Viral clearance studies …. are they needed for proteins produced using bacterial or yeast fermentation processes?

Dr. Ray Nims

In E. coli or Pichia pastoris-based bioproduction of recombinant proteins, there are no suitable host cells for amplification of viruses that are infectious for humans. Bacteria such as E. coli and yeast such as P. pastoris can only support the growth of certain bacteriophage or yeast viruses, respectively. These types of viruses are not infectious for humans or animals.

The potential for viral infection of a bacterial cell process or a yeast cell process used for production of a recombinant protein is therefore a business risk, not a patient safety risk. This is the reason why case studies of bacteriophage infection of bacterial fermentation processes or of viral contamination of yeast cell processes have not appeared in the literature (or in the news). Unlike virus contamination of animal cell-based production processes, the events involving bacterial or yeast do not have patient safety implications. When a bacteriophage or yeast virus contamination occurs, manufacturers quietly go about the business of restarting the fermentation. Remediation and prevention of future occurrences is driven primarily by business concerns, as opposed to regulatory concerns.

Manufacturers of recombinant proteins produced using bacterial or yeast cell substrates are not required to conduct cell line viral testing (though phage induction studies may be performed as part of bacterial cell line characterization – to mitigate business risk!), nor are they required to conduct lot-by-lot viral testing of bulk harvest samples. Such requirements are mandated for production processes using animal or human cells. As the title of the document “Quality of Biotechnological Products: Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin” indicates, the ICH Q5A (R1) guidance mandating such testing is clearly directed toward manufacturing processes utilizing the types of cells which may support growth of viruses infectious for human or animal cells. This ICH document also provides guidance on the conduct of viral clearance studies.

Viral clearance studies are intended to provide evidence of the ability of downstream purification steps of a manufacturing process to remove or inactivate viruses that potentially may contaminate the process, make their way into final product, and represent a health risk to patients. As such, the evaluation typically involves both relevant viruses (i.e., viruses that are known to contaminate these kinds of processes) and model viruses (i.e., viruses representative of the types of viruses that could contaminate these processes). In the case of bacteria- and yeast-based manufacturing processes, the relevant viruses (bacteriophage and yeast viruses, respectively) are not infectious for humans or animals. In addition, there are no model viruses that are capable of infecting these kinds of cells that are of concern for humans or animals. Therefore, viral clearance studies are typically not required or conducted.

Are there any circumstances where a bacterial or yeast fermentation process could be expected to harbor a virus capable of infecting humans? The only that I can imagine is a process utilizing primary or secondary animal-derived raw materials in rather large quantities. The worry would not be that viral amplification might occur, but that some carryover of surviving animal viruses to the final product might be possible. The use of such animal-derived materials by a manufacturer of a therapeutic protein must be justified based on risk analysis. If substantial risk is introduced by the use of such raw materials, then perhaps a raw material treatment approach would need to be validated.

In the absence of the use of animal derived materials or plant materials not subject to processing steps that would inactivate contaminating animal viruses, incorporation and validation of viral clearance steps into protein production processes using bacteria or yeast cell substrates is not expected, nor would this be of practical value in assuring patient safety.