Transcript Gene Therapy Workshop

Gene Therapy Production
Facility Considerations
Robert Sausville
Center for Biologics Evaluation and Research
Office of Compliance and
Biologics Quality 7/16/2015
Division of Case Management
 Overview
 General
Considerations
 Special Considerations
 Biosafety Levels
 Air Quality
 Environmental Monitoring
 Validation
 Qualification
 Miscellaneous
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

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

General Considerations cont.

As the risk for aerosol transmission increases,
higher levels of primary containment and
multiple secondary barriers may become
necessary
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

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
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)

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

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

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
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
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

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

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
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
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

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

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

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

Settling Plates Exposure times

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
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.”
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
“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
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
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
Process Validation
Sterilization
 Decontamination
 Aseptic Processing
 Cleaning
 Inactivation/removal of adventitious agents and
other contaminants

Equipment Qualification
Program in place to demonstrate that
equipment operates as expected
 Should include periodic monitoring

Equipment cont.
Air handlers
 Biological safety cabinets
 pressures
 filter integrity
 airflows; velocities
 leak testing

Equipment cont.
Incubators
 uniform temperature
 carbon dioxide
 filters
 Centrifuges
 speed
 vacuum (ultras)
 temperature

Equipment cont.
Autoclaves
 temperature
 pressure
 kill cycle
 Lyophilizers
 shelf temperature
 vacuum

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

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

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

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
