Probing of C-terminal lysine variation in a recombinant monoclonal antibody production using Chinese hamster ovary cells with chemically defined media.

Abstract:

:C-terminal lysine (C-K) variants are commonly observed in therapeutic monoclonal antibodies and recombinant proteins. Heterogeneity of C-K residues is believed to result from varying degree of proteolysis by endogenous carboxypeptidase(s) during cell culture production. The achievement of batch-to-batch culture performance and product quality reproducibility is a key cell culture development criterion. Understanding the operational parameters affecting C-K levels provides valuable insight into the cell culture process. A CHO cell line X expressing a recombinant antibody was selected as the model cell line due to the exhibited sensitivity of its C-K level to the process conditions. A weak cation exchange chromatography (WCX) method with or without carboxypeptidase B (CpB) treatment was developed to monitor the C-K level for in-process samples. The effects of operating conditions (i.e., temperature and culture duration) and media trace elements (copper and zinc) on C-K variants were studied. The dominant effect on C-K level was identified as the trace elements concentration. Specifically, increased C-K levels were observed with increase of copper concentration and decrease of zinc concentration in chemically defined medium. Further, a hypothesis for C-K processing with intracellular and extracellular carboxypeptidase activity was proposed, based on preliminary intracellular carboxypeptidase Western blot results and the extracellular HCCF holding study.

journal_name

Biotechnol Bioeng

authors

Luo J,Zhang J,Ren D,Tsai WL,Li F,Amanullah A,Hudson T

doi

10.1002/bit.24510

subject

Has Abstract

pub_date

2012-09-01 00:00:00

pages

2306-15

issue

9

eissn

0006-3592

issn

1097-0290

journal_volume

109

pub_type

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