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.
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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.
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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
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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.
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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.
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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
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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:
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(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.
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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.
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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.
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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.
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CHAPTER 5.0
Other Important Issues
Labeling
Extrapolation Interchangeability and substitution
Stability studies
Storage conditions
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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
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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
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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.
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(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.
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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.
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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.
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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.
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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
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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.
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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,”
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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
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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
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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.
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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.
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CHAPTER 6.0
Insulin
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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
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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.
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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
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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
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(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.
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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.
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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.
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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
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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.
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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.
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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).
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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.
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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
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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.
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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.
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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.
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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.
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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
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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.
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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.
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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.
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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
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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.
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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
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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.
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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.
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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.
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Chapter 10.0
Human Growth Hormone
(Somatropin)
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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
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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
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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
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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
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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.
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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.
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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.
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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
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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.
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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.
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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
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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.
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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
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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
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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
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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.
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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
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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.
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(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).
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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
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(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:
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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.
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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.
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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.
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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)
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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]).
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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]).
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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
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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
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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.
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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.
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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.
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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.
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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.
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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.
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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
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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).
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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.
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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.
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