Validation of Viral Clearance
The rationale for a validation of virus removal
and inactivation is based upon possible hazardous side effects of potential
viral contamination of a biotechnically manufactured protein product derived
from mammalian cell culture, intended for use as a therapeutical drug
or an in vivo diagnostic reagent.
Protein biosynthesis in mammalian cells features glycosylated proteins of high molecular weight with no disulphide bridge limitations and secretion of the protein as a singular, native molecule, which is folded correctly to a complex tertiar structure.
However, a potential retrovirus or adventitious virus contamination might lead to concerns on product safety. Even if no virus infectivity or reverse transcriptase activity can be detected for the master cell bank (MCB) or extended cell bank (ECB), the presence of virus-like-particles (VLP's) can often be demonstrated by electron microscopy . The microscopic evaluation gives no answer on the biological relevance of suspicious particles, especially regarding their infectivity; a prominent example is the presence of high numbers of A-type particles in hybridoma cells, where infectivity is extremely low or not detectable at all. Despite this discrepancy between the number of virus-like particles and infectivity, it is the general recommendation to calculate the overall reduction factor based on the particle number. This might induce the implementation of additional expensive but possibly unnecessary process steps.
Nevertheless, the presence of a virus of unknown origin cannot be excluded: an unidentified virus might have unknown and potentially harmful physiological effects, and it is the unknown nature of the virus contaminant which complicates the development of a specific assay. Without a specific and sensitive assay it is impossible to monitor the presence as well as the removal or inactivation of the virus along the downstream process of the protein drug.
With regard to a potential contamination by an unknown virus a number of preventive measures have to be established around and implemented into the complex manufacturing process of the protein in order to exclude the presence of any viral particle in the drug substance.
Preventive measures at the level of cell biology include extensive testing of the producer cells for specific viruses and testing for adventitious virus at a number of stages during fermentation.
The potency and efficacy of unit operations for the removal or inactivation of virus has to be demonstrated for the entire downstream process by the individual validation to those process steps that are considered to contribute to virus safety. Such validation has to be performed according to the very same concepts and criteria established for process validation in general.
With rare exceptions such as microwave induced High Temperature Short Time (•HTST•) heat treatment using the UltraTherm® System the scale of the respective validation equipment might be different from the manufacturing scale: Typically the validation needs to be performed using equipment at least of identical type and scale as intended for the manufacture.
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©1999 Charm Bioengineering, Inc.