Optimization, scale-up, and validation issues in Filtration of Biopharmaceuticals, Part 1 - - BioPharm International

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Optimization, scale-up, and validation issues in Filtration of Biopharmaceuticals, Part 1

VIRUS RETENTION Close examination of a filtration system's performance requirements, among other criteria, should determine selection. High removal efficiency (>5 to 6 LTR) has been documented with virus filters for numerous mammalian viruses >40 to 50 nm in size, including: vaccinia virus (250 nm x 300 to 450 nm), herpes simplex virus (120 to 300 nm), influenza virus (80 to 120 nm), murine leukemia virus (80 to 128 nm), HIV (80 to 100 nm), Sindbis virus (40 to 70 nm) and SV40 (40 to 55 nm). Filters with finer pores (nominally 15 to 20 nm) have demonstrated removal efficiencies >3 to 4 LTR for small viruses (18 to 30 nm) such as porcine parvovirus, human parvovirus 9, minute virus of mouse MVM (or minute mouse virus, MMV) and poliovirus. Table 5 shows typical viral clearance data in various protein solutions for virus filters nominally rated at 50 and 20 nm. Using two filters in series (double filtration) can further enhance clearance efficiency for small viruses.

PROTEIN TRANSMISSION Maximized protein recovery is a desired goal of any viral clearance strategy. Many viral retention filters are inherently hydrophilic or are hydrophilized through surface modifications to decrease protein binding and enhance product transmission. Transmission of over 95% of plasma-derived and monoclonal immunoglobulin G (IgG, ~ 160 kDa) has been reported through various virus filters, using concentrations up to 5%. Protein transmission effectiveness depends on the degree of protein aggregation, solution purity, and whether a protein gel layer forms on the membrane surface. As some product loss may be attributable to membrane adsorption or holdup within the filter assembly, flushing the filtration system with a buffer may enhance product recovery.

VALIDATION OF VIRUS FILTRATION The validation of filtration processes requires special attention, as both the filter manufacturer and the end user play a vital role in assuring specified performance. The filter manufacturer must initially qualify the membrane and filter construction, then ensure each filter will meet that specification. The filter user must demonstrate that each filter satisfies the process needs and their validated viral clearance claims.


Figure 3. Scale-up of the Depth Filtration Step Using Feed Containing 0% Solids
Viral filter retention validation studies provide two kinds of clearance. Virus-specific clearance is the direct evidence that the production process will effectively remove viruses that are either known to contaminate the starting materials or that can conceivably do so. General virus clearance is indirect evidence that the production process can remove potential novel or unpredictable viruses. Proper design of the validation study is critical to ensure its success. Usually, the retention study uses a scaled- down version of the full-scale process with small area membrane discs or modules. Some important factors in the study design include: choice of viruses; target reduction factor; virus titer; comparability of test feedstock to process feedstock with respect to concentration, temperature, chemistry, purity, and accuracy of scale-down model; volume-to-surface area ratio for the test and process filter; and inclusion of proper study controls.

FILTER PERFORMANCE PARAMETERS Several product and process parameters may affect microbial retention, including viral retention by filtration. Product parameters include pH, viscosity, surface tension, ionic strength, and osmolarity. Process parameters include batch size, temperature, time, pressure differential, and flux (flow rate per unit area). Details of these product and process parameters are outlined in PDA Technical Report No. 26, "Sterilizing Filtration of Liquids."4 Although this report focuses on retention of bacteria by "sterilizing grade" filters, many of the same considerations apply to viral retention by virus filters. When designing the validation protocol, it is important to address the effect of extreme processing factors on the filter capability; the filter validation should be conducted using worst-case conditions. In a virus filter retention validation study:

  • Maximum pressure differentials should be incorporated in model challenge conditions. A high-pressure differential might reduce viral retention. This should be the actual maximum differential across the test filter, not the total available system pressure. The pressure differential across the test filter during the validation challenge should meet or exceed the maximum pressure differential observed during processing.
  • Maximum flux (flow/area) should be used for the model challenge condition. High flux might reduce viral retention. It may not be possible to mimic pressure differential and flux simultaneously during validation studies. The user should determine which is more relevant to the specific process and develop a rationale to support the decision.
  • The longest potential processing time should be used for filter viral validation testing. Several factors related to process time may affect retention by membrane filters. These include filter compatibility, maintenance of integrity, any changes in bulk fluid during challenge, hold times, and time-dependent penetration.
  • The volume/area throughput should be at least as high as planned for the scaled-up process.
  • The lowest adsorption conditions should be used. If several versions of the same basic product are planned, virus adsorption might be affected by product composition. Product specific assays may be needed to test the lowest adsorption conditions. In general, formulations with higher ion strength and higher protein concentrations tend to reduce adsorption of viruses to membranes. However, in some cases, higher protein concentrations can enhance virus/protein interactions leading to virus aggregation.
  • The viral filter retention testing temperature should be close to the process temperature. Excessive high or low temperature variance might affect filter performance and, therefore, viral retention.


Table 5. Typical Viral Clearance with Virus Membrane Filters
It may not be justified to test each of these variables independently. A parametric approach considers possible interactions, testing a true worst-case condition. In addition to validating viral retention, the filter's capacity, compatibility, absorption, and extractables must also be validated. These concerns are comparable to validation of sterilizing filters, as outlined in PDA Technical Report No. 26, "Sterilizing Filtration of Liquids."4

FILTER INTEGRITY TESTING FDA's 1987 "Guideline on Sterile Drug Products Produced by Aseptic Processing" states, "After a filtration process is properly validated for a given product, process, and filter, it is important to assure that identical filter replacements (membrane or cartridge) used in production runs will perform in the same manner. One way of achieving this is to correlate filter performance data with filter integrity testing data."7


Figure 4. Virus Filter Integrity Test Correlation Data*
There are two types of physical integrity tests in use today: destructive and nondestructive. Destructive tests involve challenges with particulates (for example, particulate gold colloids) and are therefore limited to post-use application. In contrast, nondestructive tests based on liquid porosimetry or gas diffusion can be used both pre- and postfiltration. A nondestructive test recommended by several virus filter manufacturers is the "diffusive," or forward-flow test that quantitatively measures the diffusive flow of pressurized air or nitrogen (plus the bulk flow through any open or nonwetted pores) across a wetted membrane filter. Test sensitivity is enhanced with elevated gas pressure and reduced wetting liquid surface tension. Figure 4 shows data correlating virus retention to a forward flow diffusion limit value for a virus filter. Forward flow measurements lend themselves ideally to automation and can be performed readily with test instruments commonly used in the biopharmaceutical industry for similar integrity tests on 0.2 and 0.1 m rated "sterilizing grade" filters. Liquid porosimetry can also be conducted as a nondestructive test but requires cleaning after pre-use testing and before post-use testing and is less easily automated.

While there is common agreement that filters should be tested both before and after filtration, postfiltration integrity testing is often a regulatory requirement for product release. Because filter integrity tests must correlate with virus removal claims, performing such tests is a critical safeguard for biopharmaceutical manufacturers. In contrast to the product-specific viral clearance data generated by the user with scale-down filter models, the filter manufacturer develops conditions and data showing a correlation of virus retention by process-scale filter elements to a particular integrity test method and limit value. Often, this data is not considered by regulatory reviewers or inspectors, so users must understand the basis of a filter manufacturer's integrity test and its correlation claims, and they must determine if the test's sensitivity is adequate to detect even a marginal lack of filter integrity. Once installed in a process, the filter integrity test is the sole means of confirming validated virus clearance performance.

REFERENCES

1. A Wang, L Russell, T Tressel, A S Rathore, ACS Poster # 263, 227th ACS National Meeting, Anaheim, CA, 2004.

2. Millipore Application Note, Filter Sizing Methods- for Normal Flow Filtration Applications, Document # AN1512EN00.

3. D Uavorsky, S McGee, Selection and Sizing of Clarification Depth Filters, Genetic Engineering News, Vol. 22, No. 9, May 1, 2002.

4. Sterilizing Filtration of Liquids, Technical Report No. 26, PDA, 1998.

5. Note of Guidance on Virus Validation Studies: The design, contribution and interpretation of studies validating the inactivation and removal of viruses, CPMP EMEA, 1996.

6. Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin, ICH Harmonized Tripartite Guideline, 1997.

7. Guideline on Sterile Drug Products Produced by Aseptic Processing, FDA, 1987.


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