Biomanufacturing is tasked with producing highly pure therapeutics by ensuring the removal of process and product-related impurities, such as host cell proteins (HCP), endotoxins, viruses etc. Purification of monoclonal antibodies (mAbs), which now form ~40% of the biotherapeutic market, relies on Protein A-based adsorbents for product capture in bind-and-elute mode. However, as an inherently batch process, Protein A-based chromatography imposes high costs, large footprints, long production times, and product degradation due to harsh elution conditions. Furthermore, with the increasing breadth of biologics formats and lack of suitable affinity ligands, complexity in purification keeps increasing. Literature shows that commercial adsorbents may not remove some “high-risk” HCPs that may be immunogenic and also decrease product shelf-life, causing financial impact and delays for patient access. In response, we demonstrate a novel affinity purification adsorbent created using an ensemble of peptide ligands – LigaGuard™, that efficiently captures the spectrum of HCPs in flow-through mode, enabling truly continuous, “Flow-through Affinity” purification (FTAC), directly from clarified cell culture fluids (CCFs). We utilized FTAC coupled with proteomics to profile HCPs from various expressions systems (CHO and Pichia pastoris) to develop purification strategies. We validated the adsorbent on various industrial CHO CCFs with different mAb and HCP titers across different process conditions. Resultant product fractions were characterized for yield, HCP LRV and to identify specific “high-risk HCPs”. Our results demonstrate 95% recovery, 98% purity of mAb product, and an HCP LRV up to ~ 2 under optimized conditions. Notably, LigaGuard™ affords the removal of “high-risk” HCPs such as cathepsins, serine proteases, PLBL-2 etc. This work describes the potential of LigaGuard™ as an HCP-scrubbing adsorbent prior to Protein A chromatography, and the potential to enable a Protein A-free, truly continuous mAb purification process with highly pure product recovery.

Development of “flow-through affinity chromatography” of biotherapeutics by selective HCP capture using ensembles of peptide ligands

ACS Spring 2022

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.