12-4-2011

Immunogenicity: What Do We Know and What Can We Measure?

Meenu Wadhwa, PhD
Cytokines and Growth Factors Section, Biotherapeutics Group, NIBSC - HPA United Kingdom

Conflict of Interest Disclosure

Meenu Wadhwa, PhD

Has no real or apparent conflicts of interest to report.

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Biotherapeutic Products
Biotechnology derived products have been successful for therapy of various diseases
Hormones, enzymes, coagulation factors, cytokines Fusion proteins, monoclonal antibodies

Despite their benefits, therapeutic proteins have the potential to induce an immune response due either to their
Foreign nature if of exogenous origin

or
Due to recognition as non-self

Immunogenicity Profile of Therapeutics

Product Name
Intron A Roferon Pegasys PegIntron Betaferon Avonex Rebif Eprex Neorecormon Procrit Aranesp Neupogen Neulasta Leukine Proleukin Enbrel G-CSF GM-CSF IL-2 TNFR 2-Ig Myeloregeneration, neutropenia Myeloregeneration, immunostimulation Oncology Psoriasis, RA NHL Rituximab Anti-CD20 CLL SLE Humira Fully human anti-TNF RA 26 12 Not detected 2 95 74 15 0 Epo Anemia Rare IFN- Multiple Sclerosis IFN-2a Hepatitis C

Protein

Indication

% Patients with Immune Response

7 25 9 1 25 45 26 12 28

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Unwanted Immunogenicity
Proteins Patients Non immunogenic (G-CSF, IFN-g)

Immunogenic (induce antibodies)

No effect (growth hormone, insulin)

Neutralize biologic effects and compromise further therapy (factor VIII, IFNa2a, GM-CSF)

Alter PK/PD

Cross-react with native protein and induce adverse symptoms (Epo, MGDF)

Potential Clinical Consequences of Immunogenicity

Benign, non-significant to serious, life-threatening depending on the therapeutic Efficacyreduction of the clinical response Safetysafety issues can occur even when there is no loss of efficacy Acute consequences Infusion reactions, anaphylactic reactions Non-acute consequences Delayed-type hypersensitivity/immune complexes Cross-reactivity with an endogenous counterpart

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Factors Influencing Immunogenicity

Product & treatment related
Protein (structure, primary sequence, novel epitopes, glycosylation, oxidation, deamidation)
Product impurities, formulation, aggregation, degradation, Properties of the protein eg, immunostimulatory, replacement therapy, physiologically important? Dose, route, frequency of administration and duration of therapy

Patient related
Age, gender, genetic make-up, ethnic sensitivity (IFN-2a more immunogenic in Chinese vs Caucasian hepatitis patients 39% vs 14%), immune status, disease

Production Process
Complex manufacturing process

New manufacturer (copy product) Manufacturing and/or purification process different, no access to knowhow of the process/history of product; limited to public domain information Step-Wise Process Changes Expression system/vector Fermentation/ cell culture process Purification process Formulation and filling Drug Product

Product likely to have altered characteristics

Carson KL, Nat Biotechnol. 2005; 23 (9):1045-1048.

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Complexity of Proteins
Biosimilars in the EU
Comparability studies are needed to demonstrate the similar nature, in terms of quality, safety and efficacy, of the new similar biological medicinal product and the reference product authorized in the EU
Emphasis has been given to immunogenicity Any subtle change introduced in the manufacturing process of a given product can have enormous implications for immunogenicity

Unwanted Immunogenicity
Current Position
Testing for unwanted immunogenicity is integral to product development (clinical & post-marketing phase) for ensuring: Clinical safety of a biotherapeutic Product comparability When a biosimilar product is developed

EMA Guidance
Guideline on Immunogenicity Assessment of BiotechnologyDerived Therapeutic Proteins EMA/CHMP/BMWP/14327/2006 Guideline on Immunogenicity Assessment of monoclonal antibodies (in consultation) EMA/CHMP/BMWP/86289/2010

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EMA Guidelines on Biosimilars

In revision
(quality issues)

Consultation
mAbs

In Preparation
Revised IFN-b, FSH

Recent

LMWH

IFN-a

European Medicines Agency. http://www.ema.europa.eu/ Accessed 23 February 2011.

Unwanted Immunogenicity
Prediction of immunogenicity
In silico and T cell methods are promising but information on the true clinical utility of these approaches in a prospective manner is lacking

Determination of immunogenicity
Human clinical data neededcannot be replaced by use of animal or in vitro or in silico tools

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Unwanted Immunogenicity The Most Challenging Issues

It is impossible to predict The incidence of unwanted immunogenicity The characteristics of the immune response The clinical consequences & significance of such immunogenicity These require assessment in carefully designed studies

Immunogenicity Testing: A Tiered Approach

Screening assays: for identification of all antitherapeutic binding antibodies - Enzyme-linked immunosorbent assays (ELISAs): direct, bridging, other formats - Radioimmunoprecipitation assays (RIPA) - Surface plasmon resonance (SPR) - Other technologies Confirmatory assays: for confirming antibodies

Neutralization assays: for distinguishing neutralizing & non-neutralizing antibodies. - Cell-based assay - Non-cell-based ligand binding assay

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Strategy for Assessment of Immunogenicity

Patient samples taken at appropriate time-points

-ve samples rejected

Screening Assays Confirmatory Assays

+ve samples

Bioassay

Confirmed +ve samples

Characterization

Assess correlation of characterized antibodies with clinical responses to biologic therapeutic Assays for clinical markers and assessment of clinical response in patients

Immunogenicity Assessment Strategy Design and Interpretation

Studies need to be carefully and prospectively designed to ensure all procedures are in place prior to initiation
Selection, assessment, characterization and validation of assays Identification of appropriate sampling points, duration of testing Sample volumes and sample processing/storage Selection of statistical methods for analysis of data

This applies to all assays as shown in strategy slide Strategy needs to be established on a case-by-case basis product, patients, expected clinical parameters
In chronic use sequential sampling for a year In view of variability of antibody responses, adequate numbers of patients needed

However, unwanted immunogenicity may occur at a level, which is not detected in studies pre-approval so assessment post-approval, as part of pharmacovigilance surveillance is needed

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Comparative Immunogenicity
Studies need to be designed to demonstrate whether the immunogenicity of the products is the same or significantly different This may affect the design of the studies & their interpretation A homogeneous and clinically relevant patient population should be selected. Head-to-head studies using same assays & sampling strategy needed The consequences of immunogenicity must also be compared Post-approval assessment may be necessary, usually as part of pharmacovigilance surveillance

Immunogenicity of Biosimilars
The immunogenicity of the marketed product does not influence the need for comparative immunogenicity studies However, if the imunogenicity profiles of marketed and biosimilar products are significantly different, they may be considered DISSIMILAR

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Antibody Frequency for Biosimilars

(Presubmission Studies)
Biosimilar Omnitrope (SC) Valtropin (SC) Binocrit (IV) Silapo (IV) Silapo (SC) Ratiograstim (SC) Zarzio (IV / SC) (phase 1, crossover) Nivestim Alpheon (SC)

Ab Incidence 0/51 (0.0%) 3/98 (3.4%) 2/314 (0.6%) * 0/305 (0.0%) 0/323 (0.0%) 7/356 (2.0%) 0% 3/183 (1.6%) 31/111 (27.9%) **

Reference Genotropin Humatrope Erypo Erypo Erypo Neupogen Neupogen Neupogen Roferon-A

Ab Incidence 1/44 (2.3%) 1/49 (2.0%) 3/164 (1.8%) * 0/304 (0.0%) 0/230 (0.0%) 2/134 (1.5%) 0% 0/95 (0.0%) 38/99 (38.4%) **

* Prevalence; ** at end of treatment; Assay validation insufficient

Courtesy Martina Weise (Bfarm)

Application refused or withdrawn;

Antibodies and Adverse EffectsEPO

>Sixty PRCA cases identified in Thailand. Fourteen EPO products marketed. Link to product(s)? Safety Study for Binocrit Suspended No increased immunogenicity from IV use in patients with renal anaemia or SC use in cancer patients (both licensed) Postmarketing SC trial in previously untreated renal anaemia patients: two cases of neutralizing Ab Cause(s) ? Binocrit approved - 2007 Rigorous physico-chemical, biological characterization & clinical trial data
Brockmeyer C, et al. Eur J Hosp Pharm Prac. 2009;15(2):34-40.

Pure red-cell aplasia (PRCA) and anti-EPO antibodies in patients treated with EPO (EPREX) 2002 - 13 cases in chronic renal failure patients, rapid development of severe transfusion dependence within months of therapy, resistant to other EPO products Pre 1998 2/3 cases 1998 to June05 260+cases worldwide
Casadevall N, et al. N Engl J Med. 2002;346(7):469-475.

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Immunogenicity of Other Products

IFN- - Incidence of neutralizing antibodies varies

Avonex 2% to 6%; Rebif 12% to 28%; Betaferon 25% to 45% Loss of efficacy; abs cross-reactive and impact on disease management and QOL Monoclonal antibodies Ab incidence dependent on product, disease, age; loss of efficacy noted in some cases. Antibody Incidence % 61 8 0 65 1.9 29-63

Product Remicade

Type Anti-TNF

Disease Crohn RA Non-Hodgkin SLE CLL B-CLL RA

Rituxan

Anti-CD20

Campath

Anti-CD52

Immunogenicity Guideline
Recommendations for routine monitoring of changes in clinical response and linking immunologic findings to clinical events Immunogenicity as part of all clinical trials ( indication
specific differences possible)

Evaluate all patients ( and not in a symptom-driven manner) Standardize sampling schedule as much as possible

Specifically analyse adverse events for linkage to an unwanted immune response (ie, also in a symptom drivenmanner)
Provide guidance on how prescriber should handle patient in case unwanted immune response occurs (increase dose/discontinue, etc)

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Biosimilars: Unwanted Immunogenicity

Quote from EMA BMWP chairman: Unwanted Immunogenicity is the biggest

challenge for the approval of Biosimilars

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