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Promoter methylation and transgene copy numbers predict unstable protein production in recombinant chinese hamster ovary cell lines
Author(s) -
Osterlehner Andrea,
Simmeth Silke,
Göpfert Ulrich
Publication year - 2011
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.23216
Subject(s) - chinese hamster ovary cell , biology , transgene , methylation , dna methylation , cell culture , heterologous , recombinant dna , cpg site , microbiology and biotechnology , gene , genetics , gene expression
Abstract Mammalian cell lines for recombinant protein production need to maintain productivity over extended cultivation times. Long‐term stability studies are time and resource intensive, but are widely performed to identify and eliminate unstable candidates during cell line development. Production instability of manufacturing cell lines can be associated with methylation and silencing of the heterologous promoter. We have identified CpG dinucleotides within the human cytomegalovirus major immediate early promoter/enhancer (hCMV‐MIE) that are frequently methylated in unstable antibody‐producing Chinese hamster ovary (CHO) cell lines. We have established methylation‐specific real‐time qPCR for the rapid and sensitive measurement of hCMV‐MIE methylation in multiple cell lines and provide evidence that hCMV‐MIE methylation and transgene copy numbers can be used as early markers to predict production instability of recombinant CHO cell lines. These markers should provide the opportunity to enrich stable producers early in cell line development and allow developers to put more emphasis on other criteria, such as product quality and bioprocess robustness. Biotechnol. Bioeng. 2011;108: 2670–2681. © 2011 Wiley Periodicals, Inc.