Validation of adequate virus clearance is critical in order to obtain FDA approval for a manufacturing process. Clearance can be due to inactivation or removal from the process stream. During the purification, manufacturers of monoclonal antibodies must demonstrate reduction of 103 to 105 or more virus particles than is estimated in a single dose equivalent of the unprocessed bulk. Depending on the antibody titer, estimates of the number of virus particles in a single dose equivalent could be as high as 1010 to 1015 retrovirus-like particles per mL. In addition removal of adventitious virus using model parvovirus is required. Today unit operations such as virus filtration are frequently used to validate virus clearance. Virus filtration is conducted industrially in dead end mode. Membrane performance is shown to be affected by feed pH ionic strength and feed buffer components. In addition the presence of the product protein, especially small aggregates of the product (dimers, trimers etc.) have been found to have a significant effect on membrane performance. In addition, virus filtration experiments using model mammalian viruses are expenses, complicated to run and time consuming. The purpose of this work was to determine if model bacteriophages Phi X174 and PR772 can be used to rapidly determine the effect of operating conditions on membrane performance. Consequently parallel experiments have been conducted under the same conditions using FDA recommended model mammalian viruses Minute Virus of Mice(MVM) and Xenotropic Murine Leukemia Viruses(X-MLV). The clearance of two model bacteriophages as well as MVM and X-MLV was tested using 30, 100 and 300 kDa polyethersulfone(PES) membranes. Flux and virus clearance were measured through the dead-end filtration experiments. The presence and concentration of Bovine Serum Albumin(BSA) and Lysozyme on the virus removal were investigated. In addition, the effects of pH on virus removal and filtration flux were determined. Our results show that 100 kDa PES membrane performs the best with regard to filtration flux and virus clearance performance in the presence and absences of tested proteins. Feed concentration close to neutral leads to a higher flux while virus titer stays more or less the same compared to other pH conditions. Mechanisms and insights on the effects of proteins and pH on virus removal are obtained.

Effects of filtration condition on virus clearance

251st National Meeting (2016)

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