Validation of Virus Removal Keith O.Webber, Ph.D. Deputy Director, Div. of Monoclonal Antibodies OTRR/CBER/FDA PDA/FDA Meeting September 26, 2000

SCOPE - Products Specified Biologics q Monoclonal Antibodies q rDNA-derived products Traditional Biologics q Some Blood-products q Some Vaccines

SCOPE - Methods Column Chromatography Nano-Filtration (Viral Filtration) Other Methods Precipitation Centrifugation

Example Production Fermentor Protein A affinity chromatography Anion Exchange Chromatography Virus filtration Gel Filtration Formulation and Fill

GENERALITIES q Validation conditions must be representative of the actual manufacturing process. q Validation should use virus-spiking studies. q Viral clearance studies should not be done in the production facility.  Validations should be done in duplicate.

SCALE-DOWN q Scale Down: »Allows validation to be performed in testing labs »Maintains high titers of the spiking virus »Must be done appropriately

CHROMATOGRAPHY Parameters that should be representative of commercial-scale manufacturing: –CHROMATOGRAPHY MEDIUM –COLUMN BED HEIGHT –LINEAR FLOW RATE –BUFFER COMPONENTS AND CONCENTRATIONS –pH –TEMPERATURE –PRODUCT LOAD

CHROMATOGRAPHY and… the chromatographic profile and product recovery of the scale-down process should be comparable to that of the manufacturing process. the chromatographic profile and product recovery of the scale-down process should be comparable to that of the manufacturing process.

FILTRATION Parameters that should be representative of commercial-scale manufacturing: – VISCOSITY – VOLUME PER cm 2 OF FILTER AREA – IONIC STRENGTH – TEMPERATURE – pH – PROTEIN COMPOSITION & CONCENTRATION

FILTRATION and… the product recovery of the scale-down process should be comparable to that of the manufacturing process.

CHOOSING VIRUSES The aims of viral validation studies are: »to provide evidence that the production process will effectively remove viruses which are either known to contaminate the starting materials, or which could conceivably do so, and »to provide indirect evidence that the production process might remove novel or unpredictable virus contamination.

CHOOSING VIRUSES The clearance validation should include: »relevant viruses that may be anticipated to occur in the system »specific model viruses that are physically and chemically similar to relevant viruses »non-specific model viruses that represent the extremes of virus properties

CHOOSING VIRUSES Examples of model viruses: Retrovirus: X-MuLV Small non-enveloped virus: SV40, parvovirus, polio virus 1 Medium to large enveloped RNA virus: Parainfluenza, Sindbis virus Medium to large DNA virus: HSV-1, pseudorabies virus

PRECAUTIONS q Avoid aggregation of spiking virus q Use small volumes of spiking virus in order to retain the sample composition. q Assess the potential for assay interference by the product or buffers. q Use control assays in parallel to assess the loss of infectivity due to dilution, concentration, filtration, or storage. q Include controls to demonstrate the effect of procedures used solely to prepare the sample for assay.

General Procedure Add high Titer Spike to Sample Load Titer Sample LoadTiter Sample Load Take Hold Control sampleTake Hold Control sample Titer Hold ControlTiter Hold Control Titer Column FlowthroughTiter Column Flowthrough Titer Eluate SampleTiter Eluate Sample

Virus Assay Methodology q Infectivity is the standard method for clearance studies. q PCR assays and Polymerase Enhanced Reverse Transcriptase assays are being developed.

Evaluation of Results q Clearance factors from sequential orthogonal processes may be combined to give a Cumulative Clearance Factor q Orthogonal processes are those that clear virus by independent modes of action (e.g., Anion Exchange chromatography and Cation-Exchange chromatography)

Evaluation of Results Beware of including both a low-pH treatment step and a chromatography step that uses elution at a low pH in the Cumulative Clearance Factor.

Evaluation of Results The Cumulative Clearance Factor must be substantially greater than the estimated number of virus particles in a volume of the starting material required to produce a human dose of the drug.

Evaluation of Results q The number of virions per mL of starting material should generally be estimated by Transmission Electron Microscopy. q One “dose” of product is generally considered to be the total regimen of drug. e.g., 1 mg/week x 4 weeks = 4 mg dose

Example Product Protein A affinity chromatography Anion Exchange Chromatography Virus filtration Gel Filtration Formulation and Fill Homologous Product

Generic and Modular Validation A generic validation study demonstrates virus removal or inactivation by a process using a model product and allows the use of that process with an homologous product without the need to revalidate.

Generic and Modular Validation Generic viral clearance validation is applicable to situations when the purification process of a product is the same as a process that has already been validated for an homologous product.

Chromatography The two processes must have the same: -chromatography medium, -column geometry, -equilibration buffers, -load composition & concentration, -elution buffers, -elution parameters, -wash procedure.

Virus Filtration The two processes must have the same: q type of virus filter q solution characteristics 4 VOLUME PER cm 2 OF FILTER AREA 4 IONIC STRENGTH 4 TEMPERATURE 4 pH 4 PROTEIN COMPOSITION & CONCENTRATION

Generic and Modular Validation Example of Homologous Products: Monoclonal Antibodies of the same species, class, and subclass and derived from the same source (e.g., ascites, tissue culture, etc.) and cell substrate.

Generic and Modular Validation NOTE: GENERIC VALIDATIONS ARE NOT APPLICABLE TO HUMAN-DERIVED PRODUCTS OR PRODUCTS THAT MAY BE CONTAMINATED WITH HUMAN PATHOGENS.

Post-Approval Changes Whenever a change is made in the production or purification process, the effect of that change on the viral clearance should be considered and the system re- validated as needed.

Additional Reading q ICH Guidance on Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin,1998 ( q Points to Consider in the Manufacture and Testing of Monoclonal Antibody Products for Human Use, 1997 ( q Guideline on General Principles of Process Validation, 1987 ( q Reviewer Guidance: Validation of Chromatographic Methods, 1994 (  Validation of the Purification Process for Viral Clearance Evaluation, 1997, A.J. Darling, in Biopharmaceutical Process Validation, G. Sofer and D.W. Zabriskie, editors. Marcel Dekker, Inc (New York)

Guidance q CBER Office of Therapeutics: q CBER Office of Vaccines: q CBER Office of Blood:

Obtaining Documents q Fax (888) CBER-FAX q Internet q q Mail Training and Manufacturer Assistance (HFM-40) CBER/FDA 1401 Rockville Pike Rockville, MD