Virus elimination during the purification of monoclonal antibodies by column chromatography and additional steps

Abstract

The theoretical potential for virus transmission by monoclonal antibody based therapeutic products has led to the inclusion of appropriate virus reduction steps. In this study, virus elimination by the chromatographic steps used during the purification process for two (IgG-1 & -3) monoclonal antibodies (MAbs) have been investigated. Both the Protein G (>7log) and ion-exchange (5 log) chromatography steps were very effective for eliminating both enveloped and non-enveloped viruses over the life-time of the chromatographic gel. However, the contribution made by the final gel filtration step was more limited, i.e., 3 log. Because these chromatographic columns were recycled between uses, the effectiveness of the column sanitization procedures (guanidinium chloride for protein G or NaOH for ion-exchange) were tested. By evaluating standard column runs immediately after each virus spiked run, it was possible to directly confirm that there was no cross contamination with virus between column runs (guanidinium chloride or NaOH). To further ensure the virus safety of the product, two specific virus elimination steps have also been included in the process. A solvent/detergent step based on 1% triton X-100 rapidly inactivating a range of enveloped viruses by >6 log inactivation within 1 min of a 60 min treatment time. Virus removal by virus filtration step was also confirmed to be effective for those viruses of about 50 nm or greater. In conclusion, the combination of these multiple steps ensures a high margin of virus safety for this purification process.

Protein A chromatography: Challenges and progress in the purification of monoclonal antibodies

“…After the chromatographic process, polishing steps such as ion exchange chromatography are usually used in the platform. Finally, virus removal is achieved by means of membrane steps (ultrafiltration, diafiltration and nanofiltration) , solvent/detergent treatments , IEC , etc.…”

A Review on Downstream Processing of Monoclonal Antibodies (mAbs)

“…[47]. Peter L. Roberts employed a solvent/detergent step based on 1% triton X-100 rapidly inactivates a variety of enveloped viruses by >6 log inactivation within 1 minute of a 60 minute treatment duration [48]According to research by Maria del Rosario et al, 20% ethanol inactivated 3.7 Logs of the enveloped viruses HSV-1 and HIV-1 but was unable to inactivate the nonenveloped viruses HPV-1 and CPV. However, 0.1N HCl is a powerful chemical agent capable of inactivating >6.13 Logs of high resistance nonenveloped viruses [49].…”

Polysaccharide-based chromatographic adsorbents for virus purification and viral clearance

“…Though widely used in antibody purification processes [212,218], protein G chromatography resins have been scarcely evaluated for viral clearance. An example is given by Roberts [226] who investigated virus elimination during the purification of immunoglubulins G1 and G3 produced from human hybridoma cell-lines. The purification process involved three sequential column chromatography steps using PS-based resins, i.e., protein G AFC (protein G Seph), CEC (SP Seph FF), and SEC (Superdex 200), completed by two specific virus elimination steps (S/D treatment and NF).…”