Comparability Study (for Biosimilar) Wisit Tangkeangsirisin, PhD Faculty of Pharmacy, Silpakorn University

Content: Biosimilar Definition and Concept Regulations Quality Assessment of Biosimilar Example and Cases Assignment

Monoclonal Antibodies Biologics Biopharmaceuticals Recombinant Proteins Monoclonal Antibodies Etc. Others Gene therapy Cell therapies Tissue therapy Allergens Blood & Blood Products Vaccines & Toxoids

Monoclonal Antibodies Therapeutic Proteins Biopharmaceuticals

Required only for Biosimilarity

Major Biopharmaceuticals Recombinant Proteins Insulin, Epoetins, Interferons, Somatropin , Filgrastim Monoclonal Antibodies Trastuzumab, Rituximab, Infliximab Immunoglobulin Fusion Proteins Etanercept

Biopharmaceuticals First generation Native Proteins, Unengineered murin antibodies, simple replacement proteins Frequent Injection Second generation (biobetter) Engineered, modified, alteration of amino acid sequence, alteration of glycosylation, PEGylation Suitable PK

The complexity of biologics relates to structure and function Monoclonal antibodies belong to the most complex classes of biologics CDRs (complementarity determining regions) Selected for optimum target mediated effects (light chain and heavy chain variable binding domains) Polypeptide subunits Carbohydrate For comparison – the size and complexity of aspirin Fc region: Different isotypes with different effector functions Contains 2 glycosylation sites (1 in each of the 2 polypeptide chains) In total more than 20.000 atoms All parts contribute to the efficacy and safety in a cooperative way Slide notes taken from original slides Monoclonal antibodies belong to the most complex classes of biotherapeutics 20 000 – 25 000 atoms, MW ca. 150 000 (IgG) 4 subunits, 2 light and 2 heavy chains Glycosylated: 2 Fc glycosylation sites plus optional Fab glycosylation Multifunctionality Antigen (target) binding through unique CDRs in Fab part multiple interactions of Fc part influencing effector functions and pharmacokinetics Immunoglobulin G1 (IgG1) molecule: Padlan EA. Mol Immunol 1994;31(3):169-217.

Monoclonal Antibodies Structurally much more complex than other proteins Complexity More complex than currently developed biosimilars Biological Activity glycosylation patterns are critical-smalll differences correlate to changes in biological activity Predictability multi-functionlity (binding and immuneffector functions) coupled with an often limited understanding of structure-function relationship will limit predictability of in vitro data Extrapolation complexity and diversity of the mechanisms of action will be of particular challenge for indications and line extensions

Quality Attributes to Consider - Mab Functional characteristics Physico-chemical characteristics Fab Fc Antigen binding Effector functions complement interaction Fc recepter interaction N-terminal heterogeneity pyroglutamate formation Other modifications AA modifications deamidation, oxidation, glycation, isomerization Fragmentation Cleavage in hinge region, Asp-Pro Oligosaccharides Fucosylation, sialyation, galactosylation… Disulfide bonds Free thiols, disulfide shuffling, thioether C-terminal heterogeneity Lysine processing, proline amidation

Biologics are hard to copy

Significance and Needs Why Biosimilar??

Why Biosimilar? Ensuring safety and effectiveness Accessibility of high cost medicine Safety Effectiveness Cost of Medicine (Biologics)

Definition Global Approval Pathways Key Issues biosimilars

NDA vs. ANDA Review Process New Drug NDA Requirements Generic Drug ANDA Requirements Chemistry Manufacturing Controls Labeling Testing Animal Studies Clinical Studies Bioavailability Chemistry Manufacturing Controls Labeling Testing Bioequivalence

Generics vs. Biosimilars Patent Expired Active Pharmaceutical Ingredients Identical Never identical (Highly Similar) Proof Needed Bioequivalence Comparability

What Biosimilar are…. And are not? They are similar …… They are not the same They are different from innovator’s product. They are not generic biologics

Which biologics are in the scope of Biosimilar? Vaccines Blood Products Recombinant Proteins Monoclonal Antibodies Transgenic cells/ Cell therapy EMA guidelines – can be adapted to all biologics including blood products, vaccines WHO guidelines and others – adapted only well established and well characterized biotherapeutic products

BioMarket Trends Jun 15, 2012 (Vol. 32, No BioMarket Trends Jun 15, 2012 (Vol. 32, No. 12) Biosimilar Market Posts Steady Gains Actual Performance Varies between Regulated and Semi- and Unregulated Environments.

Biosimilarity requirements Can we use the same approval pathway of generic for biologic?

Same Approval Pathway as Generics for Biologics? For Blood Products and Vaccines No, the approval pathway treats them as New biologics. For Recombinant Proteins Yes and No Yes  Generic Approval No  New biologics Approval Somewhat between  Biosimilar

Biosimilarity is built on five indispensible pillars. Modified from lecture of Dr. Thomas Schreitmueller

Approval of Biosimilar vs. Originator Biologics Pharmacovigilance Clinical (PI,II,III for each indication) Clinical (with extrapolation) Non-clinical Non-Clinical Comparability exercise in Quality Chemistry, Manufacturing and Control Chemistry, Manufacturing and Control Originator Biologics Biosimilars

Establishment of biosimilar guidelines increased, driven by WHO But: Variability in adoption by National Regulatory Authorities 2010 2015 Biosimilar pathways in place Biosimilar pathways under development

Global regulatory environment In general, regulators require: Comparative analytical program Clinical program Post-approval commitment to Pharmacovigilance High degree of analytical similarity High degree of pre-clinical and clinical similarity Regulators may permit extrapolation when scientifically justified.

Key Issues in Biosimilar Dossier Quality Comparisons The most critical and extensive step Cell culture, impurities, glycosylations …….. Non-clinical Comparisons In vitro receptor binding & cell based assay are fundamental Clinical Comparative Studies Most sensitive population and endpoints 6-12 months safety data Extrapolation of indications

Current Thailand Biosimilar Guidelines คู่มือ และหลักเกณฑ์การขึ้นทะเบียนตำรับยาชีววัตถุคล้ายคลึง 2556 Defines philosophy and principles Biotechnology-derived proteins General guidelines Quality Non- Clinical EPO Review Unorthodox Pathway ???? 2557 2558 2559 Epoetin IFN-α Somatropin GCSF mAbs IFN-β FSH Annex guidelines- specific data requirements Non- Clinical Non- Clinical Non- Clinical Non- Clinical Non- Clinical Non- Clinical Non- Clinical Immunogenicity Guideline for biosimilars

ประเภทของยาชีววัตถุในประเทศไทย Previous New Regulations ยาชีววัตถุใหม่ ยาชีววัตถุที่ไม่ใช่ยาใหม่ ยาชีววัตถุใหม่ ยาชีววัตถุคล้ายคลึง

Extrapolation of indications key to the biosimilar concept needs to be justified in all cases

Extrapolation of Indications The Key Consequence of Biosimilar is the EXTRAPOLATION of INDICATIONS Justification of the extrapolated indication is on a case-by-case basis. Decision based upon mechanism of action, type of receptors, Affinity etc.

Interchangeability In EU, the biosimilar approval do not automatically allow interchangeability Interchangeability/switching remains a national decision. Later, after 10 yrs experience of biosimilar in the market, several countries change regulations.

Production of Biosimilar Limitations Limit access in manufacturing process information Need to Proof Similarity in Certain Items Product Monograph Specific Guidelines Scientific Evidences

Dossier requirements for a biosimilar vs. an originator Dossier requirements for a biosimilar compared with an originator (stand-alone) dossier. For quality, a more extensive dossier is required, which includes the comparability exercise to the originator, whereas the nonclinical and clinical parts of the dossier cross-reference to parts of the reference mAb dossier whenever possible. FromSetting the stage for biosimilar monoclonal antibodies Christian K Schneider, Camille Vleminckx, Iordanis Gravanis, Falk Ehmann, Jean-Hugues Trouvin, Martina Weise & Steffen Thirstrup Nature Biotechnology 30, 1179–1185 (2012) doi:10.1038/nbt.2447 The State of the Art in the Development of Biosimilars Mark McCamish and Gillian Woollett Nature Biotechnology 30, 1179–1185 (2012)

Manufacturing Process (1) Demonstrate the Consistency and Robustness of manufacturing process cGMP implementation Modern QA/QC procedures In-process controls Process validation Minimize Differences b/w SBP vs. RBP Reduction clinical testing requirements of SBP Minimize any predictable impact on clinical safety and efficacy of SBP Differences are expected and may acceptable Appropriate justification  lack of impact on clinical performance

Manufacturing Process (2) Current manufacturing process following current guidelines (ICHs) and extensive quality control is encourage Potential impact of Changing of on Quality Safety and Efficacy should be identified. Some Differences are expected and may acceptable Appropriate justification (based on scientific and clinical experience  lack of impact on clinical performance

Comparability Exercises in Quality of Drug substance General Principles Manufacturing Processes To ensure the similarity of drug substance in SBP vs. RBP (vs. Reference standard) Products should be expressed and produced in the same host cell type Expression vectors Cell banks Cell culture/ Fermentation Harvest Purification Modification reaction Filling / Storage Dosage form Container closure system

Analytical Techniques Using appropriate, state-of-the-art in Biochemical, Biophysical, Biological Provided on Primary and higher-order structures Post translational modifications Biological activity Purity Impurities Product-related substance Immunochemical properties

Justification of Analytical Methods Suitability State-of-the-art (Current and up-to-date) Able to detect any difference between SBP vs. RBP Method Validations

Physicochemical Assessment A comprehensive understanding and description of the RBP vs. SBP to support development throughout the manufacturing process to Understand impact of manufacturing changes Ensure continued and consistent biosimilar properties to the reference product Build a database to support the Quality by Design (QbD) program Orthogonal methods/multiple techniques employed to fully elucidate structure (as far as possible) including microheterogeneities

Physicochemical Assessment Extensive enough to fully characterize primary and higher order structure including post translational modifications Primary structure, sequence and bridging (disulphide, thioether) Higher (secondary, tertiary and quaternary) order structure C and N terminal variants Post translational modifications Protein concentration Surface charge Protein size Aggregation Bioassay, potency Purity and impurities

Wisit Tangkeangsirisin, Biosimilars Guideline Public Hearing Biosimilar filgrastim compared to reference product (Neupogen) using three state-of-the-art analytical techniques Biosimilar filgrastim compared to reference product (Neupogen) using three state-of-the-art analytical techniques. The State of the Art in the Development of Biosimilars Mark McCamish and Gillian Woollett The State of the Art in the Development of Biosimilars Mark McCamish and Gillian Woollett 2/22/2013 Wisit Tangkeangsirisin, Biosimilars Guideline Public Hearing

Biological Activity Assessment Biological activity based on clinically relevant assays are required The extent is, to some degree, predicated by the physicochemical data All clinically relevant functional attributes to be assessed where possible E.g., for a mAb it may be necessary to test for both antigen binding and effector binding in appropriate in-vitro bioassays Relevance may determine extent of further nonclinical (and clinical studies) Animal toxicity studies not useful if no suitable model Studies should be comparative and provide a meaningful toxicological comparison Stand-alone toxicology study can be justified Safety pharm, repro toxicity, and carc studies not generally warranted Single-dose PK and PD study in animals can be useful

Specifications ICH Q6B : Specifications: Test Procedures and Acceptance Criteria for Biotechnological/Biological Products Data from batch-to-batch consistency, pharmaceutical development, comparability exercise in quality, safety and efficacy profile Range of acceptance should not wider than RBP

Possible biophysical and clinical outcomes for biosimilars Full Comparability Studies for Safety and Efficacy Biophysical Comparison No significant differences Significant differences Clinical Results No detectable difference Bioequivalence Based Approach Possible biophysical and clinical outcomes for biosimilars

Biosimilar Development Understand a Reference Product Characterization Identify Critical Quality Attributes (CQA) Creating and Producing a Biosimilar Identify the suitable clones Production process development Throughout Comparison of the Reference Product vs Biosimilar Analytical testing and Clinical study Totality of Evidence for similarity

Biosimilar Development

Data requirements for comparability exercises for a moderate risk and a high risk manufacturing change by the same manufacturer compared with a biosimilar manufacturing process. http://www.amgenbiosimilars.com/manufacturing/comparability-vs-biosimilarity/

http://www. amgenbiosimilars http://www.amgenbiosimilars.com/~/media/amgen/full/www-amgenbiosimilars-com/content/3-1-biosimilars-critical-quality-attributes.ashx

Therapeutic Mab Critical Quality Attributes Charge Variants Glycolysation Immunogenicity Aggregation & Degradation Primary & High Order Structures

RED = always very high criticality BLUE= at least high criticality BLACK = variable criticality

Why do we need state of the art analytics?

Quality: Analytical Analytical comparison is more than “Just Meets Specs” Possible Analytical Comparison Outcomes Analytical Methods Alone Confirm High Similarity Analytical Methods Not Sufficient or Adequately Discerning Analytical Methods Confirm Differences, but Differences Are Shown to Be Not Practically Significant Eg. HX575-higher phosphorylated mannose type glycan Analytical Methods Confirm Differences, but Significance of Differences on Safety and Efficacy Cannot Be Ruled out Analytical Methods Confirm Not Comparable Analytical Methods is better

Changes in Process: Changes in Products Quality Attributes ARANESP

State of the Art Analytics http://www.amgenbiosimilars.com/the-science/state-of-the-art-analytics/

RITUXAN

ENBREL

Case studies on biosimilar approval in EU

Omnitrope (somatropin) RBP = Genotropin During development, changes by increase complexity of comparability demonstration Esp. Additional steps to reduce Host Cell Protein V. high level of non-neutralizing antibodies (about 60%, during clinical trials (old process))

Binocrit (Epoetin alpha) RBP = Eprex Extensive characterization and comparability exercise in quality Structure comparisons Qualitative similar Quantitative differences found (increase in high mannose-6-phosphate, Decrease in N-glycolyl-neuraminic acid (NGNA) Differences was justified PRCA concerns SC route contraindication in CKD

Silapo (epoetin zeta) RBP = Eprex Extensive characterization and comparability exercise in quality Structure comparisons Qualitative similar Quantitative differences found (increase in des O-glycan forms, Decrease in NGNA Differences was justified SC route initially contraindicated SC indications added post-authorization upon further clinical data

Zarzio (filgrastim) RBP = Neupogen Extensive characterization and comparability exercise in quality Clinical data Phase I: Comparative PK/PD studies in healthy volunteers. Efficacy endpoints: neutrophil and CD34 cell counts Phase III: Non-comparative (single arm) clinical trial in cancer patients undergoing chemotherapy. Safety focus G-CSF Guideline states: “The recommended clinical model for the demonstration of comparability of the test and the reference medicinal product is the prophylaxis of severe neutropenia after cytotoxic chemotherapy in a homogenous patient group (…). Alternative models, including pharmacodynamic studies in healthy volunteers, may be pursued for the demonstration of comparability if justified”

Valtropin (somatropin) RBP = Humatrope Different expression system compared to reference medicinal product (S. cerevisiae vs. E.coli). Process specific HCP (yeast) assay required Changes during development subject to additional comparability Clinical trial: Initially calculated to demonstrate non-inferiority US sourced RBP used (supportive data) Indication differ from Omnitrope Pediatric indication for Omnitrope only: Small gestational Age, Prader-Willi Syndrome May 2012 Voluntarily withdrawn for commercial reasons

Insulin Marvel (Withdrawal) RBP = Humulin S Three forms (Short, Intermediate (30/70), Long acting) Quality Concerns Incomplete comparability exercise, esp Drug Product Inadequate process validation Missing batch traceability More data required for extended release forms Clinical issues Comparative PK/PD: euglycaemic clamp – most sensitive model Similar PK Parameters, but not similar PD profiles (faster glucose absorption Applicant resorted to efficacy trial with HbA1C end-point, not sufficiently sensitive Limit Immunogenicity data

Alpheon (rhIFNalpha-2a; Negative) RBP = Roferon-A Quality concerns Stability and impurities for drug substance and drug product profile also not matching RBP Inadequate process validation in DP No comparability studies between clinical trial batches and commercial batches Non-clinical studies were inadequate and indicated differences Clinical issues Difference in virological relapse rates Inconclusive data in response rate for Genotype 1 patients Different rate of adverse events Inadequate immunogenicity documentation

Some Concerns About Biosimilars Route of Administration for approved indications of biosimilar may not be the same as Reference Biological Product Eg. Binocrit initially contraindicated in CRF by SC route

Different glycan pattern--Not biosimilar REALLY???????? HX575 has different glycan profiles from RBP. But the differences are acceptable and justified.

EPO Isoforms among Biosimilar vs. RBP are similar. Not Identical

“ More recently, two cases of neutralizing antibodies were reported during a pre-marketing clinical trial of a biosimilar epoetin (HX575) in subcutaneous administration in chronic renal failure subjects (9). In this paper, we present data showing that contamination by tungsten during the manufacture of the syringes used for primary packaging is the most likely cause of protein denaturation and aggregation, which may have led to the occurrence of two cases of neutralizing antibodies in the investigational clinical trial.

Conclusion Quality assessment is the primary and major part of biosimilar registrations. Comparability studies with RBP determine the degree of intensity in non-clinic and clinical assessment. State-of-the-Art technology should be used to detect any potential differences. How to demonstrate product comparability is challenging Case studies about EU biosimilar approval confirmed case-by-case varies

Any questions, comments and suggestions are welcomes Thank you Any questions, comments and suggestions are welcomes