Host cell protein dynamics in the supernatant of a mAb producing CHO cell line

• Published in: Biotechnology and bioengineering Vol. 109; no. 4; pp. 971 - 982
• Main Authors: Tait, A.S., Hogwood, C.E.M., Smales, C.M., Bracewell, D.G.
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.04.2012; Wiley Subscription Services, Inc
• Subjects: Animals; Antibodies, Monoclonal - biosynthesis; Antibodies, Monoclonal - genetics; Batch Cell Culture Techniques - methods; Biomarkers; Bioreactors; Biotechnology; Cell culture; Cell Survival; Cells; Chinese hamster ovary cells; CHO Cells - cytology; CHO Cells - metabolism; Chromatography, High Pressure Liquid; Cricetinae; Cricetulus; Culture Media, Conditioned - analysis; downstream processing; Electrophoresis, Gel, Two-Dimensional; Glutamate-Ammonia Ligase - biosynthesis; Glutamate-Ammonia Ligase - genetics; host cell protein; Immunoglobulin G - biosynthesis; Immunoglobulin G - genetics; Isoelectric Focusing; mammalian cell culture; Monoclonal antibodies; monoclonal antibody; Particle Size; Principal components analysis; Proteins; Proteins - analysis; Proteins - metabolism; Proteomics; Recombinant Fusion Proteins - biosynthesis; Recombinant Fusion Proteins - isolation & purification; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Time Factors
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.24383

The characterization of host cell protein (HCP) content during the production of therapeutic recombinant proteins is an important aspect in the drug development process. Despite this, key components of the HCP profile and how this changes with processing has not been fully investigated. Here we have investigated the supernatant HCP profile at different times throughout culture of a null and model GS‐CHO monoclonal antibody producing mammalian cell line grown in fed‐batch mode. Using 2D‐PAGE and LC‐MS/MS we identify a number of intracellular proteins (e.g., protein disulfide isomerise; elongation factor 2; calreticulin) that show a significant change in abundance relative to the general increase in HCP concentration observed with progression of culture. Those HCPs that showed a significant change in abundance across the culture above the general increase were dependent on the cell line examined. Further, our data suggests that the majority of HCPs in the supernatant of the cell lines investigated here arise through lysis or breakage of cells, associated with loss in viability, and are not present due to the secretion of protein material from within the cell. SELDI‐TOF and principal components analysis were also investigated to enable rapid monitoring of changes in the HCP profile. SELDI‐TOF analysis showed the same trends in the HCP profile as observed by 2D‐PAGE analysis and highlighted biomarkers that could be used for process monitoring. These data further our understanding of the relationship between the HCP profile and cell viability and may ultimately enable a more directed development of purification strategies and the development of cell lines based upon their HCP profile. Biotechnol. Bioeng. 2012; 109:971–982. © 2011 Wiley Periodicals, Inc. The dynamics of the host cell protein (HCP) profile towards the end of fed‐batch culture are investigated and show how this is a dynamic process that changes with harvest time. The authors' data suggests that the majority of HCPs in the supernatant of the cell lines investigated here arise through lysis or breakage of cells, associated with loss in viability, and are not present due to the secretion of protein material from within the cell.