters should be examined.

If possible, antibody production over time should be evaluated and retention of

plasma samples should be considered. Consideration should be given to possible

analytical interference of formed antibodies with the assays for the therapeutic

protein. Needless to say, the overriding question to address is the impact of

antibodies on the efficacy and/or safety of the drug. This includes how to treat

patients with a decreasing response to the drug due to antibodies as well as the

64

safety and efficacy of repeated treatment after a significant period of “drug

holiday”.

For further information, please refer to ICH topic S8.

4.5.1 Factors affecting immunogenicity

Many factors influencing the immunogenicity of therapeutic proteins

remain unknown and unpredictable. In general, the antibody response in

man cannot be predicted from animal studies.

The immune response against therapeutic proteins differs between

products since the immunogenic potential is influenced by many factors

such as the nature of the active substance, product- and process-

related impurities, excipients and stability of the product, dosing

regimen, the expression system in which the protein is produced, the

purification system, or its final formulation. The immune response may

depend on the dose and route of administration (subcutaneous route

more immunogenic than intravenous route).

Patient related factors may have a genetic basis such as the lack of

tolerance to the normal endogenous protein, or acquired such as

immunosuppression due to the disease or its concomitant medication.

There is considerable inter-individual variability in antibody

response in terms of different antibody classes, affinities, and

specificities Thus, data should be collected from a sufficient number of

patients to characterize the variability in antibody response.

4.5.2 Consequences of an immune response

The consequences of immunogenicity may vary considerably,

ranging from irrelevant for therapy to serious and life-threatening.

Therefore, the immunogenicity issue has become a subject of concern in

65

the development and approval of biopharmaceuticals. An immune

response to the product may have a significant impact on its clinical

safety and efficacy. Although only neutralizing antibodies directly alter

the pharmacodynamic effect, any binding antibody may affect the

pharmacokinetics. Thus, an altered effect of the product due to anti-

drug antibody formation might be a composite of pharmacokinetic,

pharmacological and safety changes. Antibody formation can cause

increased or decreased clearance of the therapeutic protein, although the

former effect is the most common.

4.5.3 Testing of immunogenicity

The applicant should present a rationale for the proposed antibody-

testing strategy. Testing for immunogenicity should be performed by

state-of-the-art methods, using assays with appropriate specificity and

sensitivity. The screening assays should be validated and sensitive

enough to detect low titre and low affinity antibodies. An assay for

neutralizing antibodies should be available for further characterization of

antibodies detected by the screening assays. Standard methods and

international standards should be used whenever possible. The possible

interference of the circulating antigen with the antibody assays should be

taken into account. The periodicity and timing of sampling for testing of

antibodies should be justified.

In view of the unpredictability of the onset and incidence of

immunogenicity, long term results of monitoring of antibodies at

predetermined intervals will be required. In case of chronic

administration, one-year follow up data will be required pre-licensing.

The applicant should consider the possibility of antibodies against

process-related impurities

66

4.5.4 Evaluation of the clinical significance of the observed immune

response

If a different immune response to the biosimilar product is observed as

compared to the innovator product, further analyses to characterize the

antibodies and their implications to clinical safety, efficacy and

pharmacokinetic parameters are required. Special consideration should

be given to those products where there is a chance that the immune

response could seriously affect the endogenous protein and its unique

biological function. Antibody testing should be considered as part

of all clinical trials protocols. The applicant should consider the role

of immunogenicity in certain events, such as hypersensitivity, infusion

reactions, autoimmunity and loss of efficacy.

4.5.5 Principles for evaluation of immunogenicity

Normally, an antibody response in humans cannot be predicted from

animal studies. Thus, immunogenicity of a biosimilar product must

always be investigated. The assessment of immunogenicity requires an

optima antibody testing strategy, characterization of the observed

immune response, as well as evaluation of the correlation between

antibodies and PK or PD effects relevant for clinical safety and efficacy

in all aspects. It is important to consider the risk of immunogenicity in

different therapeutic indications separately.

The applicant should present a rationale for the proposed antibody

testing strategy. Testing for immunogenicity should be performed by

state-of-the-art methods, using assay with appropriate sensitivity and

specificity. The screening assays should be validated and sensitive

enough to detect low titre and low affinity antibodies. An assay for

neutralizing antibodies should be available for further characterization of

antibodies detected by the screening assays. International standards

should always be followed.

67

The possible interference of the circulating antigen with the antibody

assays should be taken into account. The periodicity and timing of

sampling for testing for antibodies must be justified. In view of the

unpredictability of the onset and incidence of immunogeicity, long-term

results of monitoring of antibodies at predetermined intervals is required.

In case of chronic administration, a pre-licensing, one-year follow-up

data is required.

Antibody testing should be considered part of all clinical trial protocols.

The role of immunogenicity in certain events, such as hypersensitivity,

infusion reactions, autoimmunity and loss of efficacy should be

considered. The sponsor needs to discuss possibilities to encourage the

reporting of relevant adverse events, including events related to loss of

efficacy.

4.6 Risk management and pharmacovigilance

Data from pre-authorization clinical studies are normally insufficient to identify

all potential differences. Therefore, clinical safety of biosimilars must be

monitored closely on an ongoing basis during the post-approval phase,

including continued assessment of benefits and risks.

The applicant should give a risk specification in the application DMF for the

medicinal product under review. This includes a description of possible safety

issues related to tolerability of the medicinal product that may result from a

manufacturing process different from that of the innovator. In the DMF, the

applicant should present a risk management program or pharmacovigilance

plan in accordance with current GCC procedures and guidelines. This should

take into account risks identified during product development and potential

risks.

68

Pharmacovigilance systems and procedures to achieve this monitoring should be

in place when a marketing authorization is granted. Any specific safety

monitoring imposed to the RMP or product class should be taken into

consideration in the risk management plan.

The compliance of the marketing authorization holder with commitments

(where appropriate) and their pharmacovigilance obligations will be closely

monitored. The marketing authorization holder should address reports and any

other information on tolerability of the biosimilar that the company has

received. These reports or information must be evaluated and assessed by the

marketing authorization holder in a scientific manner with regard to causality of

adverse events or adverse drug reactions and related frequencies.

For further information on this issue, ICH topic Q9 can be used. For reporting,

the GCC Guidelines on Pharmacovigilance should be referred to.

4.7 Clinical studies for biosimilars when manufacturing changes are

introduced

Manufacturers of biosimilar products frequently introduce changes in the

manufacturing process of a given product (both before and after the granting of

a marketing authorization). It is assumed that the product‟s physicochemical

properties and in vitro/in vivo biological activity are well characterized

according to state of the art methods. For most changes to the manufacturing

process, physico-chemical and (quality related) biological testing can

demonstrate that there is no difference in quality of the product that could

adversely impact the safety and efficacy of a product. Thus the comparability

exercise may be limited to strict process validation of the change or be extended

to various quality criteria such as in-process controls, thorough analytical and

biological characterization of the product and stability data. However,

sometimes an effect on efficacy and/or safety can be expected on the basis of

69

observed difference(s) or cannot be ruled out in spite of the state of the art

physico-chemical and biological tests. In such cases, additional clinical studies

will be necessary.

The type and extent of such studies are variable and will be dependent on

numerous factors related to the drug substance and the drug product, such as:

(a) Knowledge of the molecule and of other molecules of the same class.

(b) The stage of development of products not yet authorized.

(c) The findings in the physico-chemical and biological comparability

exercise.

(d) The intended clinical use.

In principle, preclinical and clinical data, if required, need to be available before

implementation of the change in the manufacturing process, i.e. marketing the

new version of the product.

Depending on the product and the indication, approval of the process change

might be based on pharmacodynamic data. Additional clinical/safety data,

including immunogenicity data, may be provided after approval.

4.7.1 Clinical comparability after changes are introduced

Demonstration of comparability is a sequential process, beginning with

quality studies (limited or comprehensive) and supported, as necessary,

by non-clinical, clinical and/or pharmacovigilance studies. If a

manufacturer can provide evidence of comparability through

physicochemical and biological studies, then clinical studies with the

post-change product may not be warranted.

The need, extent and nature of clinical comparability studies will be

determined on a case-by-case basis in consideration of various factors

that may be associated with risk, such as:

70

(a) The process complexity, the nature of the change, the potential

impact on the molecule structure and on the final product profile.

(b) The nature, and extent of differences demonstrated by the

physico-chemical and quality-related biological characterization,

including product-related substances, impurity profile, stability

and excipients. Thus, well-characterized differences may provide

a background for a rational and focused approach with respect to

the need for preclinical and clinical studies.

(c) Product complexity, including heterogeneity and higher order

structure and the availability, capabilities and limitations of

analytical tests. If the analytical procedures used are not

sufficient to discern relevant differences that can impact the

safety and efficacy of the product, additional preclinical and/or

confirmatory clinical testing may be necessary.

(d) Structure-activity relationship and strength of the association of

quality attributes with safety and efficacy.

(e) Relationship between the therapeutic protein and endogenous

proteins and the severity of (potential) consequences for

immunogenicity; e.g. risk of autoimmunity

(f) Mode(s) of action: unknown or multiple modes of action

complicate the evaluation of the impact of changes.

(g) Therapeutic indications/target patient groups - The impact of

possible differences can vary between the target populations

covered by the different indications.

(h) Posology: dosing regimen and route of administration. For

instance, repeated administration via the subcutaneous route is

more likely to be associated with immunogenicity than

intravenous administration of a single dose.

(i) The therapeutic window/dose-response curve.

(j) Previous experience, e.g., immunogenicity, safety.

71

Experience with the pre-change product or with other products in the

same class can be relevant. However, each biosimilar should be

considered individually.

For products in development, all these points above should be taken into

consideration. However, the extent of the comparability studies will

likely increase if manufacturing changes are introduced at the later

stages of clinical development. A change after conduct of confirmatory

efficacy and safety studies represents the most challenging situation. The

selection clinical studies is product-driven, i.e. a strategy for

comparability testing should be chosen that best predicts and detects

clinically relevant differences with sufficient accuracy.

If a manufacturing change is introduced before the confirmatory trial(s),

the additional data required for the comparability exercise might be

fewer than those needed for changes introduced after the confirmatory

trial(s) or after approval.

4.7.2 Manufacturing process changes before clinical trials initiation

For this situation, adequate data from physico-chemical and biological in

vivo and in vitro, and sometimes also preclinical or clinical

comparability studies such as a single dose pharmacokinetic study, are

generally sufficient in order to demonstrate that the preclinical and

clinical data obtained before the change has been introduced are still

valid and can be extrapolated to the post-change product.

4.7.3 Manufacturing process changes during clinical trials

Changes during the clinical study are discouraged. If these occur, it is

recommended that the applicant seeks t h e c o n c e r n e d

r e g u l a t o r y a u t h o r i t y advice after proper justification to avoid

unnecessary expenditure.

72

4.7.4 Manufacturing process changes after clinical trials or after approval

If a manufacturing change takes place after a clinical trial has been

performed, or after approval, a more thorough comparability exercise is

generally required. This should include physicochemical and biological

in vitro studies, and may include clinical pharmacokinetic and/or

pharmacodynamic comparability studies. If this comparability exercise

cannot rule out an impact on the efficacy and safety profile of the drug,

additional clinical study or studies may have to be performed. Deviations

from this conceptual level should be justified.

4.7.5 Further criteria influencing the requirement of comparative clinical

data

Further important issues that should be taken into account when

designing and justifying the clinical program include results of any

preclinical study and any clinical experience gained with the pre-change

product and other products in the same category, if relevant, with respect

to:

(a) The relationship between dose/exposure and efficacy/safety

(b) Whether a dynamic marker has been accepted as a surrogate

marker for clinical efficacy/safety.

(c) The relationship between dose/exposure and this surrogate

marker.

(d) Drug/receptor(s) interaction.

(e) Disease-specific mechanisms of action.

(f) Target organ(s) for activity and toxicity.

(g) Mode of administration.

More factors to be considered in planning clinical studies are available at

ICH topic Q5E.

73

4.7.6 Requirements for risk management and pharmacovigilance of a

changed biosimilar

Even if the efficacy is shown to be comparable, the post-change product

may exhibit a difference in the safety profile (in terms of nature,

seriousness, or incidence of adverse reactions). Pre-licensing safety data

should be obtained in a number of patients sufficient to compare the

adverse effect profiles of the pre- and post-change product. Care should

be given to compare the type, severity and frequency of the adverse

reactions between the pre- and post-change product. Further studies post-

licensing may occasionally be needed, e.g. pharmaco-epidemiological

studies.

Applicants should in their discussion of adverse events not only include

the incidence, but also possible differences in clinical presentation

(duration, severity and seriousness, reversibility, response to treatment

etc.).

The applicant should give a safety specification in the application DMF

for the biosimilar under review. This includes a description of possible

safety issues related to the changes in the manufacturing process.

74

CHAPTER 5.0

Other Important Issues

Labeling

Extrapolation Interchangeability and substitution

Stability studies

Storage conditions

75

5.1 Introduction

These issues are important for the use of biosimilars. Although stability and storage

issues have generally been guided and agreed upon, other issues such as labeling and

extrapolation as well as interchangeability and substitution remain to be debatable

among different regulators worldwide and all concerned parties.

5.2 Labeling

This issue deals with the information shown on the outside package and the inside

leaflet. In both, the chosen brand name of the product must be clearly written, with the

scientific name of the product [international non-proprietary name, INN, if there is any

designated by WHO] written underneath in parentheses, with the company‟s name and

logo clearly demonstrated. Storage conditions, names and quantities of the API and

other excipients, as well as other vital instructions should be written. Each biosimilar

must have its own specific and original labels. Labels should not contain any material

that remains covered by a patent law or any other intellectual property right.

The inside leaflet must contain (in addition to the above) all needed labels, including

the name of the RMP and the summary of product characteristics (SmPCs). The same

main headings of the RMP should be in the leaflet, but the description under each

heading should be that of the company‟s findings and data as a result of the extensive

studies performed on the manufactured biosimilar by the company. Copying part(s) of

the information in the leaflet of the RMP or from other sources such as pharmacopeias

or another biosimilar product is prohibited. Differences between same biosimilar

products from different manufacturers and/or differences between the biosimilar and its

RMP during the comparability exercise must be fully reflected in the label. Additional

information that is not found in the leaflet of the RMP but found through the studies of

the manufacturer can be added.

If after marketing, pharmacovigilance mandatory excercise presented new or different

information, or changes to the product were introduced, it is the duty of the

76

manufacturing company to inform t h e r e g u l a t o r y a u t h o r i t y and update or

change the packaging and leaflets information for the proper use and safety of the

product.

In order to avoid confusion, INN or scientific name of a biosimilar should not be used

when prescribed. The brand name and company‟s name or the scientific name and

company‟s name should be clearly written in the prescription. It is essential that

prescribed biosimilars can be easily identified and traced (including batches of the same

biosimilar), and that clinicians be aware of the exact biosimilar given to a patient.

Biosimilars with the same API must have different brand names to avoid inference for

interchangeability and inadvertent substitutions. The label should contain the proper

information about interchangeability and substitution (as guided below in 5.4) to

increase the awareness of the concerned healthcare professional.

For all biosimilar products, precisely defined storage temperatures are recommended.

Specific recommendations should be stated, particularly for drug substances and drug

products that cannot tolerate freezing. These conditions, and where appropriate,

recommendations for protection against light and/or humidity, should appear on

containers, packages, and/or package inserts. Such labeling should be in accordance

with relevant national/regional requirements.

5.3 Extrapolation

The issue of extrapolation is sometimes encountered when a DMF is submitted for

registration. Some recombinant medicinal products may have more than a single

indication. The clinical study performed by the applicant for the main indication of a

biosimilar product may not be used for another indication or formulation. However, a

biosimilar product can be approved for another indication approved for the RMP when

all comparability studies to the RMP for the main indication were similar.

In this document, extrapolation refers to the submission for approval of another

indication for a biosimilar that was not evaluated by clinical studies and the request is

77

based on the indications of the RMP, even if clinical studies for the original main

indication were carried out by the applicant.

Extrapolation of a new formulation or an indication for a closely related disease maybe

granted on a case by case basis if all other comparability aspects were similar to the

RMP and the company can plausibly justify the use for another closely related

indication without the need for a clinical study.

5.4 Interchangeability and substitution

This remains a controversial issue among different regulators worldwide and all

concerned parties. Biosimilars are protein therapies similar to indigenous human

mediators, are given in microgram quantities, are not exact copies of an original

medicine, and have limited clinical experience at approval. Although interchangeability

and substitution are not encouraged and can be detrimental to pharamcovigilance and

risk management, there could be situations (financial, availability, intolerability,

hospital or country necessities) when they are needed. It is generally viewed that

changing or substituting a protein medicine produced by rDNA technology, whether

original (innovator) or a biosimilar, is the decision of the physician and the patient

when the treating doctor explains to the stakeholder the possibility of such substitution

and examine the risks versus benefits. Physicians and pharmacist should discuss the

issue before talking to the patient to prevent inappropriate substitution. Pharmacists

cannot substitute biosimilars without such consultations with treating physicians.

However, the GCC strongly recommends the followings:

(1) Changing from an innovator drug to a biosimilar drug which used that same

innovator drug as its RMP for comparability (or vice versa) can be accepted

after physician and patient discussion.

(2) Changing from a biosimilar drug to another same biosimilar drug from a

different manufacturer can be accepted after physician and patient discussion

only if they both used the same RMP for comparability purposes.

78

(3) Changing from an innovator drug to another innovator drug for the same

indication, or from a biosimilar drug to another biosimilar drug which did not

use the same innovator drug as a RMP for comparability is not acceptable in

ordinary situation. In extreme situations, physician and patient discussion, as

well as hospital administration involvement in the decision are mandatory.

In all cases, close monitoring of the patient‟s responses should be performed when

interchangeability or substitution is warranted, perhaps on a daily basis until results are

satisfactory and stable. Dosage and route of administration should be studied and

adjusted when necessary. Minute differences among biosimilars and between a

biosimilar and the innovator may affect the clinical outcomes. In addition, and for

obvious reasons, substitutions negatively affect the pharmacovigilance exercise.

5.5 Stability

The stability of a product is generally highly dependent on its storage conditions, which

must be clearly defined according to the product's characteristics. A biopharmaceutical

from the same manufacturer may also degrade at different speed and at different

conditions due to inadequate control of the production process or other reasons.

Biosimilars are rather unstable structures. Most of biosimilars have to be stored at 4oC,

and never shaken or heated. These storage and handling recommendations are based on

the innovator‟s exhaustive testing to ensure drug stability from the line of production to

the point of drug administration. Any change in the storage or handling process could

lead to protein degradation or aggregation. Aggregates (caused primarily by loss of the

original protein‟s tertiary structure) are a major source of immunogenicity.

The stability of a molecule and its degradation pathway can be studied by exposing it to

increased temperature. For example, gel-electrophoresis (or newer methods) of an

unstable product will show various bands apart from the principal molecule, indicating a

higher tendency to form aggregates. Decreased stability in comparison with the RMP

may either be due to an unstable formulation or a defective active ingredient.

79

For biosimilars, stability studies must cover the whole period of validity (shelf-life) in

real time testing fashion for all stability parameters (appearance, color, purity, identity,

potency, and others) at regular intervals and at the suggested storage temperature

(usually 5oC). Accelerated stability testing is also mandatory, to be performed for six

months at higher temperatures (usually 30oC with 65% Relative Humidity) and testing

all parameters at months 1, 3, and 6. Stress stability testing (usually 45oC) is strongly

suggested for one month. The range for identity or potency can be from 90% to 110%

of the BRP, or as described by compendial methods in reference pharmacopoeias,

keeping in mind that all other aspects of the product (particularly the clinical studies)

are complete and acceptable.

If significant change occurs between 0 and 6 months‟ testing at the accelerated storage

condition, the proposed retest period or shelf life should be based on long-term data.

Extrapolation is not considered appropriate. A retest period or shelf life shorter than the

period covered by long-term data could be called for. If the long-term data show

variability, verification of the proposed retest period or shelf life by statistical analysis

can be appropriate. In addition, a discussion should be provided to address the effect of

short-term excursions outside the label storage condition (e.g., during shipping or

handling).

Generally, there is no single stability-indicating assay or parameter that profiles the

stability characteristics of a biosimilar product. Consequently, the applicant should

propose a stability-indicating profile that provides assurance that changes in the

identity, purity and potency of the product will be specifically detected. The

determination of which tests should be included and must be product-specific. The

stability studies must be performed on the finished product in the format that will be

marketed and before the onset of clinical studies. Applicants should have validated the

advanced methods they used for the stability profile and the data available for review.

The items emphasized in the following subsections are not intended to be all-inclusive,

but represent product characteristics that should typically be documented to adequately

demonstrate product stability.

80

5.5.1 Protocol

The dossier accompanying the application for marketing authorization

should include a detailed protocol for the assessment of the stability of

both drug substance and drug product in support of the proposed storage

conditions and expiration dating periods. The protocol should include all

necessary information which demonstrates the stability of the

biotechnological/biological product throughout the proposed expiration

dating period including, for example, well-defined specifications and test

intervals. The statistical methods that should be used are described in

the tripartite ICH guideline on stability.

5.5.2 Potency

When the intended use of a product is linked to a definable and

measurable biological activity, testing for potency should be part of the

stability studies. For the purpose of stability testing of the products

described in this guideline, potency is the specific ability or capacity of a

product to achieve its intended effect. It is based on the measurement of

some attribute of the product and is determined by a suitable quantitative

method. In general, potencies of biotechnological/biological products

tested by different laboratories can be compared in a meaningful way

only if expressed in relation to that of an appropriate reference material.

For that purpose, a reference material calibrated directly or indirectly

against the corresponding national or international reference material

should be included in the assay.

Potency studies should be performed at appropriate intervals as defined

in the stability protocol and the results should be reported in units of

biological activity calibrated, whenever possible, against nationally or

internationally recognized standard. Where no national or international

reference standards exist, the assay results may be reported in in-house

derived units using a characterized reference material.

81

In some biosimilar products, potency is dependent upon the conjugation

of the active ingredient(s) to a second moiety or binding to an adjuvant

(such as pegylated products). Dissociation of the active ingredient(s)

from the carrier used in conjugates or adjuvants should be examined in

real-time/real-temperature studies (including conditions encountered

during shipment). The assessment of the stability of such products may

be difficult since, in some cases, in vitro tests for biological activity and

physicochemical characterization are impractical or provide inaccurate

results. Appropriate strategies (e.g., testing the product prior to

conjugation/binding, assessing the release of the active compound from

the second moiety, in vivo assays) or the use of an appropriate surrogate

test should be considered to overcome the inadequacies of in vitro

testing.

5.5.3 Purity and molecular characterization

For the purpose of stability testing of the products described in this

guideline, purity is a relative term. Due to the effect of glycosylation,

deamidation, or other heterogeneities, the absolute purity of a

biotechnological/biological product is extremely difficult to determine.

Thus, the purity of a biotechnological/biological product should be

typically assessed by more than one method and the purity value derived

is method-dependent. For the purpose of stability testing, tests for purity

should focus on methods for determination of degradation products.

The degree of purity, as well as individual and total amounts of

degradation products of the biotechnological/biological product entered

into the stability studies, should be reported and documented whenever

possible. Limits of acceptable degradation should be derived from the

analytical profiles of batches of the drug substance and drug product

used in the preclinical and clinical studies.

82

The use of relevant physicochemical, biochemical and immunochemical

analytical methodologies should permit a comprehensive

characterization of the drug substance and/or drug product (e.g.,

molecular size, charge, hydrophobicity) and the accurate detection of

degradation changes that may result from deamidation, oxidation,

sulfoxidation, aggregation or fragmentation during storage. As

examples, methods that may contribute to this include electrophoresis

(SDS-PAGE, immunoelectrophoresis, Western blot, isoelectrofocusing),

high-resolution chromatography (e.g., reversed-phase chromatography,

gel filtration, ion exchange, affinity chromatography), and peptide

mapping.

Wherever significant qualitative or quantitative changes indicative of

degradation product formation are detected during long-term, accelerated

and/or stress stability studies, consideration should be given to potential

hazards and to the need for characterization and quantification of

degradation products within the long-term stability program. Acceptable

limits should be proposed and justified, taking into account the levels

observed in material used in preclinical and clinical studies.

For substances that cannot be properly characterized or products for

which an exact analysis of the purity cannot be determined through

routine analytical methods, the applicant should propose and justify

alternative testing procedures.

5.5.4 Other product characteristics

The following product characteristics, though not specifically relating to

biotechnological/biological products, should be monitored and reported

for the drug product in its final container:

(1) Visual appearance of the product (color and opacity for

solutions/suspensions; color, texture and dissolution time for

83

powders), visible particulates in solutions or after the

reconstitution of powders or lyophilized cakes, pH, and moisture

level of powders and lyophilized products.

(2) Sterility testing or alternatives (e.g., container/closure integrity

testing) should be performed at a minimum initially and at the

end of the proposed shelf-life. Additives (e.g., stabilizers,

preservatives) or excipients may degrade during the dating period

of the drug product. If there is any indication during preliminary

stability studies that reaction or degradation of such materials

adversely affects the quality of the drug product, these items may

need to be monitored during the stability program.

(3) The container/closure has the potential to adversely affect the

product and should be carefully evaluated.

5.5.5 Testing frequency

The shelf-lives of biotechnological/biological products may vary from

days to several years. Thus, it is difficult to draft uniform guidelines

regarding the stability study duration and testing frequency that would be

applicable to all types of biotechnological/biological products. With

only a few exceptions, however, the shelf-lives for existing products and

potential future products will be within the range of 0.5 to 5 years.

Therefore, the guidance is based upon expected shelf-lives in that range.

This takes into account the fact that degradation of

biotechnological/biological products may not be governed by the same

factors during different intervals of a long storage period.

When shelf-lives of 1 year or less are proposed, the real-time stability

studies should be conducted monthly for the first 3 months and at 3

month intervals thereafter.

84

For products with proposed shelf-lives of greater than 1 year, the studies

should be conducted every 3 months during the first year of storage,

every 6 months during the second year, and annually thereafter.

While the testing intervals listed above may be appropriate in the pre-

approval or pre-license stage, reduced testing may be appropriate after

approval or licensure where data are available that demonstrate adequate

stability. Where data exist that indicate the stability of a product is not

compromised, the applicant is encouraged to submit a protocol which

supports elimination of specific test intervals (e.g., 9 month testing) for

post-approval/post-licensure, long-term studies.

5.5.6 Specifications

Although biosimilar products may be subject to significant losses of

activity, physicochemical changes, or degradation during storage,

international and national regulations have provided little guidance with

respect to distinct release and end of shelf-life specifications.

Recommendations for maximum acceptable losses of activity, limits for

physicochemical changes, or degradation during the proposed shelf-life

have not been developed for individual types or groups of

biotechnological/biological products but are considered on a case-by-

case basis. Each product should retain its specifications within

established limits for safety, purity, and potency throughout its proposed

shelf-life. These specifications and limits should be derived from all

available information using the appropriate statistical methods. The use

of different specifications for release and expiration should be supported

by sufficient data to demonstrate that clinical performance is not affected

as discussed in the tripartite ICH guideline on stability.

For additional information, please refer to “The GCC guidelines for

stability testing of drug substances and pharmaceutical products,”

85

published in the GCC website and the ICH-Q5C. All are listed in the

reference section.

5.6 Storage conditions

5.6.1. Temperature

Most finished biosimilar products need precisely defined storage

temperatures. The storage conditions for the real-time/real-temperature

stability studies may be confined to the proposed storage temperature.

5.6.2. Humidity

Biotechnological and biological products, including biosimilars, are

generally distributed in containers protecting them against humidity.

Therefore, where it can be demonstrated that the proposed containers

(and conditions of storage) afford sufficient protection against high and

low humidity, stability tests at different relative humidities can usually

be omitted. Where humidity-protecting containers are not used,

appropriate stability data should be provided.

5.6.3. Accelerated and stress conditions

As previously noted, the expiration dating should be based on real-

time/real-temperature data. However, it is strongly suggested that

studies be conducted on the drug substance and drug product under

accelerated and stress conditions. Studies under accelerated conditions

may provide useful support data for establishing the expiration date,

provide product stability information for future product development

(e.g., preliminary assessment of proposed manufacturing changes such as

change in formulation, scale-up), assist in validation of analytical

methods for the stability program, or generate information which may

86

help elucidate the degradation profile of the drug substance or drug

product. Studies under stress conditions may be useful in determining

whether accidental exposures to conditions other than those proposed

(e.g., during transportation) are deleterious to the product and also for

evaluating which specific test parameters may be the best indicators of

product stability. Studies of the exposure of the drug substance or drug

product to extreme conditions may help to reveal patterns of

degradation; if so, such changes should be monitored under proposed

storage conditions. While the tripartite ICH guideline on stability

describes the conditions of the accelerated and stress study, the applicant

should note that those conditions may not be appropriate for

biotechnological/biological products. Conditions should be carefully

selected on a case-by-case basis.

5.6.4. Light

Applicants should consult the appropriate regulatory authorities on a

case-by-case basis to determine guidance for testing.

5.6.5. Container/Closure

Changes in the quality of the product may occur due to the interactions

between the formulated biotechnological/biological product and

container/closure. Where the lack of interactions cannot be excluded in

liquid products (other than sealed ampoules), stability studies should

include samples maintained in the inverted or horizontal position (i.e., in

contact with the closure), as well as in the upright position, to determine

the effects of the closure on product quality. Data should be supplied for

all different container/closure combinations that will be marketed.

In addition to the standard data necessary for a conventional single-use

vial, the applicant should demonstrate that the closure used with a

multiple-dose vial is capable of withstanding the conditions of repeated

87

insertions and withdrawals so that the product retains its full potency,

purity, and quality for the maximum period specified in the instructions-

for-use on containers, packages, and/or package inserts. Such labelling

should be in accordance with relevant national/regional requirements.

5.6.6. Stability after reconstitution of freeze-dried product

The stability of freeze-dried products after their reconstitution should be

demonstrated for the conditions and the maximum storage period

specified on containers, packages, and/or package inserts. Such labeling

should be in accordance with relevant national/regional requirements.

88

Specific Guidance for Individual Biosimilar Medicines

The following Chapters deal with specific requirements for some of the most commonly

used biosimilar products. The requirements described in this Chapter are in addition to

what is described in previous Chapters (1-4) of this Guidelines. If no specific point

related to quality, efficacy, safety, risk management, and pharmacovigilance is

mentioned, it means that it is mentioned in the main Guideline (Chapter 1-4), and must

be followed. Pharmaceutical development and all other parameters (preclinical and

clinical) should represent the current state-of-the-art, and must meet relevant GCC

guidelines. Comparability to the reference medicinal (innovator‟s) product for all

aspects of production is mandatory. The choice of the RMP for each biosimilar product

should be justified.

89

CHAPTER 6.0

Insulin

90

6.1 General outline

Human insulin for therapeutic use is a non-glycosylated, disulphide-bonded

heterodimer of 51 amino acids. This Chapter discusses the manyfacturing, preclinical

and clinical requirements for recombinant soluble (short acting) rh-insulin products

similar to what are already marketed. There is extensive experience with the production

of insulin for therapeutic use from animal sources, in the form of semi synthetic insulin,

and through different recombinant techniques.

Different types of recombinant insulin are presently available, including short- and long

acting insulin-analogues, short, intermediate and long-acting human insulins, mixed

human insulins, and mixed insulin analogues. Additionally, methods of delivering

insulin orally (monopegylated insulin, spray, capsules etc.) and by inhalation

(pulmonary drug delivery systems) are being developed and should be taken into

account by regulatory bodies.

All the steps should follow the comparability exercise discussed previously. The

following are additional information required for this product.

6.2 Manufacturing considerations

Pharmaceutical development should represent the current state of art and meet

the relevant guidelines. The procedures followed should be same as that

previously discussed earlier in this chapter under manufacturing, taking into

account the differences described.

Physicochemical and biological methods are available to characterize the

primary, secondary and tertiary structures of the recombinant insulin

molecule, as well as its receptor affinity and biological activity in vitro and in

vivo. Attention should be given to product related substances/impurities

and process related impurities, and in particular to desamido forms and other

forms that may be derived from the expression vector or may arise from the

91

conversion steps removing the C-peptide and regenerating the three

dimensional structure.

6.3 Preclinical Issues

The procedures followed should be same as that previously discussed earlier in

this document under preclinical issues, taking into account the following

differences.

6.3.1 Pharmacodynamic studies

6.3.1.1 In vitro studies

In order to assess any differences in properties between

the similar biological medicinal product and the RMP,

comparative studies such as in vitro bioassays for affinity,

insulin- and IGF-1-receptor binding assays, as well as

tests for intrinsic activity should be performed.

6.3.1.2 In vivo studies

Comparative study(ies) of pharmacodynamic effects

would not be anticipated to be sensitive enough to detect

any non-equivalence not identified by in vitro assays, and

are normally not required as part of the comparability

exercise.

6.3.2 Toxicological studies

Data for local tolerance from at least one repeat dose toxicity study in a

relevant species (e.g. rat) should be provided. Study duration should be

at least 4 weeks. Other routine toxicological studies are not required

for rh-insulins developed as similar biological medicinal products.

92

6.4 Clinical studies

The procedures followed should be same as that previously discussed earlier in

this document under clinical studies, taking into account the following

differences.

6.4.1 Pharmacokinetic studies

The relative pharmacokinetic properties of the biosimilar product and the

RMP should be determined in a single dose crossover study using

subcutaneous administration. Comprehensive comparative data should

be provided on the time-concentration profile (AUC as the primary

endpoint and Cmax, Tmax, and T1/2 as secondary endpoints.

Studies should be performed preferably in patients with type 1 diabetes.

Factors contributing to PK variability (e.g. insulin dose and site of

injection/thickness of subcutaneous fat) should be taken into account.

6.4.2 Pharmacodynamic studies

The clinical activity of an insulin preparation is determined by its

time effect profile of hypoglycaemic response, which incorporates

components of pharmacodynamics and pharmacokinetics.

Pharmacodynamic data are of primary importance to demonstrate

comparability of a similar rh-insulin. The double-blind, crossover

hyperinsulinaemic euglycaemic clamp study is suitable for this

characterization.

Data on comparability regarding glucose infusion rate and serum insulin

concentrations should be made available. The choice of study population

and study duration should be justified. Plasma glucose levels should

93

be obtained as part of the PK study following subcutaneous

administration.

6.4.3 Clinical efficacy studies

Provided that clinical comparability can be concluded from PK and PD

data, there is no anticipated need for efficacy studies on intermediary or

clinical variables.

6.4.4 Clinical Safety

Antibodies to rh-insulin occur frequently, mainly as cross-reacting

antibodies. These have been rarely described to have major

consequences for efficacy or safety. The potential for development of

product/impurity-specific antibodies needs to be evaluated. Possible

patient-related risk factors of immune response are unknown.

The safety concerns with a biosimilar rh-insulin relate mainly to the

potential for immunogenicity. The issue of immunogenicity can only be

settled through clinical trials of sufficient duration, i.e. at least 12 months

using subcutaneous administration.

The comparative phase of the study should be at least 6 months, to be

completed before submission for approval. Data at the end of 12 months

could be presented as part of post-marketing commitment.

The primary outcome measure should be the incidence of antibodies to

the test and RMP. The plans for these trials should take into account the

following parameters:

(1) Justification of study population including history of previous

insulin exposure

94

(2) Definitions of pre-specified analyses of the immunogenicity data

with respect to effects on clinical findings (glycaemic control,

insulin dose requirements, local and systemic allergic reactions)

6.4.5 Local reactions

If any concern is raised through the preclinical and short-term clinical

studies outlined above, additional evaluation of local tolerability may be

needed pre-marketing. Otherwise, such reactions should be monitored

and recorded within immunogenicity trials.

6.4.6 Risk Management and pharmacovigilence

Within the authorisation procedure the applicant should present a

risk management program/pharmacovigilance plan in accordance with

national and international pharmacovigilance guidelines. This should

take into account risks identified during product development and

potential risks, especially as regards immuno-genicity, and should detail

how these issues will be addressed in postmarketing follow-up.

95

CHAPTER 7.0

Interferons

Interferons (IFNs) are glycosylated natural proteins produced by a wide variety of cells,

particularly those of the immune system, of most vertebrates including humans. They

are particularly produced in response to the existence of double stranded RNAs that are

foreign to the body. They combat materials foreign to the body, such as viruses,

parasites and tumor cells. Three types of IFNs exist, namely I, II and III. IFNs alpha ()

and beta () belonging to type I IFNs as well as IFN gamma () belonging to type II

IFNs have been used to attempt treatment of certain human illnesses.

96

7.1 Recombinant Interferon Alpha (rIFN-

7.1.1 General outline

Human interferons alpha (IFN-) 2a and 2b are well-known and characterized proteins

consisting of 165 amino acids. The non-glycosylated protein has a molecular weight of

approximately 19,240 D. It contains two disulfide bonds, one between the cysteine

residues 1 and 98, and the other between the cysteine residues 29 and 138. The

sequence contains potential O-glycosylation sites. Physicochemical and biological

methods are available for characterization of the proteins.

It is commonly used subcutaneously although it can also be used through intramuscular

or intravenous route. The sub-types IFN- 2a and 2b have different clinical use. In

general, IFN-2a or 2b use in oncology indications has been reduced considerably and

superseded by other more effective treatments.

Treatment with IFN-2a or 2b is associated with a variety of adverse reactions such as

flu-like illness, fatigue, myalgia, psychiatric, and hematological and renal disorders. It

may also induce development of autoantibodies. A variety of immunomediated

disorders such as thyroid disease, rheumatoid arthritis, systemic lupus erythematosus,

neuropathies and vasculitis have been observed with IFN therapy.

Recombinant IFN-2a or 2b (rIFN-2a or 2b) is approved to manage a wide variety of

conditions such as viral hepatitis B and C, leukaemia, lymphoma, renal cell carcinoma

and multiple myeloma. It is used alone or in combination in these indications. IFN-

may have several pharmacodynamic effects.

7.1.2 Manufacturing considerations

The active substance used to manufacture the recombinant interferon alpha as

well as the preparation and characterization techniques should be described and

justified.

97

7.1.3 Preclinical issues

7.1.3.1 In vitro pharmacodynamics (PD) studies: In order to compare

any alterations in reactivity between the biosimilar and the RMP,

data from a number of comparative bioassays (e.g., receptor-

binding studies, antiviral effects in cell culture, antiproliferative

effects on human tumor cell lines), many of which may already

be available from bioassays submitted as part of the quality

DMF, should be provided. The limitations of studying antiviral

effects in cell culture systems expressing HCV, however, should

be recognized, as the results do not correlate well with clinical

response. Standardized assays should be used to measure activity

and potency.

7.1.3.2 In vivo PD studies: To support the comparability exercise for the

sought clinical indications, the PD activity of the biosimilar

product and the RMP should be quantitatively comparable in (1)

an appropriate PD animal model (e.g. evaluating effects on PD

markers as for example serum 2 ,́5´-oligoadenylate synthetase

activity) and may be performed as part of repeat-dose toxicity

studies, (2) a suitable animal tumor model (e.g., nude mice

bearing human tumor xenografts), and/or (3) a suitable animal

antiviral model.

7.1.3.3 Toxicological studies: Data from at least one repeat dose

toxicity study in a relevant species should be provided (for

example, human IFN-may show activity in the Syrian golden

hamster). Study duration should be at least 4 weeks. The study

should be performed in accordance with the requirements of the

“Guideline on similar biological medicinal products containing

biotechnology-derived proteins as active substance: non-clinical

and clinical issues “and the "Note for guidance on repeated dose

98

toxicity" (CPMP/SWP/1042/99) and include appropriate

toxicokinetic measurements in accordance with the "Note for

guidance on toxicokinetics: A guidance for assessing systemic

exposure in toxicological studies" (CPMP/ICH/384/95). Data on

local tolerance in at least one species should be provided in

accordance with the "Note for guidance on non-clinical local

tolerance testing of medicinal products" (CPMP/SWP/2145/00).

If feasible, local tolerance testing can be performed as part of the

described repeat dose toxicity study.

7.1.4 Clinical studies

7.1.4.1 Pharmacokinetic (PK) studies: The PK properties of the

biosimilar product and the RMP should be compared in single

dose crossover studies using subcutaneous and intravenous

administration in healthy volunteers. The recommended

pharmacokinetic parameters are AUC, Cmax and T1/2.

Equivalence margins have to be defined a priori and

appropriately justified.

7.1.4.2 Pharmacodynamic (PD) studies: There are a number of PD

markers, such as -2 microglobulin, neopterin and serum 2 ,́5´-

oligoadenylate synthetase activity, which are relevant to the

interaction between IFN-and the immune system. The selected

dose should be in the linear ascending part of the dose-response

curve. Whereas the relative importance of these effects in the

different therapeutic indications is unknown, a comprehensive

comparative evaluation of such markers following administration

of the biosimilar product and the RMP could provide useful

supporting data.

99

7.1.4.3 Patient population: The mechanism of action of interferon

comprises of several different unrelated effects. Demonstration of

similar efficacy between the biosimilar product and the RMP is

required and it is recommended that this should be performed in

treatment-naïve patients with chronic hepatitis C (HCV) as

delineated by the indication for the RMP. Other patient

population(s) might be studied depending on the indications

desired.

7.1.4.4 Study design and duration: A randomized, parallel group

comparison against the RMP over at least 48 weeks is

recommended. If possible, the study should be double-blind at

least until data to complete the primary analysis have been

generated. If this not feasible, justification should be provided

and efforts to reduce and eliminate bias should be clearly

identified in the protocol. The posology (i.e., dose, route and

method of administration) should be the same as for the RMP.

The study could be designed so that the primary efficacy analysis

is performed at week 24 for all enrolled patients followed by a

secondary analysis at 48 weeks. Preferably, a homogenous and

sensitive (e.g. genotype selection) patient population is

recommended to best detect differences. The choice of the patient

population should be justified. If a mixed population is chosen,

they should be pre-stratified based on diseases parameters. For

example, in the case of HCV, parameters incluse the HCV

genotypes. The 48-week time point would constitute end-of-

treatment for those patients with HCV genotype 1. For patients

with HCV genotypes 2 and 3, week 48 would usually constitute

24 weeks post-therapy, during which time the status of antibodies

to rIFN-and the relapse rates could be assessed.

100

7.1.4.5 Endpoint(s): Primary: Virologic response as measured by the

proportion of patients with undetectable levels of HCV RNA by

quantitative PCR at week 24. The assay used to measure HCV

RNA and the cut-off applied should be justified. A 2-log

decrease in viral load may be a co-primary endpoint. Secondary:

virologic response at weeks 4, 12, 48; change in liver

biochemistry including transaminase levels and morbidity.

7.1.4.6 Safety: Safety data should be collected from a cohort of patients

after repeated dosing in a comparative clinical trial over a period

of 48 weeks and should be presented with marketing

authorization application. The number of patients should be

sufficient for the comparative evaluation of the adverse effect

profile, including laboratory abnormalities for immune mediated

disorders. The safety profile should be similar between the

biosimilar product and the RMP for the common adverse events

(such as flu-like illness, alopecia, myalgia, leucopenia, anemia

and thrombocytopenia).

101

7.2 Recombinant Interferon Beta (rIFN-

7.2.1 General outline

rIFN- is produced from Chinese Hamster Ovary (CHO) cells which contain the gene

for human rIFN-rhIFN-which is a glycosylated polypetide containing 166 amino

acid residues and is reported to be identical to that of natural human interferon beta. It is

indicated for the treatment of ambulatory patients with relapsing multiple sclerosis

(MS) characterized by at least 2 recurrent attacks of neurologic dysfunction (relapses)

over the preceding 3-year period without evidence of continuous progression between

relapses. It slows the progression of disability and decreases the frequency of relapses.

rhIFN-is also indicated for the treatment of patients who have experienced a single

demyelinating event with an active inflammatory process if it is severe enough to

warrant treatment with intravenous corticosteroids, if alternative diagnoses have been

excluded, and if they are determined to be at high risk of developing clinically definite

MS. rIFN-has not yet been investigated in patients with progressive MS and should

be discontinued in patients who develop this condition. The precise mechanism of

action of IFN-as a possible treatment for MS, however, is not known. All the steps

should follow the comparability exercise discussed in previous Chapters.

7.2.2 Manufacturing considerations

rIFN-b, with its 166 amino acids, should be glycosylated at residue 80 and

should contain a single disulfide bond. The interferon gene should be obtained

from a line of human leukocytes. The DNA should be amplified by PCR

(polymerase chain reaction) and cloned into an expression vector. A CHO cell

line can be used as host. The expression construct for IFN beta-1a and the

sequence of the selection plasmid has to be verified. The active substance used

to manufacturing as well as the preparation and characterization techniques

should be described and justified.

102

7.2.3 Preclinical issues

7.2.3.1 Pharmacodynamics: No acceptable animal model or in vitro

model for MS exists as yet in a species that is

pharmacodynamically responsive to rhIFN--1a. Since the

activities of rhIFN--1a are highly species-specific, the most

relevant information can be derived from in vitro studies in

human cell cultures and in vivo studies in rhesus monkeys.

7.2.3.2 Pharmacokinetics: Serum concentrations of rhIFN--1a can be

measured as antiviral activity in the CPE bioassay. PK should be

studied after a single intravenous, subcutaneous, or intramuscular

administration.

7.2.3.3 Toxicology: Several repeated subcutaneous dose studies can be

performed in rhesus monkeys; the duration of treatment may

range from 2 to 9 weeks (including recovery). Statistically

reliable data in reproductive toxicity testing can only be obtained

by using an excessive number of animals. In view of the data

obtained, a warning is present in the Statistical Process Control

or special purpose corporation regarding the abortifacient

potential of IFN-.

7.2.4 Clinical studies

It is not known whether the efficacy of rhIFN--1a in MS sclerosis is mediated

by the same pathway as the antiviral effect and induction of biomarkers by

rhIFN--1a. For example, IFN-gamma, an interferon with antiviral effect,

increases symptoms in MS. Consequently, markers of antiviral effect cannot

necessarily be considered as surrogate parameters establishing therapeutic

equivalence of rhIFN--1a in MS. These finding should be considered in the

103

light of the overall information concerning the biopharmaceutical

characterization as previously discussed.

7.2.4.1 Pharmacokinetics/pharmacodynamics: High baseline

variability in antiviral activity might mask the dose-response

effect. Potential accumulation in serum or tissues following a

once-weekly intramuscular administration should be investigated.

Though its sensitivity is potentially poor, cytopathic effect assay

can be used to measure serum interferon levels.

7.2.4.2 Clinical efficacy and safety: The studies should be performed in

accordance with good clinical practice, and as discussed in

previous Chapters.

7.2.4.3 Immunogenicity of rhIFN--1a: The issue of antigenicity

raised objections by the CPMP (Committee for Proprietary

Medicinal Products) of EMEA; as most of the data initially

available referred to BG 9015 and the immunogenicity of BG

9015 and BG 9418 (the substance to be marketed as rhIFN--1a)

which were not considered equivalent. As a consequence, further

data should be obtained regarding the relevance of the antibodies

for the therapeutic response.

7.2.5 rhIFN--1a for the treatment of acute lung injury

IFN-has a known anti-inflammatory action; therefore, it is expected to reduce

the leakage of blood and fluids from the capillaries in the lungs (and elsewhere

in the body). IFN-is thus likely to reduce the damage to the lungs, and

improve the oxygenation of the patient. The effects of rhIFN--1a were

evaluated in experimental models. No clinical trials in patients with acute lung

injury (ALI) had been initiated. Hence, there is a positive opinion for rhIFN--

1a to be designated as an orphan drug.

104

Currently, IFN-is not authorized anywhere in the world for the treatment of

ALI, or designated as orphan medicinal product elsewhere for this condition.

However, it authorized in the European Union for the treatment of multiple

sclerosis.

105

7.3 Recombinant Interferon Gamma (rIFN-

rIFN-is a potent cytokine that modulates IL-4-induced immune responses. It is

being researched for treatment of several conditions such as atopic dermatitis,

reduction the severity of infections such as HIV and tuberculosis, and idiopathic

pulmonary fibrosis. It is designated as an orphan drug.

A single product that is a bioengineered rIFN-1b form of interferon gamma, a

protein that acts as a biologic response modifier through stimulation of the

human immune system. It is approved by the USFDA for use in children and

adults with chronic granulomatous disease and severe, malignant osteopetrosis.

Guidelines for this product are not yet available. However, the guidelines for

IFN-and IFN-should be followed.

106

Chapter 8.0

Erythropoietin

(Epoetin, EPO)

107

8.1 General Outline

Human erythropoietin is a 165 amino acid glycoprotein mainly produced in the kidneys

and is responsible for the stimulation of red blood cell production. Erythropoietin-

containing medical products are currently indicated for several conditions such as

anemia in patients with chronic renal failure, chemotherapy-induced anemia in cancer

patients, and for increasing the yield of autologous blood from patients in a pre-

donation program.

8.2 Manufacturing considerations

Erythropoietin for clinical use is produced by recombinant DNA technology

(epoetin) using mammalian cells as expression system. All erythropoietins in

clinical use have a similar amino acid sequence as endogenous erythropoietin

but differ in the glycosylation pattern. Glycosylation influences

pharmacokinetics and may affect efficacy and safety, particularly

imunogenicity. Phisico-chemical and biological methods are available for

characterization of the protein.

When the active substance from the RMP is isolated in order to perform the

comparative analysis at the active substance level, the applicant shall

demonstrate that the isolated active substance used in the comparability exercise

is representative of the active substance present in the RMP.

Using a set of orthogonal state-of-the-art analytical methods, an extensive

characterization program shall be conducted for drug substance to elucidate

structural features of the protein backbone as well as the carbohydrate moieties.

The primary structure shall be confirmed to show that the molecule has an intact

protein structure with correctly linked disulfide bonds and integrity of the C-and

N-termini.

108

Structural analysis of N-glycans and O-glycans shall be done. It comprises

monossacharide analysis, Sialic acids characterization, sequence analysis of N-

glycans and O-glycans qualitative and quantitative basis.

The total glycan pool released from the backbone shall be subjected to

fractionation and if necessary further sub-fractionation can be performed in

order to yield sub-fractions of sufficient purity. Further analysis of the sub-

fractions shall be done to identify and/or exclude unique or unusual structures.

Site specific glycan analysis and classification of the glycans with respect to

sialylation and antennarity on each level of sub-fractionation need to be done.

Positions of N-acetyllactosamine repeats on antennae, lack of sialylation of

antennae, position of fucosylation shall be elucidated and presence of O-

acetylated groups shall be confirmed.

8.3 Preclinical issues

8.3.1 Pharmacodynamics Studies

8.3.1.1 In vitro studies

In order to assess any alterations in reactivity between the

biosimilar product and the RMP, data from a number of

comparative bioassays (e.g. receptor-binding studies, cell

proliferation assays), many of which may already be

available from quality-related bioassays, should be

provided.

8.3.1.2 In vivo studies

The erythrogenic effects of the biosimilar product and the

RMP should be quantitatively compared in an appropriate

animal assay (e.g. the European Pharmacopoeia

polycythaemic and/or normocythaemic mouse assay; data

109

may be already available from quality-related bioassays).

Additional information on the erythrogenic activity may

be obtained from the described repeat dose toxicity study.

8.3.2 Toxicological studies

Data from at least one repeat dose toxicity study in a relevant

species (e.g. rat) should be provided. Study duration should be at

least 4 weeks. In this context, special emphasis should be laid on

the determination of immune responses.

Data on local tolerance in at least one species should be provided.

If feasible, local tolerance testing can be performed as part of the

described repeat dose toxicity study.

Safety pharmacology, reproduction toxicology, mutagenicity and

carcinogenicity studies are not routine requirements for non-

clinical testing of products containing EPO as active substance.

8.4 Clinical studies

8.4.1 Pharmacokinetic studies

The relative pharmacokinetic properties of the biosimilar product and the

RMP should be determined in single dose crossover studies using

subcutaneous and intravenous administration. Healthy volunteers are

considered an appropriate study population. The selected dose should be

in the sensitive part of the dose-response curve. The primary

pharmacokinetic parameter is AUC and the secondary pharmacokinetic

parameters are Cmax and T½ or CL/F. Equivalence margins have to be

defined a priori and appropriately justified. Differences in T½ for the IV

and the SC route of administration and the dose dependence of

erythropoetin should be taken into account when designing the studies.

110

8.4.2 Pharmacodynamics studies

Pharmacodynamics should preferably be evaluated as part of the

comparative pharmacokinetic studies. The selected dose should be in

the linear ascending part of the dose-response curve. In single dose

studies, reticulocyte count is the most relevant and therefore

recommended Pharmacodynamics marker for assessment of the activity

of erythropoetin. On the other hand, reticulocyte count is not an

established surrogate marker for efficacy of erythropoietin and therefore

no suitable endpoint in clinical trials.

8.4.3 Clinical efficacy studies

The mechanism of action of erythropoietin is the same in all currently

approved indications but the doses required to achieve the desired

response may vary considerably and are highest in the oncology

indications: Erythropoietin can be administered intravenously or

subcutaneously .

Recombinant erythropoietins have a relatively wide therapeutic window

and are usually well tolerated provided, that the stimulation of bone

marrow is controlled by limiting the amount and rate of haemoglobin

increase. The rate of haemoglobin increase may vary considerably

between patients and is dependent not only on the dose and dosing

regimen of erythropoietin, but also other factors, such as iron stores,

baseline haemoglobin and erythropoietin levels, and the presence of

concurrent medical conditions such as inflammation.

Comparable clinical efficacy between the biosimilar product and the

RMP should be demonstrated in at least two adequately powered,

randomized, parallel group clinical trials.

111

Confirmatory studies should be double-blind to avoid bias. If this is not

possible, at minimum the person(s) involved in decision-making (e.g.

dose adjustment) should be effectively masked to treatment allocation.

Sensitivity to the effects of erythropoietin is higher in erythropoietin-

deficient than non erythropoietin-deficient conditions and is also

dependent on the responsiveness of the bone marrow. Patients with

renal anaemia are therefore recommended as the target study population

as this would provide the most sensitive model. Other reasons for

anemia should be excluded.

The clinical trials should include a „correction phase‟ study during

anaemia correction and a „maintain phase‟ study in patients on

erythropoietin maintenance therapy. A correction phase study is

important to determine response dynamics and dosing during the

anaemia correction phase. It should only include treatment naïve

patients or previously treated patients after a suitably long epoeitin-free

and transfusion-free period (e.g. 3 months). It is recommended that the

comparative phase be 6 months in order to establish comparable clinical

efficacy of the biosimilar product and the RMP in patients with

stabilized hemoglobin levels and erythropoietin dose. Shorter study

duration should be justified.

The study design for a maintenance phase study should minimize

baseline heterogeneity and carry over effects of previous treatments.

Patients included in a maintenance phase study should be optimally

titrated on the RMP (stable hemoglobin in the target range on stable

erythropoietin dose and regimen without transfusions) for three month.

Thereafter, study subjects should be randomized to the biosimilar

product or the RMP and followed up for a least three and, ideally, six

months to avoid carry over effects.

112

In the course of both studies, erythropoietin doses should be closely

titrated to achieve (correction phase study) or maintain (maintenance

phase study) target hemoglobin concentrations. The protocol should

clearly pre-define the dose adjustment algorithm. Hemoglobin target

range and titration schedule should be in accordance with current clinical

practice.

In the correction phase study „hemoglobin responder‟ (proportion of

patients achieving a pre-specified hemoglobin target) or „change in

hemoglobin‟ is the preferred primary endpoint. In the maintenance

phase study „hemoglobin maintenance rate‟ (proportion of patients

hemoglobin levels within a pre-specified range without transfusion) or

„change in hemoglobin‟ is the preferred primary endpoint.

Erythropoietin dosage should be a co-primary endpoint in both studies.

The fact that erythropoietin dose is titrated to achieve the desired

response reduces the sensitivity of the hemoglobin-related endpoints to

detect possible differences in the efficacy of the treatment arms.

Equivalence margins for both co-primary endpoints have to be pre-

specified and appropriately justified and serve as the basis for powering

the studies.

Transfusion requirements should be included as an important secondary

endpoint.

Since erythropoietin doses is necessary to achieve target hemoglobin

levels differ in pre-dialysis and dialysis patients, these two populations

should not be mixed in the same study.

Clinical comparability has to be demonstrated for both routes of

administration. This is best achieved by performing separate studies,

e.g. correction phase study in a pre-dialysis population using SC

113

erythropoietin and a maintenance phase study in a hemodialysis

population using IV erythropoietin.

8.4.4 Clinical Safety

Pure red cell aplasia (PRCA), due to neutralizing anti-erythropoietin

antibodies, has been observed predominantly in renal anemia patients

treated with subcutaneously administered exporting. Because antibody-

induced PRCA is a very rare event and usually takes months to years of

erythropoietin treatment to develop, such events are unlikely to be

identified in pre-authorization studies. In addition, possible angiogenic

and tumor promoting effects of erythropoietin might be of importance in

selected populations.

Comparative safety data from the efficacy trials are sufficient to provide

an adequate pre-marketing safety database.

The applicant should provide at least 12-month comparative

immunogenicity data pre-authorization. Retention samples for both

correction phase and maintenance phase studies are recommended. For

detection of anti-erythropoietin antibodies, a validated, highly sensitive

assay should be used.

8.4.5 Pharmacovigilence plan

Within the authorization procedure the applicant should present a risk

management program and pharmacovigilance plan. In order to further

study the safety profile of the biosimilar product, particularly rare

serious adverse events such as immune mediated PRCA, safety data

should be collected from a cohort of patients representing all approved

therapeutic indications.

114

Chapter 9.0

Granulocyte-Colony Stimulating Factor

115

9.1 General outline

Recombinant human granulocyte-colony stimulating factor (rhG-CSF) is a single

polypeptide chain protein of 175 amino acids and stimulates the production of white

blood cells.

Recombinant granulocyte-colony stimulating factors (rG-CSF) produced in E. coli and

in CHO cells are in clinical use. Compared to the human and to the mammalian cell

culture derived G-CSF, the E. coli protein has an additional methionine amino-terminal

and no glycosylation. The rG-CSF protein contains one free cysteinyl residue and two

disulphide bonds. rG-CSF can be used for several purposes such as (1) reduction in the

duration of neutropenia after cancer chemotherapy or myeloablative therapy followed

by bone marrow transplantation; (2) mobilisation of peripheral blood progenitor cells;

(3) treatment of severe congenital, cyclic, or idiopathic neutropenia; and (4) treatment

of persistent neutropenia in patients with advanced human immunodeficiency virus

infection. The posology varies among these conditions.

9.2 Manufacturing considerations

The active substance used to manufacture the rG-CSF as well the preparation

and characterization techniques should be described and justified.

Pharmaceutical development should represent the current state-of-the-art

methods of manufacturing and characterization, and to meet the relevant

guidelines.

The procedures followed should be same as that previously discussed earlier in

this Guideline under manufacturing consideration.

9.3 Preclinical issues

Preclinical studies should be designed to detail differences in pharmaco-

toxicological response between the biosimilar product and the RMP. At the

116

receptor level, comparability between the two should be demonstrated in in vitro

cell-based bioassays or receptor-binding assays. In vivo rodent models,

neutropenic and non-neutropenic, should compare the PD effects of the two

products. Furthermore, data from relevant toxicity studies should also be

provided. The procedures followed should be same as that previously discussed

earlier in this Guideline, taking into account the following differences:

9.3.1 Pharmacodynamic studies

9.3.1.1 In vitro studies

At the receptor level, comparability of test and RMP

should be demonstrated in appropriate in vitro cell

based bioassays or receptor-binding assays. Such data

may already be available from bioassays that were

used to measure potency in the evaluation of

biological characteristics in module 3. It is important

that assays used for comparability will have appropriate

sensitivity to detect differences and that experiments are

based on a sufficient number of dilutions per curve to

fully characterise the concentration-response relationship.

9.3.1.2 In vivo studies

In vivo rodent models, neutropenic and non-

neutropenic, should be used to compare the

pharmacodynamic effects of the test and the RMP.

9.3.2 Toxicological studies

Data from at least one repeat dose toxicity study in a relevant

species should be provided. Study duration should be at least 28

days. The study should be performed in accordance with the

requirements of the "Note for Guidance on Repeated Dose

Toxicity" (CPMP/SWP/1042/99) and include (i) pharmacodynamic

117

measurements, and (ii) appropriate toxicokinetic measurements in

accordance with the "Note for Guidance on Toxicokinetics: A

Guidance for assessing systemic exposure in toxicological studies"

(CPMP/ICH/384/95). In this context, special emphasis should be laid

on the investigation of immune responses to the products. Data on local

tolerance in at least one species should be provided in accordance with

the "Note for Guidance for Non-clinical Local Tolerance Testing of

Medicinal Products" (CPMP/SWP/2145/00). If feasible, local tolerance

testing can be performed as part of the described repeat dose toxicity

study. Safety pharmacology, reproduction toxicology, mutagenicity

and carcinogenicity studies are not routine requirements for non-

clinical testing of similar biological medicinal products containing

recombinant G-CSF as active substance.

9.4 Clinical studies

The procedures followed should be same as that previously discussed earlier in

this Guidelines under clinical studies, taking into account the following

differences:

9.4.1 Pharmacokinetic studies

The pharmacokinetic properties of the biosimilar product and the

RMP should be compared in single dose crossover studies using

subcutaneous and intravenous administration. The primary PK

parameter is AUC and the secondary PK parameters are Cmax and T1/2.

The general principles for demonstration of bioequivalence are

applicable.

9.4.2 Pharmacodynamic studies

The absolute neutrophil count (ANC) is the relevant pharmacodynamic

marker for the activity of r-GCSF. The pharmacodynamic effect of the

118

biosimilar product and the RMP should be compared in healthy

volunteers. The selected dose should be in the linear ascending part of

the dose response curve. Studies at more than one dose level may be

useful. The CD34+ cell count should be reported as a secondary PD

endpoint. The comparability range should be justified.

9.4.3 Clinical efficacy

The recommended clinical model for the demonstration of comparability

of the biosimilar product and the RMP is the prophylaxis of severe

neutropenia after cytotoxic chemotherapy in a homogenous patient

group (e.g. tumor type, previous and planned chemotherapy as well as

disease stage). This model requires a chemotherapy regimen that is

known to induce a severe neutropenia in patients. A two-arm

comparability study is sufficient in chemotherapy models with known

frequency and duration of severe neutropenia. If other chemotherapy

regimens are used, a three arms trial, including placebo, may be

needed. The sponsor must justify the comparability delta for the primary

efficacy variable, the duration of severe neutropenia (ANC below

0.5 x 109/l). The incidence of febrile neutropenia, infections and the

cumulative rG-CSF dose are secondary variables. The main emphasis

is on the first chemotherapy cycle.

Demonstration of the clinical comparability in the chemotherapy-

induced neutropenia model will allow the extrapolation of the results

to the other indications of the RMP if the mechanism of action is the

same. Alternative models, including pharmacodynamic studies in

healthy volunteers, may be pursued for the demonstration of

comparability if justified. In such cases, the sponsor should seek

for scientific advice for study design and duration, choice of doses,

efficacy and pharmacodynamic endpoints, and comparability margins.

119

9.4.4 Clinical safety

Safety data should be collected from a cohort of patients after

repeated dosing preferably in a comparative clinical trial. The total

exposure should correspond to the exposure of a conventional

chemotherapeutic treatment course with several cycles. The total follow

up of patients should be at least 6 months. The number of patients

should be sufficient for the evaluation of the adverse effect profile,

including bone pain and laboratory abnormalities.

9.4.5 Risk management

Antibodies to the currently marketed E. coli derived rG-CSF occur

infrequently. Attention should be paid to immunogenicity and potential

rare serious adverse events, especially in patients undergoing chronic

administration. Lack of efficacy should also be monitored, especially in

individuals undergoing haematopoietic progenitor cell mobilisation.

9.4.6 Pharmacovigilence plan

Within the authorization procedure the applicant should present a

risk management programme and pharmacovigilance plan in

accordance with current EU legislation and pharmacovigilance

guidelines.

Attention should be paid to immunogenicity and potential rare serious

adverse events, especially in patients undergoing chronic administration.

Lack of efficacy should also be monitored, especially in individuals

undergoing haematopoietic progenitor cell mobilisation.

120

9.5 Summary

Clinical studies should be planned to compare the pharmacokinetic (mainly AUC) and

pharmacodynamic (the absolute neutrophil count being the relevant biomarker)

properties of the biosimilar product and the RMP. Recombinant G-CSF can be used for

several purposes including mobilisation of peripheral blood progenitor cells and the

treatment of neutropenia itself, and after cancer chemotherapy, and in patients with HIV

infection. The recommended clinical model for demonstrating the comparability of the

biosimilar product and the RMP is the prophylaxis of severe neutropenia after

chemotherapy in a homogenous patient group. Demonstration of clinical comparability

in this model will allow extrapolation of the results to the other indications of the RMP

if the mechanism of action is the same. Although, if justified, alternative models,

including PD studies in healthy volunteers may be used to demonstrate comparability.

Clinical safety data should also be collected from patients after repeated dosing in a

comparative clinical trial, with a follow-up of at least six months. Immunogenicity data

should be collected, and also monitored in the pharmacovigilance plan.

121

Chapter 10.0

Human Growth Hormone

(Somatropin)

122

10.1 General outline

Human growth hormone (hGH) molecule is a single chain, non-glycosylated, 22 kD

polypeptide with 191 amino acid, produced in the anterior pituitary gland. Growth

hormone (GH) for clinical use has an identical amino acid sequence and is produced by

recombinant technology using E. coli, mammalian cells or yeast cells as expression

system (rhGH). The structure and biological activity of GH can be characterized by

appropriate physicochemical and biological methods. Several techniques and bioassays

are available to characterize both the active substance and product-related substances or

impurities, such as deamidated and oxidized forms and aggregates. GH has potent

anabolic, lipolytic and anti-insulin effects (acute insulin-like effect). The effects of GH

are mediated both directly (e.g. on adipocytes and hepatocytes) and indirectly via

stimulation of insulin-like growth factors (principally IGF-1). GH-containing medicinal

products are currently licensed for normalizing or improving linear growth and/or body

composition in GH-deficient and certain non-GH-deficient states. The same receptors

are thought to be involved in all currently approved therapeutic indications of rhGHs.

GH has a wide therapeutic window in children during the growth phase whereas adults

may be more sensitive for certain adverse effects. Antibodies to GH have been

described, including, very rarely, neutralizing antibodies. Problems have been

associated with the purity and stability of the formulations. GH is administered

subcutaneously. Possible patient-related risk factors of immune response are unknown.

10.2 Manufacturing considerations

The manufacture, preparation and characterization techniques should be

described and justified.

10.3 Preclinical issues

Before initiating clinical development, non-clinical studies should be performed.

These studies should be comparative in nature and should be designed to detect

differences in the pharmaco-toxicological response between the biosimilar

product and the RMP (the innovator‟s), and should not just assess the response

123

per se. The approach taken will need to be fully justified in the preclinical

overview.

10.3.1 Pharmacodynamics studies

10.3.1.1 In vitro studies

In order to assess any alterations in reactivity

between the biosimilar and the RMP, data from a

number of comparative bioassays (e.g. receptor-

binding studies, cell proliferation assays), many of

which may already be available from quality-

related bioassays, should be provided.

10.3.1.2 In vivo studies

An appropriate in vivo rodent model (e.g. the

weight-gain assay and/or the tibia growth assay in

immature hypophysectomized rats; data may

already be available from quality-related

bioassays) should be used to quantitatively

compare the pharmacodynamic activity of the

biosimilar and the RMP.

10.3.2 Toxicological studies

Data from at least one repeat dose toxicity study in a relevant

species (e.g. rat) should be provided. Study duration should be at

least 4 weeks. The study should be performed in accordance with

the requirements of the "Note for guidance on repeated dose

toxicity" (CPMP/SWP/1042/99) and include appropriate

toxicokinetic measurements in accordance with the "Note for

guidance on toxicokinetics: A Guidance for assessing systemic

exposure in toxicological studies" (CPMP/ICH/384/95). In this

context, special emphasis should be laid on the determination of

124

immune responses. Data on local tolerance in at least one species

should be provided in accordance with the "Note for guidance on

non-clinical local tolerance testing of medicinal products"

(CPMP/SWP/2145/00). If feasible, local tolerance testing can be

performed as part of the described repeat dose toxicity study.

Safety pharmacology, reproduction toxicology, mutagenicity and

carcinogenicity studies are not routine requirements for non-

clinical testing of biosimilar products containing rhGH as active

substance.

10.4 Clinical studies

10.4.1 Pharmacokinetic studies

The relative pharmacokinetic properties of the biosimilar product and the

RMP should be determined in a single dose crossover study using

subcutaneous administration. Healthy volunteers are considered

appropriate, but suppression of endogenous GH production (e.g. with a

somatostatin analogue) may be considered. The primary

pharmacokinetic parameter is AUC and the secondary parameters are

Cmax

and T1/2

. Comparability margins have to be defined a priori and

appropriately justified.

10.4.2 Pharmacodynamic studies

Pharmacodynamics should preferably be evaluated as part of the

comparative pharmacokinetic study. The selected dose should be in the

linear ascending part of the dose-response curve. IGF-1 is the preferred

pharmacodynamic marker for the activity of GH and is recommended to

be used in comparative pharmacodynamic studies. In addition, other

markers such as IGFBP-3 may be used. On the other hand, due to the

lack of a clear relationship between serum IGF-1 levels and growth

125

response, IGF-1 is not a suitable surrogate marker for the efficacy of a

GH in clinical trials.

10.4.3 Clinical efficacy

Clinical comparability efficacy between the similar biological medicinal

product and the RMP should be demonstrated in at least one adequately

powered, randomized, parallel group clinical trial. Clinical studies

should be double-blind to avoid bias. If this is not possible, at minimum

the person performing height measurements should be effectively

masked to treatment allocation. Sensitivity to the effects of GH is higher

in GH-deficient than non-GH-deficient conditions. Treatment-naïve

children with GH deficiency are recommended as the target study

population as this provides a sensitive and well-known model. Study

subjects should be pre-pubertal before and during the comparative phase

of the trial to avoid interference of the pubertal growth spurt with the

treatment effect. This may be achieved by limiting the age/bone age at

study entry. It is important that the study groups are thoroughly balanced

for baseline characteristics, as this will affect the sensitivity of the trial

and the accuracy of the endpoints. Change in height velocity or change

in height velocity standard deviation score from baseline to the pre-

specified end of the comparative phase of the trial is the recommended

primary efficacy endpoint. Height standard deviation score is a

recommended secondary endpoint. Adjustment for factors known to

affect the growth response to GH should be considered. During the

comparative phase of the study, standing height should be measured at

least 3 times per subject at each time point and the results averaged for

analyses. The use of a validated measuring device is mandatory.

Consecutive height measurements should be standardized and performed

approximately at the same time of the day, by the same measuring

device and preferably by the same trained observer. These

recommendations aim to reduce measurement errors and variability. For

the determination of reliable baseline growth rates, it is important that

126

also height measurements during the pre-treatment phase are obtained in

a standardized manner using a validated measuring device. Due to

significant variability in short-term growth, seasonal variability in

growth and measurement errors inherent in short-term growth

measurements, the recommended duration of the comparative phase is at

least 6 months and may have to be up to 12 months. Calculation of pre-

treatment growth rates should be based on observation periods of no less

than 6 and no more than 18 months. Comparability margins have to be

pre-specified and appropriately justified, primarily on clinical grounds,

and serve as the basis for powering the study.

10.4.4 Clinical safety

Data from patients in the efficacy trial(s) are usually sufficient to provide

an adequate pre-marketing safety database. The applicant should provide

a comparative, 12-month immunogenicity data of patients who

participated in the efficacy trial(s), with sampling at 3-month intervals

and testing using validated assays of adequate specificity and sensitivity.

In addition, adequate blood tests including IGF-1, IGFBP-3, fasting

insulin and blood glucose should be performed.

127

CHAPTER 11.0

Pharmaceutical Formulations of Biosimilars

Drug companies may choose to introduce different formulation of an existing

recombinantly-produced medicine for various purposes, the most important of which is

to make it long acting, thereby reducing the administration of injectable or other

medicines.

128

11.1 Pegylated rIFN-

11.1.1 General outline

Pegylated interferon is made with special add-on parts that help the drug stay in the

body longer. It is injected once a week, while interferon that is not pegylated needs to

be injected three times a week. The active substance is a polyethylene glycol-modified

(pegylated) derivative of IFN-. It is designated as PEG-rhIFN-. The modification

was developed in order to decrease the systemic clearance of the active moiety. The

RMPs are those of the innovators. All PEG-rhIFN- products are currently used in

combination with ribavirin to treat hepatitis C virus (HCV) infection patients, with

histologically proven chronic hepatitis C who have elevated transaminases without liver

decompensation and who are positive for serum HCV-RNA or anti-HCV.

11.1.2 Manufacturing considerations

The active substance is the same that is used to manufacture rhIFN-. Pegylated

rhIFN-is prepared by reacting rhIFN-with activated methoxypoly-[ethylene

glycol], commonly referred to as mPEG. The reaction involves the formation of

a covalent bond between the mPEG and amino groups on the IFN-molecule.

Appropriate molecular size characterization techniques (mass spectroscopy,

SDS-PAGE and size exclusion chromatography) should be used to confirm that

pegylated rhIFN-is predominantly composed of monopegylated species, with

small amounts of dipegylated species and free interferon. Studies on the

conjugation chemistry, the structural elucidation and the degradation pathways

should represent the current state of art. The resolution of the separation process

does not allow for base-line separation, and therefore fraction selection should

be controlled.

129

11.1.3 Preclinical issues

Cell or animal model for chronic hepatitis C infection is lacking. The

assessment is also restricted by the fact that rhIFN-is inactive in rodents, and

the activity is comparatively low in primates available for preclinical studies.

11.1.3.1 Pharmacodynamics

The binding of interferon to specific cell surface receptor

molecules signals the cell to produce a series of antiviral

proteins. Most of this act to inhibit the translation of viral

proteins, but other steps in viral replication is also affected. The

comparisons of in vitro antiviral and immune system related

effects should be studied between pegylated and nonpegylated

rhIFN-.

11.1.3.2 Safety pharmacology

Cardiovascular, gastrointestinal, CNS and renal effects can be

studied in rats and cynomolgus monkeys.

11.1.3.3 Pharmacokinetics

The bioavailability, plasma half-life and other PK parameters

after subcutaneous injections can be studied in rats and monkeys

and compared with that of the innovator‟s.

11.1.3.4 Toxicology

Single dose and repeated toxicity studies should be conducted in

mice, rats and monkeys using up to several hundred times the

intended clinical dose of pegylated rhIFN-and compared to

those produced by non-pegylated rhIFN-to ensure that there is

130

no unique toxicity due to the pegylation process. Reproduction

studies should be performed, especially because some interferons

have been reported to be abortifacient in primates and to ensure

that pegylation has no such effect. Genotoxicity should be

studied using a standard battery of tests. Carcinogenicity studies

are not obligatory.

11.1.4 Clinical studies

11.1.4.1 Pharmacodynamic markers

At least one PD marker should be accepted as a surrogate marker

for efficacy. In case of IFN pharmacodynamics, the changes in

concentrations of effector proteins such as serum neopterin and

2‟5‟-oligoadenylate synthetase (2‟5‟- OAS) are important PD

markers. PD markers (such as serum neopterin concentration,

neutrophil and white cell count) should respond in a dose-related

manner at the end of Week 4.

11.1.4.2 Pharmacokinetics

The ordinary crossover design is not appropriate for

pegylated proteins, because it has a long half-life. The single-

and multiple dose PK of rhIFN- should be evaluated in the

target population.

11.1.4.3 Clinical efficacy

This parameter should be investigated in various dose-regimens

of PEG-rhIFN-with the approved dose of the innovator in

patients not previously treated with an interferon. Patients with

decompensated liver function are not eligible in these clinical

studies.

131

11.2 Pegylated rhG-CSF

11.2.1 Introduction

The pegylated form is a covalent conjugate of rhG-CSF with a single 20 kDa

polyethylene glycol (PEG).

11.2.2 Manufacturing considerations

The pegylated form can be manufactured by attaching a 20 kDa methoxy-

polyethylene glycolpropionaldehyde (PEG-aldehyde) to the N-terminal amino

acid of rhG-CSF (175 amino acids). The biological activity should be

characterized and compared to innovator product. A full description of the

pegylation reaction and its controls should be provided.

Following pegylation, purification should be carried out using cation exchange

chromatography to result in purified bulk product pegylated form. The active

substance is stored in sterile polypropylene containers at 2–8 °C and shipped at

the same temperature of 2–8 °C.

11.2.3 Preclinial issues

The pegylated form has to undergo preclinical studies, which are valid both on a

stand-alone basis and also as a “bridge” to the non-pegylated product. All safety

studies were undertaken in accordance with GLP.

11.2.3.1 Pharmacodynamics

The pharmacological effects of the pegylated form have to be

investigated in in vitro and in vivo models using innovator

product for comparability purposes.

132

11.2.3.1.1 In-vitro studies

The ability of pegylated form of rhG-CSF to

stimulate mature neutrophil functions (oxidative

burst, phagocytosis, chemotaxis, etc..) should be

comparable to innovator product. In addition, the

clearance of both materials have to be via similar

receptor-mediated and non-specific mechanisms.

11.2.3.1.2 In-vivo studies

The pegylated form of rhG-CSF has to be

effective in restoring neutrophil counts in several

mouse chemotherapy models and in a monkey

model of myeloablation.

11.2.3.2 Pharmacokinetics

Single-dose kinetic studies in mice, rats, rhesus and cynomolgus

monkeys all should show a sustained dose-related increase in

blood neutrophils. Repeat-dose studies should reveal the same.

The pharmacokinetic properties of the pegylated form of the

biosimilar product and the RMP should be compared in

single dose and repeated dose studies using subcutaneous and

intravenous administration. The general principles for

demonstration of bioequivalence are applicable.

11.2.3.3 Toxicology

11.2.3.3.1 Single and repeat dose toxicity

A full set of conventional toxicity tests should be

performed for the pegylated form

133

The duration of repeat-dose toxicity studies in rats

(weekly dosing for up to 6 months) and

cynomolgus monkeys (weekly dosing for 4

weeks). Toxicokinetic investigations can be

carried out as a part of the repeat-dose studies.

11.2.3.3.2 Mutagenicity

Given the chemical structure and bioreactivity of

the pegylated form, it is considered inappropriate

to undertake genetic toxicity studies, which is

consistent with ICH Guidelines on products of

biotechnological origin.

11.2.3.3.3 Carcinogenicity

The pegylated form is most unlikely to be

carcinogenic, so no need for experimental

evaluation of carcinogenic potential

11.2.3.3.4 Immunogenicity

Immunogenicity has to be determined in

pharmacodynamic and repeat toxicity studies as

well as in clinical studies.

11.2.4 Clinical studies

As described under rhG-CSF.

134

DDRRUUGG MMAASSTTEERR

FFIILLEE

RREEQQUUIIRREEMMEENNTTSS

FOR THE

REGISTRATION OF

BIOSIMILARS (FOLLOW-ON PROTEINS)

135

Scope

This document is intended to provide guidance on the format of a registration

application for biosimilar drug substances and their corresponding drug products.

At the end of this requirements document, detailed explanation for some of the

processes (mainly the manufacturing/quality processes and the clinical studies

processes). The text following the section titles is intended to be explanatory and

illustrative only. The content of these sections should include relevant information

described in the current existing ICH guidelines, but harmonized content is not

available for all sections. The`` Body of Data`` in this document merely indicates where

the information should be located.

The type and the extent of specific supporting data must not be limited to what is

addressed in this document, which is only for explanatory purpose. The type and

the extent of specific supporting data depend on the nature of the product and on

the product specific requirements as well as advancement in the technology related

to the product manufacture and analysis.

136

GGUUIIDDEE FFOORR

RREEGGIISSTTRRAATTIIOONN

RREEQQUUIIRREEMMEENNTTSS

137

1. Administrative information

1.1 Cover letter

1.2 Trade name

1.3 Generic name

1.4 Expiry date

1.5 Other trade names of the similar product

1.6 Pharmaceutical form

1.7 Name of manufacturing company

1.7.1 Name of active substance manufacturer (if different from above)

1.8 Status of the company in the GCC country intended for

application (currently registered or not registered)

1.9 Local agent

1.9.1 Name

1.9.2 Address

1.10 Marketing status at country of origin and other countries

1.11 Similar products registered localy

1.12 Reasons for production

1.13 Advantages over other similar drugs (if any)

1.14 Reference medicinal product (RMP) – The innovator

(In addition to 2.9 and 3.6)

1.14.1 Name

1.14.2 Approval at the regulatory authority and/or EMEA

1.14.3 Company

2. Active pharmaceutical ingredient (drug substance)

2.1 General Information

2.1.1 Nomenclature

2.1.2 Structure

2.1.3 General Properties

138

2.2 Manufacturing

2.2.1 Manufacturer(s)

2.2.2 Description of manufacturing process and process specifications

2.3 Control of materials

2.3.1 Control of source and starting materials

2.3.2 Source, history, and generation of the cell substrate

2.3.3 Generation of cell substrate

2.3.4 Cell banking

2.4 Controls of critical steps and intermediates

2.4.1 Critical steps

2.4.2 Intermediates

2.5 Process validation and verification

2.6 Manufacturing process development

2.7 Comparability data of the structure elucidation and other quality

characteristics of the molecule against a reference medicinal product

2.7.1 Characterization: Elucidation of structure and other characteristics

2.7.2 Impurities

2.8. Control of drug substance

2.8.1 Specification

2.8.2 Analytical procedures

2.8.3 Validation of analytical procedures

2.8.4 Batch analyses

2.8.5 Justification of specification

2.9 Reference standards or materials

2.10 Container Closure System

2.11 Stability

2.11.1 Stability summary and conclusions

2.11.2 Post-approval stability protocol and stability commitment

2.11.3 Stability data

139

3. Drug product

3.1 Description and composition of the drug product

3.2 Pharmaceutical Development

3.2.1 Components of the drug product

3.2.2 Drug Product

3.2.3 Manufacturing process development

3.2.4 Container closure system

3.2.5 Microbiological attributes

3.2.6 Compatibility

3.3 Manufacturing

3.3.1 Manufacturer(s)

3.3.2 Batch formula

3.3.3 Description of manufacturing process and process controls

3.3.4 Controls of critical steps and intermediates

3.3.5 Process validation and/or evaluation

3.4 Control of excipients

3.4.1 Specifications

3.4.2 Analytical procedures (name, dosage form)

3.4.3 Validation of analytical procedures

3.4.4 Justification of specifications

3.4.5 Excipients of human or animal origin

3.4.6 Novel excipient

3.5 Control of drug product

3.5.1 Specification(s)

3.5.2 Analytical procedures

3.5.3 Validation of analytical procedures

3.5.4 Batch analyses

3.5.5 Characterization of impurities

3.5.6 Justification of specification(s)

3.6 Reference standards or materials

3.7 Packaging materials

3.7.1. Container closure system.

140

3.7.2. Product package insert/product leaflet.

3.8 Stability

3.8.1 Stability summary and conclusion

3.8.2 Post-approval stability protocol and stability commitment

3.8.3 Stability data

3.9 Appendices

3.9.1 Changes reporting

3.9.2 Facilities and equipment

3.9.3 Adventitious agents’ safety evaluation

3.9.3.1 For non-viral adventitious agents

3.9.3.2 For viral adventitious agents

3.9.4 Materials of biological origin

3.9.5 Testing at appropriate stages of production

3.9.6 Viral testing of unprocessed bulk

3.9.7 Viral clearance studies

3.10 List of used excipients

3.10.1 Regional information

3.10.2 Literature references

4. Pre-Clinical comparative study with the RMP

4.1 Preclinical testing

4.1.1. Selected relevant animal species (number/gender)

4.1.2. Delivery, dose and route of administration

4.2 Pharmacology/pharmacodynamics

4.3 Pharmacokinetics

4.4 Toxicological studies

4.4.1. Single dose toxicity

4.4.2. Repeated dose toxicity studies

4.4.3. Local tolerance

4.5 Immunogenicity profile

141

5. Clinical comparative study with the RMP

5.1 Protocol

5.2 Recruitment details

5.3 Eligibility criteria

5.4 Clinical studies reports

5.4.1 Reports on biopharmaceutic studies

5.4.2 Reports of studies pertinent to pharmacokinetics using human

biomaterials

5.4.3 Reports on pharmacokinetics (PK)

5.4.4 Reports on pharmacodynamic( PD)

5.4.5 Reports on efficacy and safety

5.5 Immunogenecity findings

5.6 Statistics (justification of statistical method used)

5.7 Literature references

6. Pharmacovigilance plan

6.1 Pharmacovigilance plan (track and trace)

6.2 Recall plan

6.3 Plan for adverse reactions (ADR) reports

6.4 Plan to ensure quality of the product (defect, final formulation package)

6.5 Bar-coding method

6.6 Post approval stability protocol and stability commitments

7. Certified Documents

7.1 Good manufacturing practice (GMP) certificates

7.2 Each raw material

7.3 Product analysis

142

7.4 Product composition

7.5 Diluents and coloring materials

7.6 Absence of alcohol content in the finished product

7.7 Absence of animal materials in the finished product

7.8 Package insert approval at country of origin

7.9 Registration and marketing at country of origin and other countries

7.10 Pricing at country of origin

7.11 Company from which raw material(s) was obtained

8. Other necessary activities

Site visit to the manufacturing facility, line of production and the raw

material source(s) manufacturers (if different from the drug manufacturer)

is mandatory

143

EXPLANATION

OF SOME OF THE

REQUIREMENTS

144

2. Active Pharmaceutical Ingredient (drug substance)

2.1 General Information

2.1.1 Nomenclature

• Recommended International Nonproprietary Name (INN).

• Compendial name (e.g. European Pharmacopoeia) if relevant.

• Chemical name(s).

• Company or laboratory code.

• Other non-proprietary name(s), e.g., national name, United States

Adopted Name

(USAN), Japanese Accepted Name (JAN); British Approved Name

(BAN), and

Chemical Abstracts Service (CAS) registry number.

2.1.2 Structure

The schematic amino acid sequence indicating glycosylation sites or other post-

translational modifications and relative molecular mass should be provided, as

appropriate.

2.1.3 General Properties:

A list should be provided for physicochemical and other relevant properties of

the drug substance such as the listed below:

A. Physicochemical properties: Composition, solubility, pH, osmolality,

color, clarity, and others (if any)

B. Immunochemical properties: Identity test, amino acid sequencing, N-

terminal amino acid sequencing, SDS-

PAGE (molecular weight), and Western

blot.

145

C. Biological activity: Pharmacological action, potency, in vitro

(ELISA), and in vivo (biological assay).

D. Purity: SDS-PAGE, Western blot, analytical

HPLC, host cell DNA detection assay, and

host cell protein (HCP) assay.

Isoforms determination using isoelectric

focusing (IEF) and capillary zone

electrophoresis (CZE), where applicable.

E. Carbohydrate content: Where applicable.

F. Pyrogeneicity: Rabbit test.

G. Bioburden and sterility tests

H. Test for dimers and multimers

2.2 Manufacturing

2.2.1 Manufacturer(s):

The name, address, and responsibility of each manufacturer, including

contractors, and each proposed production site or facility involved in

manufacturing and testing should be provided, as follows:

A. Name and address of manufacturer

B. Contract manufacturing laboratory (if applicable)

C. Testing facilities

D. Batch release site

E. Drug substance producing plant address

F. Drug product plant/site information address

G. Plant dedication

146

2.2.2 Description of manufacturing process and process specifications

A. Description of manufacturing process and process controls

The description of the drug substance manufacturing process represents

the applicant‟s commitment for the manufacture of the drug substance.

Information should be provided to adequately describe the

manufacturing process and process controls.

For example: Information should be provided on the manufacturing

process, which typically starts with a vial(s) of the cell bank, and

includes cell culture, harvest(s), purification and modification reactions

filling, storage and shipping conditions.

B. Batch (es) and scale definition

An explanation of the batch numbering system, including information

regarding any pooling of harvests (if any) or intermediates and batch size

or scale should be provided.

C. Cell culture and harvest:

A flow diagram should be provided that illustrates the manufacturing

route from the original inoculum (e.g. cells contained in one or more

vials(s) of the Working Cell Bank up to the last harvesting operation).

The diagram should include all steps (each unit operations) and

intermediates.

A flowchart of the process with In Process Control Tests (IPCs),

including:

i. Summaries of cell culture process

ii. Graphs for fermentation

iii. Operating Parameters

147

Relevant information for each stage, such as population doubling levels,

cell concentration, volumes, pH, cultivation times, holding times, and

temperature, should be included.

Critical steps and critical intermediates for which specifications are

established, should be identified.

A description of each process step in the flow diagram should be

provided.

Information should be included on, (for example: scale; culture media

and other additives; major equipment; and process controls, including in-

process tests and operational parameters, process steps, equipment and

intermediates with acceptance criteria.)

Information on procedures used to transfer material between steps,

equipment, areas, and buildings, as appropriate, and shipping and storage

conditions should be provided.

D. Purification and modification reactions

A flow diagram should be provided that illustrates the purification steps

(each unit operations) from the crude harvest(s) up to the step preceding

filling of the drug substance.

A flowchart of the process with In Process Control Tests (IPCs):

i. Summaries of purifications process

ii. Graphs for purification process

iii. Description for operating parameters per step

All steps and intermediates and relevant information for each stage (e.g.,

volumes, pH, critical processing time, holding times, temperatures and

elution profiles and selection of fraction, storage of intermediate, if

148

applicable) should be included. Critical steps for which specifications

are established should be identified.

A description of each process step (as identified in the flow diagram)

should be provided. The description should include information on for

example: scale, buffers and other reagents, major equipment, and

materials.

For materials such as membranes and chromatography resins,

information for conditions of use and reuse also should be provided.

Validation studies for the reuse and regeneration of columns and

membranes should be mentioned. The description should include process

controls (including in-process tests and operational parameters) with

acceptance criteria for process steps, equipment and intermediates.

If any reprocessing is required, procedures with criteria for reprocessing

of any intermediate or the drug substance should be described.

Information on procedures used to transfer material between steps,

equipment, areas, and buildings, as appropriate, and shipping and storage

conditions should be provided.

Proof of reproducibility of purification steps should include tables for

step recoveries for 3 validation batches.

F. Filling, storage and transportation (shipping)

A description of the filling procedure for the drug substance, process

controls (including in-process tests and operational parameters), and

acceptance criteria should be provided.

149

The container closure system(s) used for storage of the drug substance

and storage and shipping conditions for the drug substance should be

described. For detailed information, refer to ICH Q5A, ICH Q5B, and

ICH Q6B.

2.3 Control of materials

2.3.1 Control of source and starting materials

Materials used in the manufacture of the drug substance (e.g., raw materials,

starting materials, solvents, reagents, catalysts) should be listed, identifying

where each material is used in the process, including raw materials used in the

cell culture process and raw materials used in the purification process.

Information on the quality and control of all materials should be provided,

including the non-pharmacopoeial materials.

Information demonstrating that materials including biologically-sourced

materials (media component, like fetal bovine serum, trypsin, some other

enzymes,) meet standards appropriate for their intended use (including the

clearance or control of adventitious agents) should be provided, as appropriate.

For biologically-sourced materials, this can include information regarding the

source, manufacture, and characterization, as required by ICH Q6A and ICH

Q6B.

2.3.2 Source, history, and generation of the cell substrate

Information on the source of the cell substrate and analysis of the expression

construct used to genetically modify cells and incorporated in the initial cell

clone used to develop the Master Cell Bank should be provided as described in

150

ICH Q5B and ICH Q5D. That must include source of the cell line, species and

strain, breeding conditions, tissue or organ of origin, geographical origin, age

and sex of the donor, test for pathogenic agents, general physiological condition

of donor, documentation of cultivation history of the cells, methods and

procedures used for isolation cells, and description of any genetic manipulation.

For continuous cell line, population doubling time or number of subcultivation

at defined dilution ratio is required.

2.3.3 Generation of cell substrate

Description of such a generation should include:

Untransfected cell line

Procedures for generation of cell substrate (which ever is applicable):

Cell fusion

Transfection

Transformation

Selection

Colony isolation

Cloning

2.3.4 Cell banking

Information on the cell banking system, cell banking quality control testing

activities, and cell line stability during production and storage (including

procedures used to generate the Master and Working Cell Bank(s) should be

provided as required by ICH Q5A, ICH Q5B, ICH Q5C and ICH Q5D. Data on

the source of the cell banks (in-house developed or from an external source)

must be provided. If obtained from external source, all data for generation of cell

substrate must be provided.

151

(A) Cell banking system

Two tiered or single tiered system

Strategy for supply of cell bank (preparation of new cell bank):

Working cell bank testing and record of consumption :

Well defined master and working cell bank

Cell line history and cell bank production

Reagents used during cell culture

Methods used during cell culture

In vitro cell age

Storage conditions

Phenotypic and genotypic markers

Restriction endonuclease mapping of expression vector

Analysis of gene copy number

Analysis for insertion or deletion sequence

Number of insertion sites

For chromosomal expression system, the percent of host cell

retaining the expression construction should be determine

Confirmation of protein coding sequence in expression vector

For cell with chromosomal copies of the expression construct

the nucleotide:

o sequence of the coding sequence could e verified by

re-cloning

o sequencing of chromosomal copies

Alternatively, nucleic acid sequence for EPO could be verified by

techniques such as sequencing the pooled cDNA clones or material

amplified by polymerase chain reaction.

(B) Cell banking procedures

Contamination preventive procedures adopted during cell banking

Type of banking system (vial or ampoule).

Crypoprotectant and media used for the cell banking.

Banking procedure (cell expansion, pooling and transferring to vial or

ampoules).

152

Aliquoting and freezing process.

Storage strategy and storage conditions.

(C) General principles of characterization and testing of cell banks

Isoenzyme analysis (if applicable)

Banding cytogenecity/ species specific antisera

RFLP (DNA Banding pattern)

Bioburden test/ sterility test

Mycoplasma test

(E) Virus detection test (as per ICH guideline Q5A)

MCB Endogenous viruses

MCB Adventitious viruses

WCB or EPCB (End of Production Cell Bank ) endogenous viruses

WCB/ EPCB adventitious viruses

If more than one cell was handled during cell banking, the cell bank

should be tested for the presence of other cell lines or products

Mouse antibody production for MCB

(F) Cell substrate stability:

Comparative study WCB/EPCB (required once to be executed on one

batch of EPCB):

Consistency in DNA coding sequence in expression system:

Morphological characteristics

Growth characteristic

Biochemical markers

Immunological marker

Productivity of desired product

153

(G) Characterization of expression construct (ICH Q5B)

Nucleotide coding sequence for the protein

Source of nucleotide sequence from which it has been taken

Methods used for preparing DNA coding for the protein gene

Assembly of expression construct

Origin or replication

Antibiotic resistance genes

Promoters

Enhancer

Fusion protein

Flaking region

Site (junction) of insertions

Complete map of the plasmid sequence:

Other proteins expressed by the same plasmid (if any)

Methods used for transfer of expression vector into a host

Methods used for amplifying the expression construct

Selection criteria for selecting the clone for production

(H) End of Production Cell Bank (EPCB)

Cells should be from Pilot of full scale production batch

Coding sequence of expression system of production cells could be

verified by nucleic acid testing or final protein product.

2.4 Controls of critical steps and intermediates

2.4.1 Critical steps

Tests and acceptance criteria (with justification including experimental data)

performed at critical steps of the manufacturing process to ensure that the

process is controlled should be provided, such as:

Cell culture process monitoring:

154

Tables for in-process control tests (IPC) with controls

Summary of process controls/in-process control during

purification

Limits of in-process controls during purification

2.4.2 Intermediates

Information on the quality control of intermediates isolated during the process

should be provided, as per ICH Q6A and ICH Q6B. Stability data supporting

storage conditions should be provided, as per ICH Q5C, section 4.1, which

includes flow chart for critical parameters and sampling plan at different phases

and different process

2.5 Process validation and verification

Process validation and/or evaluation studies for aseptic processing and sterilization

should be included.

Sufficient information should be provided on validation and evaluation studies to

demonstrate that the manufacturing process (including reprocessing steps) is suitable

for its intended purpose and to substantiate selection of critical process controls

(operational parameters and in-process tests) and their limits for critical manufacturing

steps (e.g., cell culture, harvesting, purification, and modification).

The plan for conducting the validation study should be described and the results,

analysis and conclusions from the executed study(ies) should be provided.

The analytical procedures and corresponding validation should be cross-referenced or

provided as part of justifying the selection of critical process controls and acceptance

criteria.

155

For manufacturing steps intended to remove or inactivate viral contaminants, the

information from evaluation studies should be provided. The followings are examples:

Validation of media preparation

Validation of buffers preparation

Validation of cell culture process and fermentation

Validation of purification process

Validation of intermediate hold studies

Validation of virus clearance studies

Cleaning validation

Validation of mixing studies

Validation of media hold time

Validation of buffers hold time

Validation of column and resin storage studies

Validation of resin leachable studies

Validation of cell bank remote transfer

Drug substance shipping validation

Validation of sterilization processes

Process equipment qualifications

2.6 Manufacturing process development

The developmental history of the manufacturing process should be provided.

The description of change(s) made to the manufacture of drug substance batches used in

support of the marketing application should include, for example, changes to the

process or to critical equipment.

The reason for the change should be explained.

156

Relevant information on drug substance batches manufactured during development,

such as the batch number, manufacturing scale, and their use for: (e.g., stability,

preclinical, reference material) in relation to the change, should be provided.

The significance of the change should be assessed by evaluating its potential to impact

the quality of the drug substance (and/or intermediate, if appropriate), as per ICH Q6B.

For manufacturing changes that are considered significant, data from

comparative analytical testing on relevant drug substance batches should

be provided to determine the impact on quality of the drug substance

A discussion of the data, including a justification for selection of the

tests and assessment of results, should be included.

Testing used to assess the impact of manufacturing changes on the drug substance(s)

and the corresponding drug product(s) can also include nonclinical and clinical studies.

2.7 Comparability data of the structure elucidation and other

quality characteristics of the molecule against a reference

product

2.7.1 Characterization: Elucidation of Structure and other Characteristics

For desired product and product-related substances, details should be provided on

primary, secondary and higher-order structure, post-translational forms (e.g.,

glycoforms), biological activity, purity, and immunochemical properties, when relevant.

As part of the requirement of the Comparability Exercise, characterization shall be

performed in comparison with RMP, as per ICH Q6B.

157

2.7.2 Impurities

Information on impurities should be provided. Below is example of what should be

shown. Refer to ICH Guidelines for further details.

(A) Table of Content

(B) Product related protein (PRP)

Identification and charecterization

Structural and physicochemical carchterization

Biological activity (in vitro)

Identifcation of PRP arising from degradation of drug substances

Peptide mapping of drug substances stress (DSS)

SDS-PAGE anlysis for DSS

IEF analysis of DSS (if applicable)

RP-HPLC analysis

DEAE-HPLC analysis

SEC (size exclusion chromatography)-HPLC analysis

(C) Control of prodcut related proteis

(D) Potential impurities derived from host cells

Preparationof HCP (host cell proteins)

Preapration of polyclonal anti HCP antibody

Characterizationof anti HCP adtisera (antobodies)

Testing of residual HCP

Removal of HCP

Removal of Host cell DNA

Aminopeptidase

(E) Potential impurities derived from cell culture media componant

(F) Potential impurities and extracts and others, such as antifoming agent

used druing fermentation

(G) Potential impurites derived from downsream, such as column leachates

and chromatography and ultrafiltration reagents (solvent, contamiation,

bioburdden endotoxin)

158

2.8. Control of Drug Substance

2.8.1 Specification

The specification for the drug substance should be provided, as per ICH Q6B.

2.8.2 Analytical Procedures

The analytical procedures used for testing the drug substance should be provided:

Protein estimation and quantitation (ICH Q2A [section 2], ICH Q2B [section

2.2.]).

Purity and identity tests

Activity and potency test, both in vitro and in vivo

Bacterial endotoxin

Sterility and bioburden

Glycoproteins content

N-terminal aminoacid sequenceing

2.8.3 Validation of Analytical Procedures

Analytical validation information, including experimental data for the analytical

procedures used for testing the drug substance, should be provided (ICH Q2A

[section 2], ICH Q3C [section 4], and ICH Q6B [section 2.2.2]).

2.8.4 Batch Analyses

Description of batches and results of batch analyses should be provided (ICH

Q3A, ICH Q3C, ICH Q6A, ICH Q6B, ICH Q3A [section5], and ICH Q6B

[section 2 and 4]).

159

2.8.5 Justification of Specification

Justification for the drug substance specification should be provided (ICH

Q3A(R1) [section 5], ICH Q5C [section 8], and ICH Q6B [section2 and section

4.1]).

2.9 Reference standards or materials

Information on the reference standards or reference materials used for testing of the

drug substance should be provided (ICH Q6A [section 2], and ICH Q6B [section2.2]).

2.10 Container Closure System

A description of the container closure system(s) should be provided, including the

identity of materials of construction of each primary packaging component, and their

specifications. The specifications should include description and identification (and

critical dimensions with drawings, where appropriate). Non-compendial methods (with

validation) should be included, where appropriate.

For non-functional secondary packaging components (e.g., those that do not provide

additional protection), only a brief description should be provided. For functional

secondary packaging components, additional information should be provided.

The suitability should be discussed with respect to, for example, choice of materials,

protection from moisture and light, compatibility of the materials of construction with

the drug substance, including sorption to container and leaching, and/or safety of

materials of construction (ICH Q5C [section 6.5]).

160

2.11 Stability

2.11.1 Stability Summary and Conclusions

The types of studies conducted, protocols used, and the results of the studies should be

summarized. The summary should include results, for example, from forced

degradation studies and stress conditions, as well as conclusions with respect to storage

conditions and retest date or shelf-life, as appropriate (ICH Q1A [section 2.1], ICH

Q1B [sections 1 and 2], and ICH Q5C [section 5]).

2.11.2 Post-approval stability protocol and stability commitment

The post-approval stability protocol and stability commitment should be provided (ICH

Q1A and ICH Q5C [sections 7 and 8]).

2.11.3 Stability Data

Results of the stability studies (e.g., forced degradation studies and stress condit ions)

should be presented in an appropriate format such as tabular, graphical, or narrative.

Information on the analytical procedures used to generate the data and validation of

these procedures should be included (ICH Q1A, ICH Q1B, ICH Q2A, ICH Q2B, and

ICH Q5C).

3. Drug product

3.1 Description and composition of the drug product

A description of the drug product and its composition should be provided. The information

provide should include, for example:

Description of the dosage form

161

Composition

List of all components of the dosage form

Amount per-unit basis (which should also include overages, if any)

The function of the components

A reference to their quality standards (e.g., compendial monographs or

manufacturer‟s specifications)

Description of accompanying reconstitution diluent(s)

Type of container and closure used for the dosage form and accompanying

reconstitution diluent, if applicable.

3.2 Pharmaceutical Development

The Pharmaceutical Development section should contain information on the

development studies conducted to establish that the dosage form, the formulation,

manufacturing process, container closure system, microbiological attributes and usage

instructions are appropriate for the purpose specified in the application.

The studies described here are distinguished from routine control tests conducted

according to specifications.

Additionally, this section should identify and describe the formulation and process

attributes (critical parameters) that can influence batch reproducibility, product

performance and drug product quality.

Supportive data and results from specific studies or published literature can be included

within or attached to the Pharmaceutical Development section. Additional supportive

data can be referenced to the relevant nonclinical or clinical sections of the application

(ICHQ6A and Q6B).

3.2.1 Components of the drug product

(A) Drug Substance: The compatibility of the drug substance with excipients

should be discussed. Additionally, key physicochemical characteristics (e.g.,

water content, solubility, particle size distribution, polymorphic or solid state

162

form) of the drug substance that can influence the performance of the drug

product should be discussed. For combination products, the compatibility of

drug substances with each other should be addressed.

(B) Excipients: The choice of excipients, their concentration, and their

characteristics that can influence the drug product performance should be

discussed relative to their respective functions.

3.2.2 Drug Product

(A) Formulation Development: A brief summary describing the

development of the drug product should be provided, taking into

consideration the proposed route of administration and usage. The

differences between clinical formulations and the formulation (i.e.

composition) should be described.

(B) Overages: Any overages in the formulation(s) should be justified.

(C) Physicochemical and Biological Properties: Parameters relevant to the

performance of the drug product, such as pH, ionic strength, dissolution,

redispersion, reconstitution, particle size distribution, aggregation,

polymorphism, rheological properties, biological activity or potency,

and/or immunological activity, should be addressed.

3.2.3 Manufacturing process development

The selection and optimisation of the manufacturing process, in particular its

critical aspects, should be explained.

Where relevant, the method of sterilisation should be explained and justified.

Differences between the manufacturing process(es) used to produce pivotal

clinical batches and the process should be described, that can influence the

performance of the product should be discussed.

163

3.2.4 Container closure system

The suitability of the container closure system used for the storage,

transportation (shipping) and use of the drug product should be discussed.

This discussion should consider, e.g., choice of materials, protection from

moisture and light, compatibility of the materials of construction with the

dosage form (including sorption to container and leaching) safety of materials of

construction, and performance (such as reproducibility of the dose delivery from

the device when presented as part of the drug product).

3.2.5 Microbiological attributes

Where appropriate, the microbiological attributes of the dosage form should be

discussed, including, for example, the rationale for not performing microbial

limits testing for non-sterile products and the selection and effectiveness of

preservative systems in products containing antimicrobial preservatives.

For sterile products, the integrity of the container closure system to prevent

microbial contamination should be addressed.

3.2.6 Compatibility

The compatibility of the drug product with reconstitution diluent(s) or dosage

devices (e.g., precipitation of drug substance in solution, sorption on injection

vessels, stability) should be addressed to provide appropriate and supportive

information for the labeling.

164

3.3 Manufacturing

3.3.1 Manufacturer(s)

The name, address, and responsibility of each manufacturer, including

contractors, and each proposed production site or facility involved in

manufacturing and testing should be provided.

3.3.2 Batch formula

A batch formula should be provided that includes a list of all components of the

dosage form to be used in the manufacturing process, their amounts on a per

batch basis, including overages, and a reference to their quality standards.

3.3.3 Description of manufacturing process and process controls

A flow diagram should be presented giving the steps of the process and

showing where materials enter the process.

The critical steps and points at which process controls, intermediate tests or

final product controls are conducted should be identified.

A narrative description of the manufacturing process, including packaging,

that represents the sequence of steps undertaken and the scale of production

should also be provided.

Novel processes or technologies that directly affect product quality should

be described with a greater level of detail.

Equipment should, at least, be identified by type and working capacity,

where relevant.

Steps in the process should have the appropriate process parameters

identified, such as time, temperature, or pH. Associated numeric values can

be presented as an expected range. Numeric ranges for critical steps should

be justified. In certain cases, environmental conditions should be stated.

165

Proposals for the reprocessing of materials should be justified. Any data to

support this justification should be either referenced or filed, as described in

ICH Q6B.

3.3.4 Controls of critical steps and intermediates

(A) Critical Steps: Tests and acceptance criteria should be provided (with

justification, including experimental data) performed at the critical steps

of the manufacturing process, to ensure that the process is controlled.

(B) Intermediates: Information on the quality and control of intermediates

isolated during the process should be provided as described in ICH Q2A,

ICH Q2B, ICH Q6A, and ICH Q6B.

3.3.5 Process validation and/or evaluation

Description, documentation, and results of the validation and/or evaluation

studies should be provided for critical steps or critical assays used in the

manufacturing process (e.g., validation of the sterilisation process or aseptic

processing or filling), as described in ICH Q6B.

3.4 Control of excipients

3.4.1 Specifications

The specifications for excipients should be provided as described in ICH Q6A

and ICH Q6B.

3.4.2 Analytical procedures (name, dosage form)

The analytical procedures used for testing the excipients should be provided,

where appropriate, as described in ICH Q2A and ICH Q2B.

166

3.4.3 Validation of analytical procedures

Analytical validation information, including experimental data, for the analytical

procedures used for testing the excipients should be provided, where

appropriate, as described in ICH Q3A, ICH Q2A, ICH Q2B, and ICH Q6B.

3.4.4 Justification of specifications

Justification for the proposed excipient specifications should be provided, where

appropriate, as described in ICH Q3C and ICH Q6B.

3.4.5 Excipients of human or animal origin

For excipients of human or animal origin, information should be provided

regarding adventitious agents (e.g., sources, specificationsm, description of the

testing performed, viral safety data), as described in ICH Q5A, ICH Q5D, and

ICH Q6B.

3.4.6 Novel excipient

For excipient(s) used for the first time in a drug product or by a new route of

administration, full details of manufacture, characterization, and controls, with

cross references to supporting safety data (nonclinical and/or clinical) should be

provided according to the drug substance format.

3.5 Control of drug product

3.5.1 Specification(s)

The specification(s) for the drug product should be provided, as described in

ICH Q3B, ICH Q6A, and ICH Q6B.

167

3.5.2 Analytical procedures

The analytical procedures used for testing the drug product should be provided,

as described in ICH Q2A and ICH Q6B.

3.5.3 Validation of analytical procedures

Analytical validation information, including experimental data, for the analytical

procedures used for testing the drug product, should be provided as described in

ICH Q2A, ICH Q2B, and ICH Q6B.

3.5.4 Batch analyses

A description of batches and results of batch analyses should be provided, as

described in ICH Q3A, ICH Q3C, ICH Q6A, and ICH Q6B.

3.5.5 Characterization of impurities

Information on the characterization of impurities should be provided, if not

previously provided under the section of “Impurities 2.7.2,” as described in ICH

Q3B, ICH Q5C, ICH Q6A, and ICH Q6B.

3.5.6 Justification of specification(s)

Justification for the proposed drug product specification(s) should be provided,

as described in ICH Q3B, ICH Q6A, and ICH Q6B.

3.6 Reference standards or materials

Information on the reference standards or reference materials used for testing of the

drug product should be provided, if not previously provided in 2.9 as described in ICH

Q6A, and ICH Q6B.

168

3.7 Packaging material

3.7.1 Container closure system

A description of the container closure systems should be provided, including

the identity of materials of construction of each primary packaging

component and its specification.

The specifications should include description and identification (and critical

dimensions, with drawings where appropriate). Non-compendial methods

(with validation) should be included where appropriate.

For non-functional secondary packaging components (e.g., those that neither

provide additional protection nor serve to deliver the product), only a brief

description should be provided. For functional secondary packaging

components, additional information should be provided.

3.7.2 Product package insert/product leaflet: it should include the

following information but not limited to:

Drug description,

o Indications

o Dosage administration (Paediatric patients, adult

patients, for all patients, and stability and storage)

o Dosage form

o Side effects

o Drug interactions

o Precautions

o Use in specific condition (Pregnancy, Nursing

Mothers, Geriatric Use etc)

o Over dosage

o Missed dose

o Contraindications

o Clinical pharmacology

Mechanism of action

169

Pharmacodynamics

Pharmacokinetics

o Instruction for use (pictorial stepwise presentation)

o Storage.

3.8 Stability

3.8.1 Stability summary and conclusion

The types of studies conducted, protocols used, and the results of the studies

should be summarized. The summary should include, for example, conclusions

with respect to storage conditions and shelf-life, and, if applicable, in-use

storage conditions and shelf-life (ICH Q1A, ICH Q1B, ICH Q3B, ICH Q5C,

and ICH Q6A).

3.8.2 Post-approval stability protocol and stability commitment

The post-approval stability protocol and stability commitment should be

provided (ICH Q1A and ICH Q5C).

3.8.3 Stability data

Results of the stability studies should be presented in an appropriate format (e.g.

tabular, graphical, narrative). Information on the analytical procedures used to

generate the data and validation of these procedures should be included.

Information on characterisation of impurities is located in 2.7.2 (ICH Q1A, ICH

Q1B, ICH Q2A, ICH Q2B and ICH Q5C).

170

3.9 Appendices

3.9.1 Changes reporting

Pre and post approval changes,

Pre and post or during clinical trial changes

o Detailed description of change

o At what stage of the manufacturing process change was introduced.

o What is foreseeable impact of such changes in the current process (it

should be address under risk based approach, ICH Q9)

3.9.2. Facilities and equipment

A diagram should be provided illustrating the manufacturing flow including

movement of raw materials, personnel, waste, and intermediate(s) in and out

of the manufacturing areas.

Information should be presented with respect to adjacent areas or rooms that

may be of concern for maintaining integrity of the product.

Information on all developmental or approved products manufactured or

manipulated in the same areas as the applicant's product should be included.

A summary description of product-contact equipment , and its use

(dedicated or multi-use) should be provided. Information on preparation,

cleaning, sterilization, and storage of specified equipment and materials

should be included, as appropriate.

Information should be included on procedures (e.g., cleaning and production

scheduling) and design features of the facility (e.g., area classifications) to

prevent contamination or cross-contamination of areas and equipment,

where operations for the preparation of cell banks and product

manufacturing are performed.

3.9.3 Adventitious agents’ safety evaluation

Information assessing the risk with respect to potential contamination with

adventitious agents should be provided in this section.

171

3.9.3.1 For non-viral adventitious agents

Detailed information should be provided on the avoidance

and control of non-viral adventitious agents (e.g.,

transmissible spongiform encephalopathy agents, bacteria,

mycoplasma, fungi).

This information can include, for example, certification

and/or testing of raw materials and excipients, and control of

the production process, as appropriate for the material,

process and agent (ICH Q5A, ICH Q5D, and ICH Q6B).

3.9.3.2 For viral adventitious agents

Detailed information from viral safety evaluation studies

should be provided in this section.

Viral evaluation studies should demonstrate that the materials

used in production are considered safe, and that the

approaches used to test, evaluate, and eliminate the potential

risks during manufacturing are suitable (ICH Q5A, ICH Q5D,

and ICH Q6B).

3.9.4 Materials of biological origin

Information essential to evaluate the virological safety of materials of animal

or human origin (e.g. biological fluids, tissue, organ, cell lines) should be

provided.

For cell lines, information on the selection, testing, and safety assessment for

potential viral contamination of the cells and viral qualification of cell banks

should also be provided.

172

3.9.5 Testing at appropriate stages of production

The selection of virological tests that are conducted during manufacturing (e.g.,

cell substrate, unprocessed bulk or post viral clearance testing) should be

justified. The type of test, sensitivity and specificity of the test, if applicable,

and frequency of testing should be included. Test results to confirm, at an

appropriate stage of manufacture, that the product is free from viral

contamination should be provided.

3.9.6 Viral testing of unprocessed bulk.

Results for viral testing of unprocessed bulk should be included (ICH Q5A and

ICH Q6B).

3.9.7 Viral clearance studies

In accordance with Q5A, the rationale and action plan for assessing viral

clearance and the results and evaluation of the viral clearance studies should be

provided. Data can include those that demonstrate the validity of the scaled-

down model compared to the commercial scale process; the adequacy of viral

inactivation or removal procedures for manufacturing equipment and materials;

and manufacturing steps that are capable of removing or inactivating viruses

(ICH Q5A, ICH Q5D, and ICH Q6B).

3.10 List of used excipients

3.10.1 Regional information

Any additional drug substance and/or drug product information specific to each

region should be provided in section R of the application. Applicants should

consult the appropriate regional guidelines and/or regulatory authorities for

additional guidance. Some examples are as follows:

173

Executed batch records, method validation package, comparability

protocols.

Process validation scheme for the drug product, where validation is still to

be completed, a summary of the studies intended to be conducted should be

provided.

3.10.2 Literature references

Key literature referenced should be provided, if applicable.

4. Pre-Clinical comparative study with RMP

Pre-clinical overview should present an integrated and critical assessment of

the pharmacologic, pharmacokinetic, and toxicologic evaluation.

For pharmacokinetic studies for biotechnology-derived pharmaceuticals,

single and multiple dose pharmacokinetics, toxicokinetics, and tissue

distribution studies in relevant species are useful; however, routine studies

that attempt to assess mass balance are not useful.

This section should include details of pre clinical tests performed on the

similar biological medicinal product and differences with relevant attributes

of the RMP.

Changes introduced during development which could affect comparability

should be highlighted and tests versus the RMP for quality, safety and

efficacy should be described.

The RMP used throughout the quality, safety and efficacy development

programme (as appropriate) should be defined.

This is not intended to indicate what studies are required. It merely indicates

an appropriate format for the preclinical data that have been acquired.

Detection of occurrence of glycosylation, contamination and changes to 3D

structure, which may affect potency/response and immunogenicity are of

primary concern.

174

Glycosylation of recombinant proteins can influence their degradation, their

exposure of antigenic sites and their solubility, as well as their

immunogenicity.

Changes to three-dimensional structure, protein aggregation, oxidation and

deamidation, can have major affect on its degradation and should be

detected

4.1 Preclinical testing should mention

4.1.1 Selected relevant animal species (number/gender)

Due to the species specificity of many biotechnology-derived pharmaceuticals,

it is important to select relevant animal species for toxicity testing. Both

genders should generally be used or justification given for specific omissions.

A variety of techniques (e.g., immunochemical or functional tests) can be used

to identify a relevant species. Knowledge of receptor/epitope distribution can

provide greater understanding of potential in vivo toxicity.

4.1.2 Delivery, dose and route of administration

The route and frequency of administration should be as close as possible to that

proposed for clinical use. Consideration should be given to pharmacokinetics

and bioavailability of the product in the species being used, and the volume

which can be safely and humanely administered to the test animals. Dosage

levels should be selected to provide information on a dose-response

relationship.

4.2. Pharmacology/pharmacodynamics

4.2.1 In vitro studies used to assess any alterations in reactivity between

the similar biological medicinal and the medicinal product (data from a

number of comparative bioassays) should be provided

4.2.2 In vitro cell lines derived from mammalian cells, used to predict

specific aspects of in vivo activity and to assess quantitatively the

relative sensitivity of various species to the biosimilar, should be

documented.

175

4.2.3 Studies designed to determine receptor occupancy, receptor affinity,

and/or pharmacological effects, if appropriate, should be explained.

4.2.4 In vivo studies that assess pharmacological activity, including

defining mechanism(s) of action and support rationale of proposed use,

should be explained.

4.3 Pharmacokinetics

4.3.1 The relative pharmacokinetic properties of the biosimilar product and the

RMP should be determined in single submaximal dose crossover studies

using subcutaneous and intravenous administration.

4.3.2 The primary pharmacokinetic parameter AUC and the secondary

pharmacokinetic parameters Cmax and T1/2 or CL/F should be

determined. Equivalence margins have to be defined and appropriately

justified.

4.3.3 The behaviour of the biosimilar in the biologic matrix and the possible

influence of binding proteins effect on pharmacodynamic effects, should

be mentioned.

4.4 Toxicological studies

Conventional approaches to toxicity testing of pharmaceuticals are not

appropriate for biopharmaceuticals due to the unique and diverse structural

and biological properties of the latter that may include species specificity,

immunogenicity, and unpredicted pleiotropic activities.

The onset, severity, and duration of the toxic effects, their dose-dependency

and degree of reversibility (or irreversibility), and species- or gender-related

differences should be evaluated

Pharmacodynamics, toxic signs, causes of death, pathologic findings should

be investigated.

176

4.4.1 Single dose toxicity

Significant side effects that may arise should be evaluated.

The onset, severity, and duration of the toxic effects, their dose-

dependency and degree of reversibility (or irreversibility), and

species- or gender-related differences should be evaluated and

important features discussed.

The biosimilar should be tested in pharmacologically relevant

species.

4.4.2 Repeated dose toxicity studies

Data from at least one repeat-dose toxicity study in a relevant species

should be provided.

Special emphasis should be laid on determining immune responses.

For biopharmaceuticals that induce prolonged

pharmacological/toxicological effects, recovery group animals

should be monitored until reversibility is demonstrated.

The duration of repeated dose studies should be based on the

intended duration of clinical exposure and disease indication.

4.4.3 Local tolerance

Data on local tolerance in at least one species should be evaluated.

4.5 Immunogenicity profile

Due to their immunogenicity, antibody determinations should be

measured if more than a single dose is administered in the toxicology

studies, to help gauge the relevance of the toxicity data collected.

Antibody responses should be characterised (e.g., titer, number of

responding animals, neutralising or non-neutralising), and their

177

appearance should be correlated with any pharmacological,

pharmacokinetic and/or toxicological changes.

Possible pathological changes related to immune complex formation

and deposition should be evaluated.

Inflammatory reactions at the injection site should be recorded since

it may be indicative of a stimulatory response (may stimulate or

suppress the immune system and affect not humoral and/or cell-

mediated immunity.

Routine tiered testing approaches or standard testing batteries are not

recommended (including guinea pig anaphylaxis tests (generally

positive for protein products).

5. Clinical comparative study with RMP

5.1 Protocol

It is recommended to generate the required clinical data for the comparability study

with the test product as produced with the final manufacturing process and, therefore,

representing the quality profile of the batches to become commercialized. Equivalent

therapeutic efficacy should be demonstrated. Frequently, clinical studies should be

randomized and double blind to avoid bias.

Possible differences in efficacy should normally be investigated in studies with the

highest probability of showing a difference. Any deviation from this recommendation

should be justified and supported by adequate additional data.

178

5.2 Recruitment details

5.2.1 Informed consent document(s)

A copy of the “Informed Consent” documents(s) to be used in conjunction with

the clinical trial(s), including a statement regarding the risks and anticipated

benefits to the clinical trial subjects as a result of their participation in the

clinical trial(s); Informed Consent document(s) to be used in conjunction with

the clinical trial(s) should be prepared in accordance with ICH-E6.

5.2.1 Clinical trial site information

A complete clinical trial site information form for each clinical trial site should

be furnished.

5.3 Eligibility criteria

A patient population should be chosen where differences are best distinguishable, i.e.

the most sensitive model for efficacy. Concerns regarding human gender, women with

child-bearing age potential, pregnant women, children, and individuals with chronic

diseases must be taken into consideration

5.4 Clinical studies reports

5.4.1 Reports on biopharmaceutical studies

Bioavailability studies evaluate the rate and extent of release of the active

substance from the medicinal product. Comparative bioavailability or

bioequivalence studies may use PK, PD, clinical, or in vitro dissolution

endpoints, and may be either single dose or multiple dose. Bioavailability

studies in this section should include:

179

Studies comparing the release and systemic availability of a drug substance

from a solid oral dosage form to the systemic availability of the drug

substance given intravenously or as an oral liquid dosage form.

Dosage form proportionality studies.

Food-effect studies.

5.4.2 Reports of studies pertinent to pharmacokinetics using human

biomaterials

Of particular importance is the use of human biomaterials such as hepatocytes

and/or hepatic microsomes to study metabolic pathways and to assess drug-drug

interactions with these pathways. Reports should include:

Plasma Protein Binding Study Reports

Reports of Hepatic Metabolism and Drug Interaction Studies

Reports of Studies Using Other Human Biomaterials

5.4.3 Reports on pharmacokinetics (PK)

These reports should provide a description of the body‟s handling of a drug over

time, focusing on maximum plasma concentrations (peak exposure), area-under-

curve (total exposure), clearance, and accumulation of the parent drug and its

metabolite(s), in particular those that have pharmacological activity.

The PK studies are generally designed to (1) measure plasma drug and

metabolite concentrations over time, (2) measure drug and metabolite

concentrations in urine or faeces when useful or necessary, and/or (3) measure

drug and metabolite binding to protein or red blood cells.

Healthy subject PK and initial tolerability study reports

Patient PK and initial tolerability study reports

Intrinsic factor PK study reports

180

Extrinsic factor PK study reports

Population PK study reports

5.4.4 Reports on pharmacodynamic (PD)

This section should include reports of [1] studies of pharmacologic properties

known or thought to be related to the desired clinical effects (biomarkers), [2]

short-term studies of the main clinical effect, and [3] PD studies of other

properties not related to the desired clinical effect.

5.4.5 Reports on efficacy and safety

This section should include reports of all clinical studies of efficacy and/or

safety carried out with the drug, conducted by the sponsor, or otherwise

available, including all completed and all ongoing studies of the drug in

proposed and non-proposed indications. The study reports should provide the

level of detail appropriate to the study and its role in the application. ICH E3

describes the contents of a full report for a study contributing evidence pertinent

to both safety and efficacy.

5.5 Statistics (justification of statistical method used)

5.6 Reports of post-marketing experience

Data from pre-authorization clinical studies are normally insufficient to identify all

potential differences. Therefore, clinical safety of biosimilars must be monitored

closely on an ongoing basis during the post-approval phase, including continued

assessment of benefits and risks.

The applicant should give a risk specification in the application DMF for the medicinal

product under review. This includes a description of possible safety issues related to

tolerability of the medicinal product that may result from a manufacturing process

181

different from that of the innovator. In the DMF, the applicant should present a risk

management program or pharmacovigilance plan in accordance with current GCC

procedures and guidelines. This should take into account risks identified during product

development and potential risks.

Pharmacovigilance systems and procedures to achieve this monitoring should be in

place when a marketing authorization is granted. Any specific safety monitoring

imposed to the RMP or product class should be taken into consideration in the risk

management plan.

For further information on this issue, ICH topic Q9 can be used. For reporting, the

GCC Guidelines on Pharmacovigilance should be referred to.

5.6. Testing of immunogenicity

The applicant should present a rationale for the proposed antibody-testing

strategy. Testing for immunogenicity should be performed by state-of-the-art methods,

using assays with appropriate specificity and sensitivity. The screening assays should be

validated and sensitive enough to detect low titre and low affinity antibodies. An assay

for neutralizing antibodies should be available for further characterization of antibodies

detected by the screening assays. Standard methods and international standards should

be used whenever possible. The possible interference of the circulating antigen with the

antibody assays should be taken into account. The periodicity and timing of sampling

for testing of antibodies should be justified.

In view of the unpredictability of the onset and incidence of immunogenicity,

long term results of monitoring of antibodies at predetermined intervals will be

required. In case of chronic administration, one-year follow up data will be required

pre-licensing. The applicant should consider the possibility of antibodies against

process-related impurities

If a different immune response to the biosimilar product is observed as compared to the

innovator product, further analyses to characterize the antibodies and their

182

implications to clinical safety, efficacy and pharmacokinetic parameters are required.

Special consideration should be given to those products where there is a chance that the

immune response could seriously affect the endogenous protein and its unique

biological function. Antibody testing should be considered as part of all clinical

trials protocols. The applicant should consider the role of immunogenicity in certain

events, such as hypersensitivity, infusion reactions, autoimmunity and loss of efficacy.

5.7 Literature references

Copies of referenced documents, including important published articles, official

meeting minutes, or other regulatory guidance or advice should be provided. Only one

copy of each reference should be provided. Copies of references that are not included

here should be immediately available on request.

183

References

EMEA

http://www.emea.europa.eu/

http://www.emea.europa.eu/htms/human/human guidelines/

Quality of Biotechnological Products: Stability Testing of Biotechnological/Biological

Products. July 1996

http://www.emea.europa.eu/pdfs/human/bwp/3ab5aen.pdf

Guideline on comparability of medicinal products containing biotechnology-derived

proteins as active substance: Quality issues (EMEA/CPMP/BWP/3207/00/Rev1*)

http://www.emea.europa.eu/pdfs/human/bwp/320700en.pdf

Note for Guidance on the Investigation of Bioavailability and Bioequivalence

(CHMP/EWP/QWP/1401/98) - 2001

http://www.emea.europa.eu/pdfs/human/ewp/140198en.pdf

Guideline on similar biological medicinal products (CHMP/437/04)

http://www.emea.europa.eu/pdfs/human/biosimilar/043704en.pdf

Guidelines on similar biological medicinal products containing biotechnology-derived

proteins as active substances: Quality issues (EMEA/CHMP/BWP/49348/2005)

http://www.emea.europa.eu/pdfs/human/biosimilar/4934805en.pdf

Guidelines on similar biological medicinal products containing biotechnology-derived

proteins as active substances: Nonclinical and clinical issues

(EMEA/CHMP/BMWP/42832/2005)

http://www.emea.europa.eu/pdfs/human/biosimilar/4283205en.pdf

184

Guidelines on similar biological medicinal products containing biotechnology-derived

proteins as active substances: Nonclinical and clinical issues

(EMEA/CHMP/BMWP/42832/2005). February 2006

http://www.emea.europa.eu/pdfs/human/biosimilar/4283205en.pdf

Concept paper on guideline on comparability of biotechnology derived medicinal

products after a change in the manufacturing process: Non-clinical and clinical issues

(EMEA/CHMP/BMWP/9437/2006/corr)

http://www.emea.europa.eu/pdfs/human/biosimilar/943706en.pdf

Guideline on Comparability of Biotechnology-Derived Medicinal Products after a

Change in the Manufacturing Process: Non-Clinical and Clinical Issues

(EMEA/CHMP/BMWP/101695/2006) 19 July 2007

http://www.emea.europa.eu/pdfs/human/biosimilar/10169506enfin.pdf

Guideline on the Clinical Investigation of the Pharmacokinetics of Therapeutic Proteins

(CHMP/EWP/89249/2004). January 2007

http://www.emea.europa.eu/pdfs/human/ewp/8924904enfin.pdf

Guideline on immunogenecity assessment of biotechnology-derived therapeutic

proteins (EMEA/CHMP/BMWP/14327/2006) - 2007

http://www.emea.europa.eu/pdfs/human/biosimilar/1432706en.pdf

Guideline on Similar Medicinal Products Containing

Recombinant Interferon Alpha EMEA/CHMP/BMWP/102046/2006

www.emea.europa.eu/pdfs/human/biosimilar/10204606en.pdf

European Public Assessment Report (EPAR)

Viraferonpeg: EPAR summary for the public EMEA/H/C/329

http://www.emea.europa.eu/humandocs/PDFs/EPAR/Viraferonpeg/137200en1.pdf

European Public Assessment Report (EPAR). Viraferonpeg: Scientific Discussion

http://www.emea.europa.eu/humandocs/PDFs/EPAR/Viraferonpeg/137200en6.pdf

185

European Public Assessment Report (EPAR)

Avonex: EPAR summary for the public EMEA/H/C/102

http://www.emea.europa.eu/humandocs/PDFs/EPAR/avonex/106396en1.pdf

European Public Assessment Report (EPAR)

Avonex: Scientific Discussion

http://www.emea.europa.eu/humandocs/PDFs/EPAR/avonex/106396en6.pdf

PUBLIC SUMMARY OF POSITIVE OPINION FOR ORPHAN DESIGNATION

Interferon gamma for the treatment of idiopathic pulmonary fibrosis

(EMEA/COMP/468704/2007)

http://www.emea.europa.eu/pdfs/human/comp/opinion/46870407en.pdf

PUBLIC SUMMARY OF POSITIVE OPINION FOR ORPHAN DESIGNATION

Interferon beta for the treatment of acute lung injury (EMEA/COMP/473691/2007)

http://www.emea.europa.eu/pdfs/human/comp/opinion/47369107en.pdf

Guidance on Similar Medicinal Products Containing Somatropin,

EMEA/CHMP/BMWP/94528/2005.

http://www.emea.europa.eu/pdfs/human/biosimilar/9452805en.pdf.

Guideline on risk management systems for medicinal products for human use

(EMEA/CHMP 96286/2005).

http://www.emea.europa.eu/pdfs/human/euleg/9626805en.pdf.

Note for Guidance on Good Clinical Safety Data Management: Definitions and

Standards for Expedited Reporting (CPMP/ICH/377/95).

www.emea.europa.eu/pdfs/human/ich/037795en.pdf

ICH Note for Guidance on Planning Pharmacovigilance Activities (CPMP/ICH/5716/03

- Final approval by CHMP on PHV).

www.emea.europa.eu/pdfs/human/ich/571603en.pdf

186

Annex To Guideline On Similar Biological Medicinal Products Containing

Biotechnology-Derived Proteins As Active Substance: Non-Clinical And Clinical

Issues. Guidance On Similar Medicinal Products Containing Recombinant Granulocyte-

Colony Stimulating Factor (EMEA/CHMP/BMWP/31329/2005) London, 22 February

2006.

http://www.tga.gov.au/DOCS/pdf/euguide/bmwp/3132905en.pdf

European Public Assessment Report (EPAR) of Neupopeg.

http://www.emea.europa.eu/humandocs/PDFs/EPAR/neupopeg/296202enl.pdf

Scientific Discussion for the Approval of Neupopeg

http://www.emea.europa.eu/humandocs/PDFs/EPAR/neupopeg/296202en6.pdf

Annex to Guideline on Similar Biological Medicinal Products Containing

Biotechnology-Derived Proteins as Active Substance: Non-Clinical and Clinical Issues

Guidance on Similar Medicinal Products Containing Recombinant Erythropoietins. 22

March 2006.

http://www.emea.europa.eu/pdfs/human/biosimilar/9452605en.pdf

Annex To Guideline On Similar Biological Medicinal Products Containing

Biotechnology-Derived Proteins As Active Substance: Non-Clinical And Clinical

Issues. Guidance On Similar Medicinal Products Containing Recombinant Human

Soluble Insulin (EMEA/CHMP/BMWP/32775/2005) London, 22 February 2006

http://www.emea.europa.eu/pdfs/human/biosimilar/3277505en.pdf

ICH Topic S 3 A. Toxicokinetics: A Guidance for Assessing Systemic Exposure in

Toxicology Studies (CPMP/ICH/384/95) June 1995

http://www.emea.europa.eu/pdfs/human/ich/038495en.pdf

Note for guidance on non-clinical local tolerance testing of medicinal products

(CPMP/SWP/2145/00). March 2001

http://www.emea.europa.eu/pdfs/human/swp/214500en.pdf

187

Directive 2001/83/Ec Of The European Parliament And Of The Council Of 6

November 2001 On The Community Code Relating To Medicinal Products For Human

Use, As Amended London 28/11/2004

http://www.emea.europa.eu/pdfs/human/pmf/2001-83-EC.pdf

Guidance On Similar Medicinal Products Containing Recombinant Human Soluble

Insulin (EMEA/CHMP/BMWP/32775/2005) London, 22 February 2006

http://www.emea.europa.eu/pdfs/human/biosimilar/3277505en.pdf

Note for guidance on non-clinical local tolerance testing of medicinal products

(CPMP/SWP/2145/00). March 2001

http://www.emea.europa.eu/pdfs/human/swp/214500en.pdf

Directive 2001/83/Ec Of The European Parliament And Of The Council Of 6

November 2001 On The Community Code Relating To Medicinal Products For Human

Use, As Amended London 28/11/2004

http://www.emea.europa.eu/pdfs/human/pmf/2001-83-EC.pdf

ICH

http://www.ich.org/cache/compo/363-272-1.html

ICH Topic E1 - The Extent of Population Exposure to Assess Clinical Safety for Drugs

Intended for Long-term Treatment of Non-Life-Threatening Conditions

(CPMP/ICH/375/95)

http://www.emea.europa.eu/pdfs/human/ich/037595en.pdf

ICH Topic E6 - Good Clinical Practice: Consolidated Guideline 10 June 1996

http://www.ich.org/LOB/media/MEDIA482.pdf

ICH Topic E8 - General Considerations for Clinical Trials - 17 July 1997

http://www.ich.org/LOB/media/MEDIA484.pdf

188

ICH Topic E 9 - Statistical Principles for Clinical Trials (CPMP/ICH/363/96)

September 1998

http://www.emea.europa.eu/pdfs/human/ich/036396en.pdf

ICH Topic E10 - Choice of control group and related issues in clinical trials

(CPMP/ICH/364/96) - London 2001

http://www.emea.europa.eu/pdfs/human/ich/036496en.pdf

ICH topic Q5A(R1) - Viral safety evaluation of biotechnology products derived from

cell lines of human or animal origin (CPMP/ICH/295/95)

http://www.ich.org/LOB/media/MEDIA425.pdf

ICH topic Q5B - Quality of biotechnological products: Analysis of the expression

construct in cells used for production of R-DNA derived protein products

(CPMP/ICH/139/95)

http://www.ich.org/LOB/media/MEDIA426.pdf

ICH Topic Q5C - Quality of Biotechnological Products: Stability Testing of

Biotechnological/Biological Products (CPMP/ICH/138/95)

http://www.emea.europa.eu/pdfs/human/ich/013895en.pdf

ICH topic Q5D - Derivation and characterisation of cell substrates used for production

of biotechnological/biological products (CPMP/ICH/294/95)

http://www.ich.org/LOB/media/MEDIA429.pdf

ICH Topic Q5E - Comparability of Biotechnological/Biological Products Subject to

Changes In Their Manufacturing Process (CPMP/ICH/5721/03) - 18 November 2004

http://www.ich.org/LOB/media/MEDIA1196.pdf

ICH Topic Q 6 B - Specifications: Test Procedures and Acceptance Criteria for

Biotechnological/Biological Products (CPMP/ICH/365/96)

www.emea.europa.eu/pdfs/human/ich/036596en.pdf

189

ICH Topic Q 8 - Note For Guidance on Pharmaceutical Development

(EMEA/CHMP/167068/2004)

http://www.emea.europa.eu/pdfs/human/ich/16706804en.pdf

ICH Topic Q9 – Guidance for the Industry

Quality risk management. June 2006

http://www.fda.gov/CBER/gdlns/ichq9risk.pdf

ICH Topic S 6 - Preclinical Safety Evaluation of Biotechnology-Derived

Pharmaceuticals. (CPMP/ICH/302/95)

http://ww.emea.europa.eu/pdfs/human/ich/030295en.pdf

The Common Technical Document For The Registration Of Pharmaceuticals For

Human Use, Efficacy M4e(R1). Clinical Overview And Clinical Summary Of Module

2 Module 5: Clinical Study Reports.

http://www.ich.org/LOB/media/MEDIA561.pdf

GCC

http://www.sfda.gov.sa

Clinical Trials Requirements Guidelines. May 2005

http://www.sfda.gov.sa/NR/rdonlyres/858DFB23-47E8-4838-854A-

EF3076A05481/0/ClinicalTrialRequirementGuidelines.pdf

The GCC Guidelines for Stability Testing of Drug Substances and Pharmaceutical Products.

Edition two. 2007.

http://www.sfda.gov.sa/NR/rdonlyres/E4FCA44E-84B4-4DA6-9B28-

A42CCEA5FE8D/0/GCC_Stability_Guidelines_Dec_2007Final.pdf

190

OTHER GUIDELINES

Annex To Guideline On Similar Biological Medicinal Products Containing

Biotechnology-Derived Proteins As Active Substance: Non-Clinical And Clinical

Issues. Guidance On Similar Medicinal Products Containing Recombinant Granulocyte-

Colony Stimulating Factor (EMEA/CHMP/BMWP/31329/2005) London, 22 February

2006

http://www.tga.gov.au/DOCS/pdf/euguide/bmwp/3132905en.pdf

“Notice to applicants and regulatory guidelines medicinal products for human use, The

rules governing medicinal products in the European Union, Chapter 1 of Vol. 2B: Module

4. Presentation and format of the dossier Common Technical Document (CTD).

http://ec.europa.eu/enterprise/pharmaceuticals/eudralex/homev2.htm

“Note for guidance on preclinical safety evaluation of biotechnology-derived pharmaceuticals”

(CPMP/ICH/302/95).

“Toxicokinetics: the assessment of systemic exposure in toxicity studies” (CPMP/ICH/384/95)

(ICH S3A).

“Pharmacokinetics: Guidance for repeated dose tissue distribution studies”

(CPMP/ICH/385/95) (ICH S3B).

“Guideline on similar biological medicinal products containing biotechnology-derived proteins

as active substance: non-clinical and clinical issues” (EMEA/CHMP/42832/05).

"Note for guidance on repeated dose toxicity" (CPMP/SWP/1042/99).

“Guideline on immunogenicity assessment of biotechnology derived therapeutic proteins”,

(CHMP/BMWP/14327/2006).

"Note for guidance on non-clinical local tolerance testing of medicinal products"

(CPMP/SWP/2145/00).

191

Selected Literature

Mahmood I (2009) Methods to determine pharmacokinetic profiles of therapeutic

proteins. Drug Discovery Today. In Press.

Horikawa H, Tsubouchi M, Kawakami K (2009) Industry views of biosimilar

development in Japan. Health Policy 91: 189-194.

Schellekens H (2009) Assessing the bioequivalence of biosimilars: The Retacrit®

case.

Drug Discovery Today 14: 495-499.

Wafelman AR (2008) Symposium Report – Development of safe protein therapeutics:

Pre-clinical, clinical and regulatory issues. European Journal of Pharmaceutical

Sciences 34: 223-225.

Tsuji K, Tsutani K (2008) Approval of new biopharmaceuticals 1999-2006:

Comparison of the US, EU and Japan situations. European Journal of Pharmaceutics

and Biopharmaceutics 68: 496-502.

Schellekens H (2008) Immunogenicity of therapeutic proteins and the Fabry antibody

standardization initiative. Clinical Therapeutics 30: S50-S51.

Nakazawa T, Kurokawa M, Kimura K, et al. (2008) Safety assessment of

biopharmaceuticals: Japanese perspective on ICH S6 guideline maintenance. Journal of

Toxicological Sciences 33: 277-282.

Mellstedt H, Nierderwieser D, Ludwig H (2008) The challenge of biosimilars. Annals

of Oncology 19: 411-419.

Hwang I, Park S (2008) Computational design of protein therapeutics. Drug Discovery

Today.

192

Bohlega S, Al-Shammari S, Al Sharoqi I, et al. (2008) Biosimilars: Opinion of an

expert panel in the Middle East. Current Medical Research and Opinion 24: 2897-2903.

Roger SD, Goldsmith D (2008) Biosimilars: It‟s not as simple as cost alone. Journal of

Clinical Pharmacy and Therapeutics 33: 459-464.

Wurm FM (2007) Manufacturing of biopharmaceuticals and implications for biosimilars.

Kidney and Blood Pressure Research 30: 6-8.

Roger S, Mikhail A (2007) Biosimilars: Opportunity or cause for concern? Journal of

Pharmacy and Pharmaceutical Sciences 10: 405-410.

Wright E (2007) Generic and biosimilar medicinal products in the European Union. Chemistry

Today 25: 4-6.

Nowicki M (2007) Basic facts about biosimilars. Kidney and Blood Pressure Research 30: 267-

272.

Walle IV, Gansemans Y, Parren PWHI, et al. (2007) Immunogenicity screening in protein drug

development. Current Opinion in Biology and Therapeutics 7: 405-418.

Trouvin J-H (2007) Introductory notes to the three-part series of papers by B Sharma on:

Immunogenicity of therapeutic proteins: How to assess and the role of pharmaceutical quality.

Biotechnology Advances 25: 307-309.

Barbosa MDFS, Celis E (2007) Immunogenicity of protein therapeutics and the

interplay between tolerance and antibody responses. Drug Discovery Today 12: 674-

681.

Kuhlmann M, Covic A (2006) The protein science of biosimilars. Nephrology Dialysis and

Transplantation 21 [Suppl 5]: v4–v8.

193

Locatelli F, Roger S (2006) Comparative testing and pharmacovigilance of biosimilars.

Nephrology Dialysis and Transplantation 21 [Suppl 5]: v13–v16.

Narhi M, Nordstrom K (2005) Manufacturing, regulatory and commercial challenges of

biopharmaceutical production: A Finnish perspective. European Journal of

Pharmaceutics and Biopharmaceutics 59: 397-405.

Schellekens H (2005) Follow-on biologics: challenges of the „next generation.‟ Nephrology

Dialysis and Transplantation 20 [Suppl 4]: v31–v36.