(336e) Host Cell Protein Removal Via Flow-through Chromatography: A Proteomics-Based Approach

Manufacturing costs represent a major barrier limiting the access to biotherapeutics by the global patient population. As purification processes particularly contribute to biomanufacturing costs, the development of affordable and more efficient purification technologies is a topic of growing interest. Platform processes in current downstream processing rely heavily on â??bind-and-eluteâ? chromatography, an inherently batch process that requires long processing times, significant buffer storage capacity, and often expensive chromatography resins. As an alternative to this methodology, we aim to develop a new purification paradigm for biotherapeutics hinged on a â??truly continuousâ? process comprising only flow-through or continuous mode steps. In applying this strategy, removal of host cell protein is a primary concern due to the wide variety in sequence and structure. In this work, we explored methods for removing host cell protein (HCP) by chromatography in flow-through mode. Commercially available chromatography resins commonly used in flow-through mode for polishing applications were evaluated over a range of different loads of HCP obtained from CHO-S culture harvest to (1) determine order of magnitude of protein removal and (2) identify and characterize what protein species are removed with each technology. A high-throughput protein detection process by mass spectrometry enabled us to identify and quantify the HCPs in both cell supernatant and process fluids. Our results indicate that commercially available technologies are not sufficient for removing extracellular HCP when operating strictly in flow-through mode, as the maximum log reduction observed under the conditions tested was ~1 LRV. HCP clearance was then evaluated over an immobilized combinatorial peptide library to determine the expected binding capacity range using affinity interactions of the library with the HCP mixture, a promising strategy for capture of host cell proteins.