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History of Viral Contamination of Biologics

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Evaluation of Viral Clearance
Studies
Mahmood Farshid, Ph.D.
Div. Of Hematology
OBRR/ CBER/FDA
Biologics
• Monoclonal antibodies and recombinant
products produced in cell culture
• Animal derived products
• Blood and blood products and other human
derived products
Risk Reduction Strategies
• Donor Screening:
– donor history assessment,
– written and oral questionnaire
• Donors Testing:
– Anti- HIV-1/2, HIV-1 p24 Ag ,anti-HCV, HBsAg , anti
HBc, anti-HTLV-1/2, syphilis
– (NAT for HCV and HIV)
• Pharmacovigilance/ look back studies
• Inactivation/Removal
– Validating the manufacturing processes for removal /
inactivation of viruses
The Aim of Viral Validation
• To provide evidence that the production
process will effectively inactivate/remove
viruses which could potentially be
transmitted by the product
• To provide indirect evidence that the
production process has the capacity to
inactivate/remove novel or yet
undetermined virus contamination
Virus Clearance Methods
Virus inactivation:
• Chemical: organic
solvents; pH extremes;
solvent/detergent;
alcohol
• Physical: Heat
treatment (dry heat or
pasteurization)
• Combined Methods:
Photochemical
Virus removal:
• Precipitation: ammonium
sulfate etc.
• Chromatography: ion
exchange; gel filtration;
affinity; reverse phase
• Membrane filtration:
Omega, Planova, DV50
Validation of Virus
Removal/inactivation
• Scaling down process steps
• Spiking appropriate steps with high titer of
infectious virus (relevant or model)
• Determining virus reduction factors for each
step
• Summing reduction factors to give a total
log10 reduction value (LRV)
Evaluation of Viral Clearance
Steps
• Test viruses used
• The design of the validation studies
– Validity of scaled-down process
– Kinetics of inactivation
– Robustness
– Assay sensitivity
• The log reduction
Virus Selection
• Viruses that can potentially be transmitted
by the product (relevant or specific model
viruses)
• Viruses with a wide range of
physicochemical properties to evaluate
robustness of the process (non-specific
model viruses)
Virus Selection
• The nature of starting material
– Cell lines
– Human derived
– Animal derived
• Feasibility
– Availability of a suitable culture system
– Availability of high-titer stocks
– Reliable methods for quantification
Model viruses for human Blood-Derived Products
Virus
Model
Envelope/
Genome
Size
(nm)
Resistance
HIV/HTLV
HIV-1
Yes / RNA
80-130
Low
HBV
DHBV
Yes / DNA
~ 40
Medium
HCV
BVDV
Yes / RNA
40-50
Medium
HAV
HAV
No / RNA
28-30
High
CMV
CMV/HSV
/PRV
Yes / DNA
150-200
Low-Med
B19
PPV
No / DNA
18-26
Very high
Viruses Used to Validate Product Derived from
Cell Lines
Virus
Genome
Size(nm)
Enveloped
Resistance
MVM
ss-DNA
18-26
No
Very high
Reo-3
ds-RNA
60-80
No
High
MuLV
ss-RNA
80-130
Yes
Low
PRV
ds-DNA
150-200
Yes
Low-med
Virus Selection
• DNA and
RNA genome (single and double-stranded)
• Lipid-enveloped and nonenveloped
• Large, intermediate, and small size
• From very highly resistant to inactivation to very
easily inactivated
Scale-Down of Purification Steps
• Usually 1/10 to 1/100 scale
• Must keep buffers, pH, protein
concentration, and product the same as full
scale manufacturing
• Must keep operation parameters as close to
full scale as possible
• Must show product is identical to
production scale
Important Factors for Validation of
Photochemical Processes
• Concentration of the chemical with changes in
donor plasma/cell volume
• Lipemia and other impurities in the donor unit
• The degree of impurity removal prior to treatment
• The total quantity (fluence) of light as well as its
intensity and wavelength
• Plastic bag transparency
• Sample depth
• Mixing efficiency
• Residual level of chemical and its breakdown
products
Criteria for An Effective Virus
Clearance Step
• Significant viral clearance
• Reproducible and controllable at process
scale and model-able at the laboratory scale
• Should have minimal impact on product
yield and activity
• Not generate neo-antigens or leave toxic
residues
Other Considerations
• Manufacturing processes for blood derived
products must contain two effective steps
for removal/inactivation of viruses
• At least one step should be effective
against non-enveloped viruses
• At least one stage in a production process
must inactivate rather than remove viruses
Limitations of Viral Validation Studies
• Laboratory strains may behave differently
than native viruses
• There may exist in any virus population a
fraction that is resistant to inactivation
• Scale-down processes may be differ from
full-scale
• Source plasma or Igs may have neutralizing
antibodies
Limitations of Viral Validation Studies
• Total virus reduction may be overestimated
because of repeated and similar process
steps
• The ability of steps to remove virus after
repeated use may vary
How Much Clearance?
• The total viral reduction should be greater
than the maximum possible virus titer that
could potentially occurs in the source
material
• A manufacturing process must be validated
to remove/inactivate three to five orders of
magnitude more virus than is estimated to
be present in the starting materials
Factors influencing TSE
clearance
• Selection of TSE agent strain
– CJD, vCJD or GSS
• Infectivity assay
– Animal species
– Genotype
– Period observed
• Spiking preparation
– Crude brain homogenate
– Microsomal preparation
– Bolton preparation
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