Trish Greenhalgh discusses: “Chromatography for virus clearance”
What we are seeing as we work with customers is a drive to simplify and compress processes as much as possible. Perhaps five years ago, there was a protein A column, low pH step, cation exchange (CEX), anion exchange and a virus filtration step. What we see now is customers who don’t include, or don’t want to include the cation exchange step or they may be doing direct capture with the CEX resin or a mixed-mode resin rather than a traditional protein A. There is a definite shift to reduce the number of steps, which basically means that the expectations for the virus validation are that you have to have to demonstrate greater clearance across each unit operation from a reduced number of steps than you might have had to before. Instead of validating maybe four steps or five steps in a process, some people are going into a validation study in the hope of getting enough clearance across as few as three steps with the expectation that the chromatographic media are going to deliver greater log reduction at each step. This is particularly challenging for parvoviruses or non-enveloped viruses especially which tend to have less predictable clearance across protein A or CEX. In Europe, the regulatory guidelines indicate that manufacturing processes for biologics should include two orthogonal steps for virus removal, each capable of reducing virus by at least 4 logs which is the measure of ‘effective removal’. It’s not as explicitly stated in the U.S. guidelines, but it’s assumed that there will be more than one effective virus removal step. The virus filter if generally considered an effective step for parvovirus removal, but if you reduce the number of chromatography steps it could be challenging to robustly deliver at least four logs of parvovirus clearance across one other chromatography step.
There is also a real need to understand the mechanism of action of virus removal for the different purification unit operations. Small differences in the molecule characteristics, buffers, impurity levels, and the preceding purification steps can all have a profound effect on the performance of a subsequent chromatography step and also the levels of virus reduction that can be achieved. As such, real process understanding is critical particularly as the FDA moves towards a more holistic approach of processes in terms of QbD; there is a far greater expectation on manufacturers today to really understand how small changes in running or manufacturing conditions impacts both the product quality and the safety testing in terms of virus removal.