ABSTRACT
Viral clearance steps are essential for maintaining the safety and integrity of biopharmaceutical products. ICH Q5A mandates
that the manufacturing process remove or inactivate contaminants based on a process-specific virus clearance strategy. A 20-nm
retentive virus removal filter can clear both large and small viruses, but a virus spiking trial is needed to validate the
effectiveness of such a step. Virus retention studies were run with three lots of Virosart CPV, a 20-nm polyethersulfone virus
filter, over a flow decay range of 0 to 90%. The model virus used was bacteriophage PP7 using Pseudomonas aeruginosa as the target and indicator cell. Four different protein solutions were spiked with PP7 and tested in triplicate runs. The
retention goal of 4 log10 was met and exceeded over the entire flow decay profile.
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Innovative technologies in biopharmaceutical processing have resulted in greater production capabilities, and at the same
time, growing concern over the viral safety of the products. Virus contamination of products derived from human or animal
cell lines can have disastrous clinical consequences. Although there have been no reported infections or transmissions of
Chinese hamster ovary (CHO) cell-related type A and C virus particles to date, viral clearance steps are vital for ensuring
the safety and integrity of biopharmaceutical products.
The ICH Q5A regulatory guideline mandates that therapeutic biological product manufacturers implement technologies into their
manufacturing process that remove or inactivate known or unknown contaminants based on a process-specific virus clearance
strategy.1 Such viral clearance steps must be validated and must also demonstrate that the implemented technologies effectively remove
a range of known and unpredictable viruses.2 Due to these stringent regulatory demands, bioprocess companies have struggled to find innovative methods to effectively
eliminate and inactivate viruses in the bio-feedstream.
Virus filtration is an established and robust method for effectively reducing a range of viruses within a single stage of
the downstream purification process.2 As a part of the purification process of a biopharmaceutical, 20-nm retentive virus removal filters can clear both large
and small viruses. A virus spiking trial must be performed to validate the efficiency of these 20-nm filters.3 This article describes trials with a particular 20-nm filter and four different protein solutions.
PROPERTIES OF A VIRUS FILTER
The ideal virus filter should retain all viruses and allow high protein transmission while maintaining a high flow rate without
significant virus breakthrough. However, virus breakthrough seems to be a general phenomenon among current virus filters
and it has been suggested that pore plugging causes a decline in the flow rate: "With virus breakthrough a general phenomenon
for virus removal filters, one could argue that fouling by contaminants in virus spikes should be minimized so that conditions
in filter validation most closely represent those in the manufacturing process."4 Others have shown that the virus reduction capability of some virus removal filters decreases with increasing flux decay.5,6
Although contaminants and other various parameters may be main causes of filter breakdown, some nanofilters still efficiently
remove viruses at high Log Reduction Value (LRV) even when experiencing high flow decay. A recent paper outlined the various
titer reduction capabilities of virus retentive nanofilters.7 The data showed that not all filters tested for their LRV versus flow decay profile experienced a significant loss of titer
reduction with increasing flow decay.
The existing virus filtration technologies available on the market have similarities and differences with respect to the physical
parameters that affect small virus retention:
- pleated versus hollow fiber configuration
- pore size and pore size distribution
- pore density
- membrane symmetry
- membrane thickness
- number of membrane layers
- membrane material used.
Care should be taken with general statements about virus filter performance and LRV predictions. The filter configuration
is not the only factor that influences the overall virus reduction capability. A potential decrease in virus reduction may
depend on several factors, including protein concentration, buffer composition, product purity, and plugging mechanism (adsorptive
versus pore plugging). The influence of these parameters on the overall flow decay profile of each virus filter has to be
examined case by case.
Flow Decay Study Protocol
This article presents virus retention data for Virosart CPV, a 20-nm polyethersulfone (PESU) nanofilter from Sartorius AG
(Gottingen, Germany). Different protein types and concentrations have been used to determine LRV versus flux decay profiles.
Spiking studies were performed using a laboratory scale model of Virosart CPV (Virosart CPV Minisarts with an effective filter
area of 5cm2).
