The impacts of residence time and bed height on virus and aggregate clearance with a multimodal chromatography resin.

Process intensification has become a priority for the biopharmaceutical industry. Mixed-mode polishing chromatography holds the potential to streamline mAb purification through clearance of key product and process related impurities in a single step. Additionally, by operating in flow-through mode, increased capacities can be achieved, and the unit operation fits more conveniently into a continuous process. This study presents a unique, systematic exploration of the impacts of residence time versus bed height on viral clearance for a single-step, flowthrough polishing process, alongside additional development ensuring sufficient product recovery and impurity clearance. The mixed mode chromatography resin, Capto™ adhere, along with high titer, high quality feeds and optimized upstream processes provided through collaboration with Celltheon Corporation are leveraged to demonstrate a process development strategy for mAb polishing in flow-through mode to achieve reduction of aggregate, host cell proteins (HCP), and adventitious virus. A two-stage process development strategy was followed. First, a design of experiments investigating the impact of pH and conductivity on product recovery and aggregate removal enabled identification of a design space to remove the majority of aggregates. To further optimize the process and explore a single step polishing process, a second round of experiments introduced residence time and bed height as additional factors and mock virus removal as an additional response. These factors, which are not traditionally studied systematically in early process development, informed process setpoints where pool aggregate levels were brought to the target of 1% and >4 log reduction values (LRVs) of mock virus clearance were achieved. Contour and breakthrough plots indicate that residence time and bed height are significant for aggregate and mock virus removal, and furthermore, have fundamentally differing impacts on the separation of these two impurities.