Gene Therapy Production Facility Considerations

1. Gene Therapy Production Facility Considerations Robert Sausville Center for Biologics Evaluation and Research Office of Compliance and Biologics Quality Division of Case Management

2. Overview • General Considerations • Special Considerations • Biosafety Levels • Air Quality • Environmental Monitoring • Validation • Qualification • Miscellaneous

3. Overview • New manufacturing areas • retrofits of existing facilities • located within existing facilities • Designed to minimize potential for contamination • between products • between early and late stages of production • Operations should be well controlled • Personnel should be appropriately trained • someone responsible for facility operations, equipment validation and maintenance, record keeping

4. General Considerations • Good techniques and appropriate equipment used to minimize exposure to infectious agents • Strict adherence to standard practices and techniques (consistent manufacture of product) • Safety equipment (primary barrier) • biological safety cabinet • closed containers • other designs to minimize aerosols • Facility design (secondary barrier) • amount of protection depends on the nature of the infectious agent and production associated risks (aerosols)

5. General Considerations cont. • As the risk for aerosol transmission increases, higher levels of primary containment and multiple secondary barriers may become necessary

6. General Considerations cont. • Facility designed for aseptic processing • Smooth surfaces, seamless tile, etc. • Hard, easily cleanable and impervious finishes • Non shedding ceilings • HEPA filtered air from seperate air handling unit • separation of production from other areas of the facility (often a hospital) • Phase 1/II • Class 100,000: general manufacturing areas (10,000 III) • Class 100: all open manipulations

7. General Considerations cont. • Separate entrance/gowning area • Air locks (containment) • No personnel access between vector production and transduction areas • Room(s) for support areas • buffer media prep, glass/equipment wash • Storage areas • raw materials, cell banks • adjacent to production if possible to minimize traffic in and out of the facility

8. General Considerations cont. • Material and personnel flows designed to maximize efficiency and minimize mix-ups • unidirectional flows where possible • also controlled by procedures or temporally • Emergency power for critical systems (UPS)

9. General Considerations; Control of the Facility • Production area(s) should be separate from other activities (research, testing) • Potential cross-contamination should be minimized • between process steps, and • other production activities • Access into the production area should be limited • Equipment used in production should not be shared with non-production activities • Environmental monitoring and product testing should be performed to ensure adequate control of the process/area • Spill/accident procedures should be in place

10. Special Considerations; Multi-use scenarios • Concurrent vs. campaign production will impact design considerations • Personnel work on one cell line at a time • Procedures in place to prevent cross contamination • Appropriate changeover procedures • Adequate segregation of concurrent activities • color coded labeling • bar coded • The sponsor is responsible for assuring that the contract manufacturer has all the procedures in place

11. Special Considerations Cont. • Potential Routes of Cross Contamination • Centrifuges (generation of aerosols) • one sample processed at a time • cleaning/sanitization of centrifuges between “lots” • Pipettors • effective cleaning procedures • filters • Incubators

12. Appropriate BioSafety Level • In general, BioSafety Level 2 • Transduction • Higher risk, BioSafety Level 3 • Vector Preparation • Defined in CDC-NIH publication Biosafety in Microbiological and Biomedical Laboratories

13. BioSafety Level 2 • Transduction and non-viral vector preparation • access limited • personnel trained in handling pathogenic agents • infectious wastes are decontaminated before disposal • gowning required (lab coat, hair cover) • gloves should also be worn for aseptic manipulations • Class I or II Biosafety Cabinets to be used: • for procedures potentially creating aerosols • aerosol generation should be minimized • with high concentrations or large volumes of infectious agents

14. BioSafety Level 3 • Viral Vector prep areas, higher levels of containment • Negative pressure or “sink” for containment • All activities with infectious materials are conducted in biological safety cabinets • Class I, II or III may be used • Passage between two sets of doors is a basic requirement • An autoclave for decontaminating waste is available • preferably in the laboratory • Ducted exhaust provided • not recirculated • may be discharged to the outside without being filtered

15. BioSafety Level 4 • Class III biosafety cabinets • or personnel in suits with life support systems • All materials must be autoclaved before leaving BSL4 area • Exhaust air HEPA filtered

16. Air Quality • Recommend that production areas receive single pass air (no recirculation) for vector production • Dedicated air handler where possible • Segregating air supply from rest of facility important, (hospital setting) • HEPA-filtered air • Objective: to meet Class 100,000 specifications for Phase I/II • In-line vs. terminal

17. Air Quality cont. • Air pressure differentials between areas should be balanced to maintain cleanliness or containment • Positive (aseptic processing) • Negative (containment) for steps needing greater than BSL-2 • Open steps in biosafety cabinets (Class II) • Quality monitored to assure facility is acceptable for production

18. Environmental Monitoring • Testing of surfaces and for viable and non-viable particulates • Demonstrate facility under control • as part of validation • routine monitoring program • Frequency and intensity dependent on how close to GMPs the facility will operate • specifications based on desired Class

19. Environmental Monitoring cont. • Viable particulates • active air monitoring devices (slit to agar, centrifugal samplers) • settling plates (passive, less desirable) • Non-viable particulates • particle counters • room classifications, certification of biosafety cabinets • Surface “contact” plates or swabs • monitor cleaning efficacy and personnel asepsis

20. NASA Standards; Viable Air Particulates • Room classification, defined by Federal Standard 209E, determined by non-viable particulate monitoring under dynamic conditions. • Class 100,000 • 2.5 CFU/ ft3 • Class 10,000 • 0.5 CFU/ ft3 • Class 100 • 0.1 CFU/ ft3

21. Class 100,000 1 CFU/ 9cm plate 0.11 hours 2 CFU/ 9cm plate 0.21 hours 1 CFU/ 14cm plate 0.04 hours 2 CFU/ 14 cm plate 0.09 hours Class 100 1 CFU/ 9cm plate 2.65 hours 2 CFU/ 9 cm plate 5.31 hours 1 CFU/ 14 cm plate 1.10 hours 2 CFU/ 14 cm plate 2.21 hours Settling Plates Exposure times

22. Validation • “Establishing documented evidence which provides a high degree of assurance that a specific process will consistently produce a product meeting its predetermined specifications and quality attributes.”

23. Validation • “Sliding Scale” approach for clinical manufacturing • Facilities involved in clinical manufacturing should be in compliance with the concepts of cGMPs • Do not expect full validation in early stages (may not have 3 repetitive runs, worst case configuration, etc.) • facility supplying clinical material to other institutions implies that it meets cGMPs and should approach what is expected for commercial facilities • Phase 3 material should be manufactured at close to full cGMP

24. “Sliding Scale” Approach, An Example • Autoclave used to prepare sterile materials • Early (I/II) • demonstrate proper cycle achieved • monitor temperature, pressure, and time • use of biological indicators for verification • loads not well defined

25. Autoclave cont. • Middle (II/III) • temperature mapping done to determine cold/hot spots • biological indicators placed to verify cycle at problems points • loads are somewhat defined

26. Autoclave cont. • Late (III+) • lethality of cycle determined at monitored points • loads are well defined and standardized • each load configuration has been mapped or worst-case load has been validated • Another example • Sanitizer effectiveness • Phase I/II supported by literature • Phase III supported by validation

27. Process Validation • Sterilization • Decontamination • Aseptic Processing • Cleaning • Inactivation/removal of adventitious agents and other contaminants

28. Equipment Qualification • Program in place to demonstrate that equipment operates as expected • Should include periodic monitoring

29. Equipment cont. • Air handlers • Biological safety cabinets • pressures • filter integrity • airflows; velocities • leak testing

30. Equipment cont. • Incubators • uniform temperature • carbon dioxide • filters • Centrifuges • speed • vacuum (ultras) • temperature

31. Equipment cont. • Autoclaves • temperature • pressure • kill cycle • Lyophilizers • shelf temperature • vacuum

32. Raw Materials • Critical raw materials - established criteria for acceptance from vendors: • sterility • adventitious agents • activity/purity • Vendor’s Certificate of Analysis • identity test where possible • Inventory Control • proper storage • FIFO

33. Water • Should meet Water for Injection (WFI) specifications : • microbial <10 CFU/100ml • endotoxin <0.25 EU/ml • chemical tests per USP • WFI for all product contact surfaces and formulations • May be purchased

34. Personnel • Designated person in charge of facility • Responsible for: • limiting access • training • maintenance/safe operations • writing and enforcing procedures • Production personnel are trained (periodic retraining) • Appropriately gowned for production step

35. Final Thoughts • Design facility for worst-case = maximum flexibility • Consider filing a Master File for facilities handling several IND products • Meet with FDA to discuss your Phase I/II (or III!) facility