Streamlining Biopharmaceutical Cell Line Development - Reducing risk and decreasing timelines through optimized reagents and processes
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The webinar by Merck KGaA discusses strategies for streamlining biopharmaceutical cell line development, focusing on optimizing processes, reducing timelines, and mitigating risks associated with cell line development. Key highlights include the benefits of using glutamine synthetase auxotroph cell lines to eliminate the need for amplification and drug selection, thereby shortening development times and lowering costs. The presentation also covers advanced genetic engineering techniques that enhance the production process and improve clone stability and consistency.
Streamlining Biopharmaceutical Cell Line Development - Reducing risk and decreasing timelines through optimized reagents and processes
- 1. Merck KGaA Darmstadt, Germany Reducing risk and decreasing timelines through optimized reagents and processes Streamlining Biopharmaceutical Cell Line Development Kate Achtien, Sr. Scientist Webinar 30th November 2017
- 2. 2 The life science business of Merck KGaA, Darmstadt, Germany operates as MilliporeSigma in the U.S. and Canada. Streamlining Biopharmaceutical Cell Line Development | Webinar
- 3. Webinar Goals Streamlining Biopharmaceutical Cell Line Development • Finding the needle in the haystack What are the goals and challenges of cell line development? • Foundation blocks Defining the components of a robust cell line development platform • Striking a balance Optimizing the cell line development process • The future of cell line development processes Advancing through genetic engineering Streamlining Biopharmaceutical Cell Line Development | Webinar3
- 4. Finding the needle in the haystack The Goals and Challenges of Cell Line Development Streamlining Biopharmaceutical Cell Line Development | Webinar4
- 5. Streamlining Biopharmaceutical Cell Line Development | Webinar Poll Question 5
- 6. Streamlining Biopharmaceutical Cell Line Development | Webinar What makes a robust biotherapeutic production process? Cell culture Harvest Clarification Chromatography Virus removal Tangential flow filtration Sterile filtration Formulation and Final Fill Cell line development Process EconomicsSpeed to Clinic Business Continuity 6
- 7. Fast development process Low clone screening efforts Screen molecules earlier in discovery process Maintain/enable favorable PQ attributes Scalable process Keep development costs low High titers and productivity Product homogeneity Culture longevity Clone Stability Exclusion of unnecessary additives Increased process robustness Concentrated feeds High growth rate and cell densities Reduce product and process impurities Control/reduce metabolic waste Clear technology IP path Regulatory compliance Reliable supply chain for paired media Reduce regulatory and risk considerations Streamlining Biopharmaceutical Cell Line Development | Webinar A robust cell line development process leads to strong benefits for development & production Process EconomicsSpeed to Clinic Business Continuity 7
- 8. Expression Vector Gene of Interest Transfection Cell line Cloning The cell line development challenge Finding the needle in the haystack Need for robust processes to Reduce Time, Resources and Cost Streamlining Biopharmaceutical Cell Line Development | Webinar8
- 9. Streamlining Biopharmaceutical Cell Line Development | Webinar Poll Question 9
- 10. Traditional methods Wild Type CHO Glutamate + Ammonia Glutamine Biological Function MSX Glutamine SynthetaseMethionine sulfoximine Folic acid Dihydrofolic acid Tetrahydrofolic acid Dihydrofolate Reductase Dihydrofolate Reductase Purine Metabolism X X X DG44 or DUXB11 (CHO) MTX MTX Wild Type CHO Neomycin Puromycin Selection achieved through addition of a selection agent Dihydrofolate Reductase based selection Glutmine Synthetase based selection Antibiotic based selection methods Streamlining Biopharmaceutical Cell Line Development | Webinar10
- 11. Weaknesses of additive-based selection methods Technical, economic and ecological challenges Requirement for amplification Significant clone screening efforts Increase of gene copy and risk for clone instability Time consuming process Application of antibiotics Regulatory review less smooth - clearance to be proven Disposal of hazardous drug for environmental compliance Absence of antibiotic-based selection pressure in the production process increases the risks for reduced productivity Longer timelines Increased resources needs Increased development costs Higher risk for loss of productivity Streamlining Biopharmaceutical Cell Line Development | Webinar11
- 12. Additive independent selection Wild Type CHO Glutamate + Ammonia Glutamine Biological Function MSX Glutamine SynthetaseMethionine sulfoximine Glutmine Synthetase based selection GS knock out cell line allows additive independent selection CHO GS-/- Glutamate + Ammonia Glutamine Biological Function Glutamine Synthetase Glutamine Synthetase KO based selection X Streamlining Biopharmaceutical Cell Line Development | Webinar12
- 13. Benefits of a glutamine synthetase auxotroph cell line GS-/ provides extensive benefits for development & production processes Fewer clones need to be evaluated No amplification needed Drug selection not required Stringent selection Streamlining Biopharmaceutical Cell Line Development | Webinar Shortened timelines Reduced cost and lowered regulatory hurdles Increased clone stability Lower resources required ✓ ✓ ✓ ✓ 13
- 14. Building a Foundation Defining the components of a robust cell line development platform Streamlining Biopharmaceutical Cell Line Development | Webinar14
- 15. Fast development process Low clone screening efforts Screen molecules earlier in discovery process Maintain/enable favorable PQ attributes Scalable process Keep development costs low High titers and productivity Product homogeneity Culture longevity Clone Stability Exclusion of unnecessary additives Increased process robustness Concentrated feeds High growth rate and cell densities Reduce product and process impurities Control/reduce metabolic waste Clear technology IP path Regulatory compliance Reliable supply chain for paired media Reduce regulatory and risk considerations A robust cell line development process leads to strong benefits for development & production Process EconomicsSpeed to Clinic Business Continuity Streamlining Biopharmaceutical Cell Line Development | Webinar15
- 16. Building a strong foundation for an efficient platform process Fast development process Low clone screening efforts Screen molecules earlier in discovery process Maintain/enable favorable PQ attributes Scalable process Keep development costs low High titers and productivity Product homogeneity Culture longevity Clone Stability Exclusion of unnecessary additives Increased process robustness Concentrated feeds High growth rate and cell densities Reduce product and process impurities Control/reduce metabolic waste Clear technology IP path Regulatory compliance Reliable supply chain for paired media Reduce regulatory and risk considerations Cell line design parameters Media design parameters Streamlining Biopharmaceutical Cell Line Development | Webinar Process EconomicsSpeed to Clinic Business Continuity 16
- 17. Streamlining Biopharmaceutical Cell Line Development | Webinar Step 1 Develop robust host Supension adapted • Adapted to fast and high suspension growth Strong origin Cells originated from ECACC CHO K1 Ease of use Optimized for cell line development, scale up and production processes Regulatory friendly • Grown in chemically defined media • Animal component free • cGMP banked • Full viral testing, complete traceability 2 1 43 17
- 18. Technical Benefits • Introduces site specific double strand breaks in the DNA • High specificity • Limited off target effects Regulatory Benefits • Clear IP path • Regulatory acceptance • Non-viral based technology Targeted Mutagenesis Step 2 Genetic engineering of host through regulatory friendly means Zinc Finger Nucleases Genetic engineering using Zinc Finger Nucleases (ZFNs)
- 19. Development of a paired media for efficient production processes Step 3 Business Continuity • Robust and sustainable supply chain • Proven manufacturability and consistency • Multisite manufacturing redundancy • Scalable GMP ready products • Animal component free Streamlining Biopharmaceutical Cell Line Development | Webinar19
- 20. Comprehensive expression platform & services To accelerate your development CHOZN® GS-/- cell line GS auxotroph cell line with clear IP path Traceability Documentation Comprehensive cell line history documentation to support regulatory filing Expression Vector IP free GS expression vector suitable for mAbs/ recombinants Process Guidance & Protocols Protocols for entire workflow Media and Feeds Fed Batch & Perfusion media produced under GMP Technical Support Comprehensive product and process technical support to ensure your success Streamlining Biopharmaceutical Cell Line Development | Webinar CHOZN® GS-/- Expression Platform 20
- 21. Striking a balance: Titer and Speed Developing a Robust Cell Line Streamlining Biopharmaceutical Cell Line Development | Webinar21
- 22. Striking a balance Development requires the right balance of clone performance and resources Speed Resources Equipment costs Throughput Clone performance Streamlining Biopharmaceutical Cell Line Development | Webinar22
- 23. Time investment to develop a stable pool reduces resources needed in single cell cloning stage Titer Heterogeneity Early screens for protein quality Proving “clonality” High throughput early screening High titers Scalable process Protein quality attributes Streamlining Biopharmaceutical Cell Line Development | Webinar Stable Pool Development Single Cell Clone Isolation Time intense Resource intense 23
- 24. Optimized cell line development process Balancing Resources and Performance Transfect Selection via “minipools” Screen minipools in 96-well plates Scale-up top producing minipools Screen 7-day TPP/shake flask Fed-batch assay on top minipools (TPP/shake) Single Cell Cloning 200 pools 100 pools 100 pools 20 pools 3-4 weeks 1 week 2 weeks 1 week 2 weeks 10-12 weeks Streamlining Biopharmaceutical Cell Line Development | Webinar24
- 25. CHOZN® GS-/- platform performance High expressing stable pools and clones 0 1 2 3 4 5 Stable Pools - Fed Batch Clones - Fed Batch 2 – 2.5 g/L 4 – 4.5 g/L CHOZN® Platform Benefits • High expressing pools can be quickly identified for early protein evaluation • Clones isolated typically produce at least twice the titer of the pool • Clones commonly produce 4- 5g/L prior to process development Titer Range of Top IgG Expressing GS-/- Pools and Clones mAbs (IgG1 and 4, IgM), Fc Fusions, Fabs, r-proteins, Bi-specifics Streamlining Biopharmaceutical Cell Line Development | Webinar25
- 26. CHOZN® GS-/- platform performance Performance of 4 CHOZN® GS-/- clones isolated from mini-pools 0 500 1000 1500 2000 2500 3000 3500 4000 4500 Productivity,Peak(mg/L) Peak Productivity 0 20 40 60 80 100 120 CellSpecificProductivity pg/Cell/Day Cell Specific Productivity (Qp) Streamlining Biopharmaceutical Cell Line Development | Webinar26
- 27. CHOZN® GS-/- platform stability Confidence in consistent production 75%75% High stability Studies performed on top ten producing clones show above industry standard stability. 80-90% of CHOZN® clones are found to have stable productivity. Streamlining Biopharmaceutical Cell Line Development | Webinar27
- 28. Streamlining Biopharmaceutical Cell Line Development | Webinar Process Scalability Consistent performance between vessel sizes 0 5 10 15 20 25 - 2 4 6 8 10 12 14 16 18 20 1e6Cells/ml Days Viable Cell Density 0 500 1000 1500 2000 2500 - 2 4 6 8 10 12 14 16 18 20 mg/L Days Titer Shake Flask EX-CELL® Advanced CHO Fed Batch No process development or optimization performed Mobius® 50L Bioreactor Mobius® 3L Single Use bioreactor 28
- 29. Streamlining Biopharmaceutical Cell Line Development | Webinar Process Predictability Glycan profiles similar across multiple scales Shake Flask Mobius® 3L Single Use bioreactor Mobius® 50L Bioreactor 29
- 30. The Future of Cell Line Development Processes Advancing Cell Line Development Through Genetic Engineering Streamlining Biopharmaceutical Cell Line Development | Webinar30
- 31. Cell Line Engineering by application of gene editing technologies Targeted Integration Reduce CLD timelines; increase consistency and stability Glycoengineering Removal of immunogenic and half-life reducing glycoforms Expression Platform Enhanced selection process, isolate high producing stable clones Manufacturing Security Reduce risk of viral contamination Robust Cell Line Suspension adapted to chemically defined medium CHO K1 CHOZN® GS-/- Protein Quality Mitigate Risk TI Streamlining Biopharmaceutical Cell Line Development | Webinar31
- 32. Streamlining Biopharmaceutical Cell Line Development | Webinar Poll Question 32
- 33. Targeted Integration Fast, Easy, Stable Reduced variability/enhanced cell line performance Clone to clone consistency Decreased characterization Decreased cell line development timelines More homogeneous stable pools More top clones in your pools Increased pool and clonal stability Use of well characterized safe harbor sites Remove stability from the critical path Aron M. Geurts, and Carol Moreno, Clin. Sci. 2010:119:303-311 GOI GOI Streamlining Biopharmaceutical Cell Line Development | Webinar33
- 34. Targeted Integration Ongoing Research 32 Increased Efficiency 1 Feasibility Hot Spot Identification Short RFLP donors GFP reporters IgG and Fc r-proteins Decreasing NHEJ Increasing HR Tagged ZFNs Optimized donor designs TI landing pads Transcriptomic analysis of constitutively expressed genes Reverse engineering of high expressing clones Random integration of a landing pad followed by characterization Streamlining Biopharmaceutical Cell Line Development | Webinar34
- 35. Cell culture Harvest Clarification Chromatography Virus removal Tangential flow filtration Sterile filtration Formulation and Final Fill Cell line development Process EconomicsSpeed to Clinic Business Continuity Streamlining Biopharmaceutical Cell Line Development | Webinar A robust cell line development process leads to strong benefits for development & production Finding the needle in the haystack The goals of cell line development Foundation blocks: Designing a robust platform Striking a balance Optimizing the cell line development process The future of cell line development processes Advancing through genetic engineering 35
- 36. Questions Streamlining Biopharmaceutical Cell Line Development | Webinar36
- 37. Kate Achtien Senior R+D Scientist 2909 Laclede Ave St. Louis, MO 63103 USA Phone: +1 314 289 8496 ext. 1939 Email: Kate.Achtien@sial.com CONtact: The vibrant M, CHOZN, Mobius, SAFC, MilliporeSigma and EX-CELL are trademarks of Merck KGaA, Darmstadt, Germany or its affiliates. All other trademarks are the property of their respective owners. Detailed information on trademarks is available via publicly accessible resources. © 2017 Merck KGaA, Darmstadt, Germany and/or its affiliates. All Rights Reserved.