Bottlenecks in Biopharmaceutical Development: Chemistry , Manufacturing, and Control

1. Bridging the Gap – Overcoming Bottlenecks in the Development of Therapeutics for Infectious Diseases Bottlenecks in Biopharmaceutical Development: Chemistry, Manufacturing, and Control Steven Giardina, Ph.D., RACSenior Scientist SAIC August 2012

2. Topics • Brief history of cGMP for biopharmaceuticals • CMC Issues – Perils of not planning backwards • Building – Not testing – Quality into the Product • Product Safety – How soon do I need to start worrying? • Case Study – Ch14.18 – A monoclonal antibody for treatment of late-stage neuroblastoma (or making your own biosimilar without intending to) • Lessons Learned

3. Foundations of GMP • All drugs (small molecule and biologic) have to be shown to be safe and effective. • It was not always so. • The history of drug law shows us that most of the laws and regulations we have today were in response to tragic events. • A sampling…

4. It all started with a horse named “Jim”

5. Diphtheria Antitoxin • Made in old milk horses – like Jim. In St. Louis, Missouri. The blood of a tetanus-infected retired milk-wagon horse was used for this purpose. Thirteen children who were given the antitoxin died from tetanus. *Led to the Passage of the Biologics Control Act of 1902.

6. Guidance Documents • Put meaning to the ambiguities in the statutes and regulations. • Points to consider • ICH (International Conference on Harmonisation) • Do not have the force of law but the FDA does expect investigators to adhere to them unless they have an equally sound approach. • Can be found on FDA web site

7. GMP Rules of Thumb • If it is not written down – it never happened. • Until you know it does not matter – it matters. • GMPs really begin at the bench prior to what would normally be perceived as the true beginning of GMP PD and manufacture. • Important to keep good records of anything that is needed for inclusion in the Chemistry, Manufacturing, and Control (CTD Module 3). • What would affect a decision on product safety? Can reach back into the origins of the discovery phase of PD. • You cannot test quality into the product – it must be built into all aspects of the drug’s manufacture (quality systems and risk management). • A product is adulterated – by definition – if it is not made in conformance with cGMP requirements even if it tests to spec.

8. Example from ICH-Q5D – Cell Substrate – Cell Origins • The cultivation history of the cells should be documented. The method originally used for the isolation of the cells should be described as well as the procedures used in the culturing of the cells in vitro and any procedures used to establish cell lines (for example, use of any physical, chemical, or biological procedure, or added nucleotide sequences). A description of any genetic manipulation or selection should be provided. All available information regarding the identification, characteristics, and results of testing of these cells for endogenous and adventitious agents should be provided.

9. Common CMC Pitfalls – Biologics • No traceability on what was used to manufacture the drug. • Origin of DNA used to make cell substrate? • Chain of custody/identity of cell lines, vector, plasmids, etc. • Exposure to animal-derived raw materials • No scientifically valid assays for purity, potency, etc. • No agreement on what pure and potent even mean • Impurities/residuals • LPS levels? • Genomic DNA? • Host Cell Proteins? • Aggregates? Clipped forms? Other product-related “impurities.” • No qualified reference standard for establishing “scientifically sound” assays before they are needed to track process development and product quality during manufacture, release, and stability period.

10. Common CMC Pitfalls – Biologics (cont.) • What are the stability-indicating assays? Not just a list of what is easy to do – e.g., SDS-PAGE may be easy and needed but not necessarily stability indicating. • Drug does not meet obvious GMP requirements. • Inadequate supply. • Formulation does not ensure stability over time and during shipment. • Optimal formulation/fill-finish/concentration/amount per vial, etc. for actual use over duration of the trial? • No agreement on release criteria. • Inadequate documentation of what was done and when. • Poor quality systems (investigations, deviations, not completed in a timely manner). • Executive management and QA are “missing in action.” • Hubris.

11. Drug Substance and Drug Product • Release vs. characterization (FIO) assays. • Reference standard – why is it suitable for the purpose? • Impurity profiles – product or process?? • Stability in bulk or primary container – outsourcing to CMOs for different aspects of the product manufacture? • Shipping stability – cold chain? Lyophilization? Mechanical stress? • Drug product stability for toxicology studies? GLP requirement. • Drug product stability under proposed conditions of administration – each center may be quite different despite what is in the Investigator’s Brochure. • Lost due to adsorption on the IV bag and infusion set tubing? • No toxicity = no drug administered? Proof?

12. Purity • 21 CFR 610.13 • “Products shall be free of extraneous material except that which is unavoidable in the manufacturing process described in the approved biologics license application.”

13. Identity • 21 CFR 610.14 • “The contents of the final container of each filling of each lot shall be tested for identity after all labeling operations shall have been completed. The identity test shall be specific for each product in a manner that will adequately identify it as the product designated on the final container and package labels and circulars, and distinguish it from other product being processed in the same laboratory. Identity may be established either through physical or chemical characteristics of the product, inspection by microscopic or macroscopic methods, specific cultural tests, or in vitro or in vivo immunological tests.”

14. Certificate of Analysis • Product Name: XX Project Number: YY Part Number: ZZ • Date of Manufacture: 08/10/12 Lot Number: L123456 Lot Size: 3000 vials • Container Size/Fill Volume: 3ml/0.5 mL Storage Temperature: 2-8 °C • Description: A chimeric monoclonal antibody (hIgG1) to the GD-2 disialoganlgioside antigen. Colorless liquid at a concentration of 5mg/mL in a solution of 10 mM sodium phosphate, 150 mM sodium chloride, pH 7.4 TEST DESCRIPTION SPECIFICATION SOP OR QC RESULT STUDY REPORT # Identity Appearance Colorless liquid 123 456 Colorless liquid • Essentially-free of particulates Essentially-free of particulates SDS-PAGE/R and NR/ Report Major Bands 444 777 166.8 kDa (NR) Densitometry HC @ 51.29 kDa (R) LC @ 28.58 kDa (R) Human IgG Subclass human IgG1 – kappa subclass 333 888 human IgG1κ Content Protein Concentration E @ 1mg/mL = 1.0 222 111 5.14 (5.1) by Absorbance @ 280nm Potency GD-2 ELISA Binding within 50-200% of 000 222 Binding is 104% of Standard Standard 1

15. Certificate of Analysis (con’t) • Product Name: XX Project Number: YY Part Number: ZZ • Date of Manufacture: 08/10/12 Lot Number: L123456 Lot Size: 3000 vials • Container Size/Fill Volume: 3ml/0.5 mL Storage Temperature: 2-8 °C • Description: A chimeric monoclonal antibody (hIgG1) to the GD-2 disialoganlgioside antigen. Colorless liquid at a concentration of 5mg/mL in a solution of 10 mM sodium phosphate, 150 mM sodium chloride, pH 7.4 TEST DESCRIPTION SPECIFICATION SOP OR QC RESULT STUDY REPORT # Purity HPLC-SEC Monomer purity ≥ 95% 666 222 Monomer purity:97.2% cIEF Report % and pI of major forms 222 666 Single peak at pH 5.5 Safety Sterility No Growth USP 26 <71> 555 No Growth Endotoxin < 5EU/mg 333 777 1EU/mL(0.19 EU/mg) gDNA Content Report Results 111 999 0.5 pg/ML murinegDNA pH 7.5 ± 0.4 XXX YYY 7.3 1. reference standard is lot L098765

16. Certificate of Analysis (con’t) • For Information Only – Possible Additions – Comparability? • Silver Stain – impurity/degradation • Western Blot with Product Specific Antibody -Identity • N-terminal Sequencing – Identity • BiolayerInterferometry or Surface Plasmon Resonance (kinetics/KD) w/ conformation sensitive ab or antigen or ?? – changes in structure = telegraphing a change in potency?? • Carbohydrate Composition – glycoform distribution and identity • Host Cell Proteins – Important and a problem when using commercial kit s • Process Impurities – BSA?, Bovine IgG ( if using serum-containing medium) • Other chemicals/reagents that should be removed – antifoams? Selective agents?

17. COA – Last Page • Signatures and More Signatures – • QC – attests that data are true and accurate • Project Scientist • Program Director (Go Directly to Jail) • QA • Customer • Caution Statement – proscibed based on intended use • Not for use in humans? ( as opposed to not for human use) • For investigational use… • Name and Address of the Manufacturer of Record – not the subs

18. The Challenges of Making a Biologic • Biological molecules – like immunoglobulins – are complex and large. • Complexity can be increased due to processing such as glycolsylation. • How do we define “pure” if there are multiple forms of the product? • How do we define its biological activity/potency? • Which is not the same as efficacy!

19. Case Study – When is a product difference = a different product?

20. New Therapy for Pediatric Neuroblastoma • Most common extracranial solid tumor in infancy. • One of the most common malignancies of childhood. • About 600–700 cases per year in the United States. • Patients with low/intermediate risk neuroblastoma have excellent prognosis and outcome. • High risk = poor outcome despite intensive therapy. • 70%–80% of patients older than 18 months present with metastatic disease. Less than half are cured even with the use of high-dose therapy followed by autologous bone marrow or stem cell rescue.

21. Ch14.18 Milestones • MAb against GD-2 disialoganglioside is shown to be safe and effective in refractory neuroblastoma. • Treatment includes the use of IL-2 and GM-CSF to increase efficacy via enhanced ADCC in addition to 13-cis-retinoic acid. Not a benign therapy (pain, allergic reactions) • Multiple lots of ch14.18 manufactured in support of nonpivotal Phase III trial sponsored by the NCI over a period of ~7 years. • All product lots manufactured by SAIC-Frederick’s Biopharmaceutical Development Program (BDP).

22. Ch14.18 Milestones • In 2009 results of Phase III randomized trial showed event-free survival at 2.1 years was significantly higher (66%±5% vs 46% %±5%). • Overall survival also significantly higher (~86% vs. ~75%). • Based on these data ch14.18 randomization of the trial was stopped and ch14.18 + 13 CRA + CM-CSF + IL-2 became the standard of care.

23. New Challenges • Methods Legacy: The process used by the BDP in support of the NCI trials remains the only manufacturing process. • Undemonstrated Technology Transferability: The BDP was the only manufacturer of the product being used in the NCI-sponsored current trials. • System Capacity: Increased anticipated demand – still have existing process using hollow fiber production system. • Manufacturing Reproducibility: The challenge was to meet the demand focusing on lot-to-lot consistency using a process that was not originally designed to meet the new material requirements.

25. Reference Standard • Produced from a batch of material manufactured in 2003. • FDA has accepted the specification of “conforms to standard” for a number of attributes. • This became a problem when comparing pre-2010 lots with the most recent lots. • Quantitative specifications were developed as PD moved along. • Potency assays continue to be reported as the ED50 as well as the percentage of the reference standard.

26. In the Interest of Time • A large menu of standard assays (SDS-PAGE, SEC, etc.) showed no differences between the old and new lots • So – how about potency? • Binding to GD-2 expressing cells? • C’ mediated killing (Fc structure/function) • Binding to FcγIIIa receptor (effector-cell receptor) • ADCC (performed by Jackie Hanks/Paul Sondel, University of Wisconsin)

27. Relative Activity of Ch14.18 Lots (Potency Tests)

28. Comparison of KDs

29. Everything Looked Great • ADCC – not QC assay – also normal • No issues until…..

30. Early Lots of CH14.18 Show Comparable cIEF Profile

31. Current Lots of Ch.14.18 Show Quantitative Differences in Relative Peak Intensities Compared to Earlier Reference Lot signal pl

32. MS of Intact Ch14.18Reference Lot (Green) vs. Old Clinical Lot L0512003(Blue) = Similar

33. MS of Intact Ch14.18Reference Lot (Green) vs. New Clinical Lot L1001001 (Blue) = Not Similar

34. Relative Percent of Four Major N-Oligosaccharides by CE

35. Reason for the Change? • Too many years and changes in raw materials to pinpoint precise root cause. • No new charge species or glycoforms – only a change in relative distribution • Important to recall that there was no evidence of any change to the product by routine tests and no differences were reflected in 4 different potency tests. • Product made in same facility and by same people – still a change. • Time will tell if any of these changes have any impact on efficacy.

36. Consequences? • FDA allowed the product to be used transparently to the earlier lots but there was enhanced monitoring for AEs, stability changes. • Health Canada initially viewed the changes as a new product but also allowed the use of the new material in the ongoing trial. • Enhanced reporting. • Required that there be a new reference standard that looked more like the new clinical material and its stability be monitored under a formal stability protocol. • Many technical reports, ad hoc meetings to bridge/explain the differences and how to handle them both in the lab and, more importantly, in the clinic.

37. Take-Away Message • Biologicals are complex and, therefore, have unique CMC challenges that must be understood and anticipated long before the formal product development phase begins in earnest = Knowledge Management. • All data (productivity, stability, activity) can inform decision making down the road and have to be of the highest quality and retrievable. • Not having a useful (think QC friendly) potency assay, reference material, or an understanding of a product’s stability issues can have a profound impact on PD time and costs.

38. Take-Away Message (cont.) • AskWhat am I doing NOW that can affect the safety, identity, purity, potency, and overall quality of a drug I may want to make LATERfrom the cells, etc. I am handling in the lab (traceability of genetic material and contact with animal-derived RMs, cross-contamination?)? • Is this really the concept candidate I want to take forward into expensive tox and clinical trials? Is it “druggable?” • The consequences of not planning backward from the first-in-man research goal can be….

39. Consequence of Poor Planning

40. And then..

41. SAIC’s Virtual Pharmaceutical and Product Development Support Service Capabilities Project Planning Assay Development Process Development GMP Manufacturing • Expression Platform Evaluation • CMC Characterization • Product Development Plans • Facility Audits • Assay Design, Development, and Validation • Stability-Indicating Assay Design, Development, and Qualification • Upstream Manufacturing Optimization • Downstream Processing and Purification Development • Pre-Formulation Studies • Release, Analytics, and Stability Assays • Technology Transfer • Facility Audit • Characterization of Final Drug Product • Product Stability Program • Finish and Fill • cGMP Drug Product, Vialing, and Labeling Repository Your Product Candidate at SAIC RegulatorySupport Tox and Safety Testing • Inventory Storage of Biological Materials • Shipment of Samples and Permits • All Tasks Under One Umbrella • Cost-Effective • No IP Issue • IT Infrastructure and Data Security • Complete Services from Bench to Bedside • Pre-IND Meeting • IND Preparation and Submission to FDA • Stabiltiy Monitoring During Clinical Life Cycle • In-House Quality Assurance and Oversight • Mouse Immunogenicity Dose Ranging and Potency • GLP Rabbit Toxicity • General Safety

42. Thank You • Opinions expressed in this presentation are mine and do not represent those of SAIC, NIAID, FDA, or other regulatory bodies. • Thanks to the organizers.