The effect of feed quality due to clarification strategy on the design and performance of protein A periodic counter‐current chromatography

Abstract

The impact of two different quality feeds, derived using two different harvest clarification processes, on protein A periodic counter‐current chromatography (PCC) design and performance is investigated. Data from batch experiments were input into a model to design optimal PCC operating parameters specific to each feed material. The two clarification methods were: depth filtration using a wetlaid matrix which has Q‐functionality; and a combination of depth filtration and chromatographic clarification, using a Q‐functional nonwoven with a high anion exchange capacity (Emphaze™ AEX Hybrid Purifier) in which key impurities such as host cell DNA (HCDNA) and host cell proteins (HCP) are removed. The model predicted 34% better productivity for the chromatographically clarified cell culture fluid (CCCF) using a 4 column system, and productivity gains of 28% using only 3 columns enabling the option to simplify the protein A PCC strategy. Experimental validation of the predicted optimized PCC operating parameters using industrially relevant monoclonal antibody (mAb) CCCF feedstock over 100 cycles showed productivity gains of 49% for the chromatographically clarified material. HCP concentration was 11‐fold lower, and HCDNA concentration was reduced by 4.4 Log Reduction Value (LRV) in the protein A PCC eluates. This work, therefore, demonstrates that the removal of HCDNA and HCP during clarification is an effective strategy for improving protein A PCC performance. This was achieved using the Emphaze™ AEX Hybrid Purifier which can be easily incorporated into a batch or continuous process, in a scalable fashion, without adding additional separate unit operations. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:1380–1392, 2018

In situ neutron scattering of antibody adsorption during protein A chromatography

“…For protein A chromatography, a silica-based resin (Prosep Ultra Plus, Millipore, Hertfordshire, UK) was used. The model protein for this study was IgG1, protein A purified humanised IgG1 produced in CHO cell culture, as described in El-Sabbahy et al, [19]. It was dialysed into a running buffer (sodium phosphate, pH 7.2) with a final concentration of 1 mg/mL.…”

Chromatographic clarification overcomes chromatin‐mediated hitch‐hiking interactions on Protein A capture column

“…This approach can reduce DNA complexes in harvest CCF down to <100 ppb levels (Koehler, Jokondo, Narayan, Voloshin, & Castro‐Forero, 2019). Numerous previous studies have demonstrated strongly reduced HCP impurity levels in the Protein A elution pool, compared to traditional depth filters (Castro‐Forero, Jokondo, Voloshin, & Hester, 2015; El‐Sabbahy et al, 2018; Gilgunn et al, 2019; Singh et al, 2017). One of these studies also demonstrated removal of histone proteins to below the detection limit (Gilgunn et al, 2019).…”

Recent Developments in Bioprocessing of Recombinant Proteins: Expression Hosts and Process Development

“…The impact of two different quality feeds (one from depth filtration and other from a combination of depth filtration and chromatographic clarification) on Protein A PCC was studied, with the result that there was 49% increased productivity for the chromatographically clarified material over 100 cycles, with 11-fold lower HCP and a 4.4 LRV for HCDNA (El-Sabbahy et al, 2018). Upscaling of Protein A continuous chromatography using the Cadence TM BioSMB PD and the Cadence TM BioSMB Process 80 system was successfully carried out for a 10-day run time using feed from a perfusion culture and resulted in a 400-500% increase in vs. batch mode (Ötes et al, 2018).…”