Identification and classification of host cell proteins during biopharmaceutical process development

Abstract

As significant improvements in volumetric antibody productivity have been achieved by advances in upstream processing over the last decade, and harvest material has become progressively more difficult to recover with these intensified upstream operations, the segregation of upstream and downstream processing has remained largely unchanged. By integrating upstream and downstream process development, product purification issues are given consideration during the optimization of upstream operating conditions, which mitigates the need for extensive and expensive clearance strategies downstream. To investigate the impact of cell culture duration on critical quality attributes, CHO-expressed IgG1 was cultivated in two 2 L bioreactors with samples taken on days 8, 10, 13, 15, and 17. The material was centrifuged, filtered and protein A purified on a 1 ml HiTrap column. Host cell protein (HCP) identification by mass spectrometry (MS) was applied to this system to provide insights into cellular behavior and HCP carryover during protein A purification. It was shown that as cultivation progressed from day 8 to 17 and antibody titer increased, product quality declined due to an increase in post-protein A HCPs (from 72 to 475 peptides detected by MS) and a decrease in product monomer percentage (from 98% to 95.5%). Additionally, the MS data revealed an increase in the abundance of several classes of post-protein A HCPs (e.g., stress response proteins and indicators of cell age), particularly on days 15 and 17 of culture, which were associated with significant increases in total overall HCP levels. This provides new insight into the specific types of HCPs that are retained during mAb purification and may be used to aid process development strategies.

Adaptive modeling optimized by the data fusion strategy: Real‐time dying cell percentage prediction using capacitance spectroscopy

“…Additionally, as the viability declines, cell lysis occurs, releasing various host cell proteins (HCPs) from the fragmented plasma membrane 5 . This poses challenges in product purification due to the potential risks related to immunogenicity and regulatory compliance 6–9 . Some residual HCPs, for example, reductase and lipase, may also deteriorate the antibody quality or degrade the formulation components 10–14 .…”

Development and validation of an analytical procedure for the determination of residual protein A in active substances of therapeutic monoclonal antibodies

“…Известно, что производственные примеси, присутствующие после очистки продукта (белки клетки-хозяина, липиды или ДНК клеток-продуцентов, белки контаминирующих агентов и др. ), могут индуцировать иммунный ответ или, выполняя роль адъювантов, стимулировать иммунные реакции на целевой рекомбинантный белок [3].…”