Analysis of filtration behavior using integrated column chromatography followed by virus filtration

Abstract

We evaluated filtration behavior and virus removal capability for a monoclonal antibodies (mAb) and plasma IgG under constant flow rate directly following flow‐through column chromatography in an integrated process. mAb solution with quantified host cell protein (HCP) content processed in flow‐through mode on in‐series mixed‐mode AEX and modified CEX columns connected to the Planova BioEX filter (pool‐less) achieved HCP logarithmic reduction value (LRV) of 2.3 and 93.9% protein recovery, demonstrating comparable or higher HCP LRV with high protein recovery compared to previous reports. For 5–15 mg/ml plasma IgG run to 100 L/m2, similar filtration behavior was achieved for flux of 10–100 LMH, and lower flux runs remained well below the maximum operating pressure, suggesting that higher throughput in continuous processing is achievable. Comparison of fit of plasma IgG and mAb filtration behavior to four blocking models showed little differences but slightly better fit to the cake filtration model. Viral clearance of the filtration step tested by in‐line spiking X‐MuLV or MVM into purified plasma IgG following the chromatography step showed robust removal at low flux. Integrating the Planova BioEX filter into continuous processes with column chromatography can achieve efficient downstream processing with reduced footprint and process time.

Viral clearance in end-to-end continuous process for mAb purification: Total flow-through integrated polishing on two columns connected to virus filtration

“…The total flow-through integrated polishing process has been reported to achieve high mAb purification and robust virus reduction (Shirataki et al, 2021a). In this report, we verify the effectiveness of this integrated process when incorporated into a practical scaled-down end-to-end continuous process.…”

“…Validation of the virus filter incorporated into a continuous process cannot be evaluated individually, unlike in a batch process; rather, it is necessary to consider an appropriate virus validation method for continuous process. Proposals for various inline virus spiking methods adapted for virus validation in a continuous process and confirmation of robust virus removal by these methods has been reported by Lute et al (2011), Bohonak et al (2021, Shirataki et al (2021a) and Malakian et al (2022).…”

“…After low pH inactivation, the pH-adjusted mAb solution is supplied to a total flowthrough two column polishing process including direct connection to a virus filter. The combination of the AEX and CEX columns used here with the virus filter has already been reported to show very high impurity removal, recovery rate and filterability of the virus filter in a total flow-through process (Shirataki et al, 2021a). Though activated carbon is reported to be effective for impurity and virus clearance by flow-through (Ichikawa et al, 2019;Kikuchi et al, 2022), we adopted flow-through two column chromatography in this study.…”

Development of a transient inline spiking system for evaluating virus clearance in continuous bioprocessing—Proof of concept for virus filtration

“…Bohonak et al (2021) used this inline spiking method to evaluate the performance of a continuous virus filtration step, with undetectable levels of XMuLV and MVM using the Viresolve® Pro virus filter operated at a low filtrate flux typical of what might be encountered in continuous bioprocessing. A similar approach was used by Shirataki et al (2021) to evaluate virus clearance for an integrated process consisting of a flow‐through ion exchange chromatography step directly coupled to a Planova™ BioEX virus filter. However, in both studies, the feed was continuously spiked with virus at a uniform level, while the virus clearance was evaluated from the virus titer in pooled samples obtained downstream of the virus filter.…”

“…For example, Lute et al (2020) showed that both the Planova 20N and BioEX hollow fiber virus removal filters provided very effective virus removal (>4-log clearance under all conditions) over 4 days of continuous operation. In contrast, David, Niklas et al (2019) A similar approach was used by Shirataki et al (2021) to evaluate virus clearance for an integrated process consisting of a flow-through ion exchange chromatography step directly coupled to a Planova™ BioEX virus filter. However, in both studies, the feed was continuously spiked with virus at a uniform level, while the virus clearance was evaluated from the virus titer in pooled samples obtained downstream of the virus filter.…”