Generation Bio — BPI East 2024
21 slides from Generation Bio's conference presentation, Sep 26, 2024
Slide 1: Rapid Enzymatic Synthesis and Scaling of DNA Gene Therapy Molecules
Rapid Enzymatic Synthesis and Scaling of DNA Gene Therapy Molecules BPI East 2024 Non-Viral Delivery Development and Manufacturing 26 September 2024
Slide 2: Disclosure Statements
Disclosure Statements • Anthony Dawson, Ph.D. o I am a current employee of Generation Bio Co. o I hold employee Incentive Stock Options (ISO) in Generation Bio Co. o I am listed as an inventor of patent applications covering the Technology disclosed o I have not received a separate speaking fee for
Slide 3: Forward Looking Statements
Forward Looking Statements 3
Slide 4: Today's Talk
TODAY’S TALK Limitations of traditional AAV manufacturing Approach to overcome common gene therapy quality and manufacturing limitations Generation Bio’s cell-free DNA manufacturing process: Rapid Enzymatic Synthesis (RES) Modularity RES enables modularity to enable preclinical speed and impart desi
Slide 5: Traditional AAV therapies rely on cellular DNA and capsid production
Traditional AAV therapies rely on cellular DNA and capsid production DNA payload and capsid delivery vector produced simultaneously intracellularly Cell-based (viral) manufacturing limitations • Long production times (weeks/months) • Low yields (1014 vg/L or 0.25 mg/L (DNA)) and high variability • R
Slide 6: How to address and overcome common gene therapy quality and manufacturing limitations
How to address and overcome common gene therapy quality and manufacturing limitations Viral gene therapy Non-viral gene therapy (cell-based manufacturing) Increase Process control (synthetic manufacturing) Sf9-produced drug substance RES produced drug substance Drug Sub- monomer Rapid substance Enzy
Slide 7: Developing a cell-free, enzymatic DNA production method to increase speed and quality
Developing a cell-free, enzymatic DNA production method to increase speed and quality Goal Approach Increase throughput, yield, quality and Develop an enzymatic process to scalability of DNA drug substance eliminate dependence on cellular manufacturing manufacturing Enhance: Process control Speed Sc
Slide 8: Considerations for designing an enzymatic process
Considerations for designing an enzymatic process Synthetic: Enable modularity to impart functionality Platform: Build based on standard inputs and constituents Tunability: Enzyme activity is more readily controlled than cell- based processes Scalability: Take advantage of highly processive nature o
Slide 9: Considerations for designing an enzymatic process
Considerations for designing an enzymatic process Synthetic: Offers Enable modularity to impart functionality construct flexibility and quality control Platform: Build based on standard inputs and constituents Tunability: Enzyme activity is more readily controlled than cell- based processes Scalabil
Slide 10: Rapid Enzymatic Synthesis (RES) offers drug substance flexibility
Rapid Enzymatic Synthesis (RES) offers drug substance flexibility + + Plasmid Amplified Plasmid Oligo Ends Functional elements 10
Slide 11: RES flexibility offers functional modifications not achievable in cell-based processes
RES flexibility offers functional modifications not achievable in cell-based processes Enabled by RES Transgene Structured Modified Positional Site-specific flexibility elements nucleotides modifications ligations BASE O O H H H H R O O P X O 11
Slide 12: Amplification during RES enables rapid scaling of input DNA materials
Amplification during RES enables rapid scaling of input DNA materials Step 1: Plasmid Amplification Amplification Reaction > 1000x µg → mg or mg → g Amplification reduces material lead times and allows scaling for candidate discovery and validation 12
Slide 13: RES-derived drug substance using amplified or non-amplified material displays comparable quality
RES-derived drug substance using amplified or non-amplified material displays comparable quality Step 2: Digest DNA to isolate GOI Step 3: Assemble and purify drug substance RES can be applied directly to plasmid or amplified plasmid Purified (drug substance) Plasmid | Amplified Plasmid | Amplified
Slide 14: Scalability and speed of RES allows iterative screening and platform manufacturing
Scalability and speed of RES allows iterative screening and platform manufacturing by the same process Iterative functional screening Platform manufacturing process Construct / 1 2 3 4 5 Design 5.0kb RES 3.0kb 1.5kb RES enables construct screening RES enables rapid scaling of • Transgene lead candid
Slide 15: Modularity of RES iqDNA synthesis enables end stabilization against exonucleases
Modularity of RES iqDNA synthesis process enables end stabilization against exonucleases 100 Modified iqDNA Unmodified iqDNA % of intact iqDNA 75 50 25 iqDNA + SVPD Agarose gel 0 densitometry exonuclease 0 30 60 90 120 Time (Min) 15
Slide 16: RES enables accelerated high-quality DNA production across scales
RES enables accelerated high-quality DNA production across scales Plasmid for throughput and screening Starting Scale (plasmid) microgram - milligram µg mg Plasmid receipt Processing Volumetric Process time time scale ~ 2 wk 3d 1 – 10 mL Sf9 Cell Based ~ 3 wk (bacmid) 3 - 4 wk 100 - 1000 mL Reduced
Slide 17: RES enables accelerated high-quality DNA production across scales
RES enables accelerated high-quality DNA production across scales mg g Amplification for rapid scaling Amplification Scale (utilizes plasmid template from starting scale) 100 milligrams 1 gram Plasmid Plasmid Processing Volumetric Processing Volumetric Process Amplification time scale Amplification
Slide 18: RES offers comparable drug substance quality across scales
RES offers comparable drug substance quality across scales Factor VIII 50 mL scale Factor VIII 2L Stirred-tank reactor 40x volume expansion uV 105000 monomer 100000 95000 monomer 90000 85000 80000 75000 70000 65000 60000 55000 50000 45000 40000 35000 30000 25000 20000 15000 10000 5000 0 9.0 10.0 11.
Slide 19: RES derived DNA provides expression enhancements over cell-derived DNA
RES derived DNA provides expression enhancements over cell-derived DNA. RES outperforms Sf9 Amplified material performs manufacturing equivalently to standard plasmid 2.0 mg/kg i.v. injection 2.0 mg/kg i.v. injection FVIII (LNP) Luciferase (LNP) FVIII Expression RES Luciferase Expression (Day 7) 1 1
Slide 20: RES demonstrated to be exceptional method for DNA gene therapy production
Rapid enzymatic synthesis (RES) demonstrated to be exceptional method for DNA gene therapy production Platform application • Standardized enzymes, DNA inputs, reaction conditions, and processing methods • Synthetic makeup provides valuable engineerability and modular functionality Heightened R&D pre
Slide 21: Acknowledgements
Acknowledgements Leadership Teams Geoff McDonough DS Process Sciences Matthew Stanton Analytical Development Antoinette Paone Molecular Biology Platform CMC Russell Monds Adam Villa Kelly Miller Daniel Blackstock Andrew O’Hagan John Durant Joris Cipi