Comparability Assessment of BioTherapeutics: Paving the Way for Late-Stage Projects

Transcription

1 Comparability Assessment of BioTherapeutics: Paving the Way for Late-Stage Projects Olga Friese, PhD Associate Research Fellow Pfizer Biotherapeutics PharmSci 28 January 2014

2 Overview Manufacturing process changes throughout product life cycle Types of process changes and potential product impact Establishing Comparability Challenges and expectations for mab Biotherapeutics Risk assessment Analytical considerations and tools utilized Case Study Process 1 to 2 improvements Summary 2

3 Typical Manufacturing Process Changes Throughout Product Life Cycle Batch scale up for clinical trials Increased need for clinical trial material for later phase studies Larger bioreactor suite at the same site Process improvements as knowledge is gained Reduction in impurities or improvement in quality Increase in efficiency or yield Change of raw materials and/or equipment Change in business strategy Site changes for resourcing optimization Internal facility at different location External CMO Change in formulation/dosage form Improved stability profile/shelf life Increased strength Change in container closure/introduction of device Improved dosing regimen Improved product profile for the market 3

4 Representative Antibody Manufacturing Process B. Minow, P. Rogge, K. Thompson, BioProcess International, Vol. 10, No. 6, 2012, pp

5 Process Changes and Potential Impact to Product Quality Type of Change Examples Potential Impact to Product Quality DS - manufacturing site and/or scale New CMO, new suite in same facility DS - upstream process DS downstream process DP - manufacturing site and/or scale DP - process DP - formulation or dosage form Cell line/clone, raw materials, feed strategy, ph, time, and fermentation temp, cell culture conditions, scale Column/resin, loading, cycling and hold times, reagents, gradient New CMO, new fill/finish line in same facility Shipping, equipment, container/closure/device Liquid to lyo or vice versa, strength, excipients, ph Purity/impurities, stability, primary structure; isoforms; PTMs Stability, higher order structure 5

6 Comparability Challenges and Expectations for mab Biotherapeutics Challenges Proteins are inherently heterogeneous Biosynthetic processes in living organisms result a mixture of post-translationally modified forms Examples of mab heterogeneities are glycosylation, deamidation and disulfide isoforms State-of-the art analytical technologies often required to detect product quality changes Expectations Establish comparability post-change as per ICH Q5E The demonstration of comparability does not necessarily mean that the quality attributes of the prechange and post-change products are identical; but that they are highly similar and that the existing knowledge is sufficiently predictive to ensure that any differences in quality attributes have no adverse impact upon safety and efficacy of the drug product. Process improvements are likely to impact the product Perform a risk-based assessment - no adverse effects on product quality, safety or efficacy Focus on changes to quality attributes (critical or non-critical) Rely on capacity to detect significant product quality changes Understand the magnitude of the process changes Leverage platform experience with the product Build on overall clinical experience the likelihood of impact on safety and efficacy 6

7 Process Changes and Potential Risk to Comparability Low Risk Moderate Risk High Risk Examples of Changes Replicate equipment Reagent storage conditions Change specifications Updating tests to meet new compendial requirements Minor formulation change New WCB from approved MCB Manufacturing site change (internal move lower risk) Process scale-up Change purification step Change filtration step Change test methods Substitute raw materials Change cell line Major change to fermentation recipe New recovery and purification procedures Major formulation and/or strength change Dosage form change Analytical data Possible Supporting Data Scientific rationale Analytical data Analytical data (Non-clinical data)* (Non-clinical data)* (Clinical data)* (If necessary)* 7

8 Comparability Assessment Attributes and Methods Attribute* Primary Structure Characterization Molecular mass Amino acid sequence Amino acid composition Methods* Mass spectrometry intact mass, LC/MS - peptide mapping, AAA, Edman sequencing Post-translational Modifications Disulfide bonds, thiols Glycosylation: N- and O-linked Isoforms/Degradation Products Aggregate Product-related substances Charge and size isoforms Degradation products Higher-order Structure Secondary and tertiary structure Biological Activity Receptor and/or ligand binding Bioactivity LC/MS - subunit analysis, LC/MS - peptide mapping, Glycan assays, Ellman s Assay SEC, RP-HPLC, IE-HPLC, ice, Electrophoresis, LC/MS - peptide mapping, AUC, LS, FFF CD spectroscopy, DSC, Fluorescence spectroscopy, FTIR Binding assays, SPR, Cell based assays, in vivo assays * Not inclusive 8

9 Comparability Assessment - Strategy Assess analytical comparability through Release methods -measure quality attributes that define identity, purity, potency, safety -compare batch release data pre- and post-change, potential statistical analysis -if assay has changed may repeat side-by-side analysis of comparability samples Heightened characterization methods - similar to methods used for reference material characterization (ICH Q6B) with some exceptions - potential changes based on process changes, not full characterization of multiple batches - in-process testing may be used to compare process changes - typically 3 batches from each process, tested side-by-side - it is similar to the strategy for process changes prior to Phase 3 Stability studies - accelerated stability studies may be used to establish degradation profiles and provide additional direct comparison of pre- and post-change product - if no accelerated stability data available, side-by-side forced degradation studies: thermal stability at accelerated temperatures and photo-degradation Note: To mitigate the risk, a supportive comparability assessment of pilot scale run of new process is recommended 9

10 Comparability Assessment Data Evaluation Release methods assess quantitative assays results for Process 2 against both pre-established Process 2 specification and Process 1 historical ranges no widening of the Process 2 acceptance criteria Heightened characterization methods primary structure, posttranslational modifications, higher order structure results will be assessed qualitatively in a side-by-side comparison Stability studies quantitative assays results for Process 2 will be assessed against historical ranges of Process 1 if accelerated stability studies data is used if side-by-side forced degradation study is performed, compare ranges for Process 2 and Process 1 assess qualitatively heightened characterization data in both cases Note: If Process 2 data is outside of historical ranges for Process 1, risk assessment of impact on safety and efficacy needs to be performed; Difficult to implement statistical analysis due to a small number of lots for late stage process. 10

11 Example of DS Process Changes - Process 1 to Process 2: Case Study for a Late-Stage Project Process 1 Process 2 Process Changes Rationale for Process Change 2500-L Scale L Scale Commercial scale MCB WCB Introduce working cell bank High Seeding Density Production Process Protein A Chromatography Increased Titer Process Protein A Chromatography new resin Increase process titer; nutrient delivery based on process understanding, increased amounts of nutrients delivered to sustain productivity Capture mechanism does not change; improved capability due to resin properties Potential Product Impact Primary structure, PTM, isoforms, purity, stability Primary structure, PTM, isoforms, purity, stability Primary structure, PTM, isoforms, purity, stability Purity/impurities 1 conc. 1.5 conc. DS concentration increase to manage commercial scale DS volumes (formulation remains constant between supplies) Higher order structure EVA bags Stainless Steel Cryovessel Batch volume, scale appropriate, improve logistics Stability 11

12 Release Test Showed Increase in Basic Species in Process 2 Pilot Run Material Quality Attribute Process 1 (n=3) Process 2 pilot (n=1) Charge Isoforms Acidic species Basic species 27.6% 2.3% 27.1% 2.1% 27.9% 2.0% 28.9% 8.5%* Heightened characterization by the three-part subunit domain assay showed that increased basic species correspond to the heavy chain C-terminal Pro amide * ice data 4 x10 3 G0F x G0F- GlcNAc Man5 G0 Pro amide G0F G1F Process Man5 G0F- GlcNAc G0 Process m/z G1F 12

13 Risk Assessment for Increase in the Heavy Chain C- terminal Amidated Proline in mabs Literature Overview Several papers since original discovery by Johnson et al. (2007) The heavy chain C-terminal amidation reaction is catalyzed by peptidyl glycine alpha-hydroxylating monooxygenase (PAM) Copper (Cu) is critical for the catalytic function of the PAM Supplementing copper in the production medium helps to maintain cell viability and improve mab titers often needed for late stage projects Effect of copper concentrations on the basic peak levels T. Kaschak et al, mabs 3:6, ;

14 The Heavy Chain C-terminal Pro Amide Does Not Impact FcRn Binding Affinity or Animal PK ice profile of tested materials Subcutaneous administration of 10 mg/kg in normal rats Charge variants of IgGs do not affect the in vitro potency, FcRn binding affinity or the PK properties in rats. Thus no impact on safety or potency is anticipated L. A. Khawli et al, mabs 2:6, ;

15 Disulfide Isoform Distribution Difference Due to Process Changes Process 2 lot 2 #1 #2 #3 #4 Lot 02 Lot 05 Lot 08 %Peak# %Peak# %Peak# %Peak# Process 2 lot %Peak#1 %Peak#2 %Peak#3 %Peak# Process 1 lot 8 Area% * RP-HPLC profiles of disulfide isomers 0.0 Lot 02 Lot 05 Lot 08 Change in disulfide isomers due to increase in mab residence time in production bioreactor and longer hold time in broth; No significant impact on potency measured by binding ELISA and cell-based assay for materials with different relative levels of disulfide isomers; * RP-HPLC method: Dillon, T., et al., J. Biol. Chem. 2008, 283,

16 Risk Assessment of the Disulfide Isomer Changes in mabs Literature Overview and Scientific Rationale Mixture of IgG2 disulfide isomers found in endogenous antibodies isolated from human blood (Dillion et al., 2008 and Wypych et al., 2008); Mild redox conditions redistribute the relative ratio of A, A/B, and B isomers in vitro and in vivo (Dillion et al., 2008); Disulfide reshuffling occurs in vivo with decrease in IgG2-A and increase in IgG2-B (Liu et al., 2008); Typically, late stage process changes result in increase in IgG2-B isomer; 16

17 Change in the Host Cell Leads to a Different Glycan Profile an Example of Late Stage Process Change Glycan profile of IgG2 mab G0F G1F G2F NS0 G0F minus GlcNAc Man5 G1F minus GlcNAc G0F G1F Man5 G1F minus GlcNAc+NeuGc G2F G0F minus GlcNAc G0 G0 minus GlcNAc Man6 G0-Hybrid CHO G0 minus GlcNAc G2F+α-Gal Minutes 17

18 Risk Assessment for Changes in Glycan Profile Scientific Rationale and Literature Overview Both processes have same major N-linked glycans; Some difference in the abundance of major and minor species; CHO derived glycans are less complex and more aligned with those in human mabs: lower levels of Man5; absence of Gal-α(1-3)-Gal (alpha-gal) species; absence of N-glycolyl neuraminic acid (NeuGc) moeities; Slight variations in the relative abundance of major glycans and elimination of the minor species is not expected to have any impact to the clinical safety; Composition of the Fc N-glycan is not essential for MOA of the IgG 2 subclass of antibodies - impact on clinical efficacy is not expected; Literature suggests a possible impact on PK non-clinical PK study to mitigate the risk should be considered. 18

19 Summary Manufacturing process changes often require comparability studies to be performed Site change, scale-up, process improvements, etc. Nature of the change dictates the comparability study design Predict potential impacts to product quality, safety, efficacy Analytical methods employed need to have sufficient resolution to determine changes Focused analytical methods based on change; powered appropriately Analytical comparability strategy needs to be defined release testing, heightened characterization, stability Differences assessed to evaluate their impact on safety and efficacy Consider whether methods are capable of detecting important differences Non-clinical or clinical studies may be required to assess comparability Requires regulatory discussion 19

20 Acknowledgement Jennifer Feng Andrew Saati Thomas Porter Bruno Figueroa James Carroll Jason Rouse Laura Bass 20

21 Back Up 21

22 Comparability Assessment by Release Testing for Process 1 and Process 2 DS and DP Drug Substance Drug Product Assay Side-by-Side Analysis Comparison to Historical Range Side-by-Side Analysis Comparison to Historical Range Appearance (Clarity) Appearance (Coloration) Appearance (Visible Particulates) ph Protein concentration by UV Spectroscopy (A 280 ) Sub Visible Particles Extractable Volume Peptide Map (Identity) N-linked Oligosaccharide SE-HPLC (Purity) CGE (Reducing) CGE (Non-reducing) ice (Charge Heterogeneity) Binding ELISA Cell-Based Bioassay Protein A ELISA Host Cell Protein ELISA Host Cell DNA Assay (qpcr) Sterility Bioburden Endotoxin Osmolality 22

23 Comparability Assessment by Heightened Characterization for Process 1 and Process 2 DS and DP Assay Drug Substance Drug Product Side-by-SideAnalysis Side-by-SideAnalysis Peptide Map (Reduced and Alkylated, LC/MS) Subunit Analysis (LC/MS) N-Linked Oligosaccharide (HILIC/MS) Intact Analysis (LC/MS) Disulfide Linkages (NR Peptide Map, RP HPLC) Sialic Acid Assay Near-UV Circular Dichroism Far-UV Circular Dichroism Differential Scanning Calorimetry (DSC) Sulfhydryl Analysis (Ellman s Assay) Target binding by SPR (Biacore) Up to 3 Process 1 DS batches/dp lots and 3 Process 2 DS batches/dp lots to be tested 23

24 Comparability Assessment by Forced Degradation for Process 1 and Process 2 DS and DP Degradation Pathway Time points Drug Product, side-by-side Photo-exposure 0, 8 (1152 K lux-hrs) and 14 (2015 K lux hrs) days Thermal Stability at 40 C 0, 7, 14, 28 days 3 Process 1 DS batches and 3 Process 2 DS batches formulated as DP to be tested 24