Document Detail


Enhancement of transient gene expression and culture viability using Chinese hamster ovary cells overexpressing Bcl-x(L).
MedLine Citation:
PMID:  18727128     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Transient gene expression (TGE) provides a method for quickly delivering protein for research using mammalian cells. While high levels of recombinant proteins have been produced in TGE experiments in HEK 293 cells, TGE efforts in the commercially prominent CHO cell line still suffer from inadequate protein yields. Here, we describe a cell-engineering strategy to improve transient production of proteins using CHO cells. CHO-DG44 cells were engineered to overexpress the anti-apoptotic protein Bcl-x(L) and transiently transfected using polyethylenimine (PEI) in serum-free media. Pools and cell lines stably expressing Bcl-x(L) showed enhanced viable cell density and increased production of a glycosylated, therapeutic fusion protein in shake flask TGE studies. The improved cell lines showed fusion protein production levels ranging from 12.6 to 27.0 mg/L in the supernatant compared to the control cultures which produced 6.3-7.3 mg/L, representing a 70-270% increase in yield after 14 days of fed-batch culture. All Bcl-xL-expressing cell lines also exhibited an increase in specific productivity during the first 8 days of culture. In addition to increased production, Bcl-x(L) cell lines maintained viabilities above 90% and less apoptosis compared to the DG44 host which had viabilities below 60% after 14 days. Product quality was comparable between a Bcl-xL-engineered cell line and the CHO host. The work presented here provides the foundation for using anti-apoptosis engineered CHO cell lines for increased production of therapeutic proteins in TGE applications.
Authors:
Brian S Majors; Michael J Betenbaugh; Nels E Pederson; Gisela G Chiang
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Biotechnology and bioengineering     Volume:  101     ISSN:  1097-0290     ISO Abbreviation:  Biotechnol. Bioeng.     Publication Date:  2008 Oct 
Date Detail:
Created Date:  2008-09-01     Completed Date:  2008-11-06     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  7502021     Medline TA:  Biotechnol Bioeng     Country:  United States    
Other Details:
Languages:  eng     Pagination:  567-78     Citation Subset:  IM    
Copyright Information:
(c) 2008 Wiley Periodicals, Inc.
Affiliation:
Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, 3400 North Charles Street, 221 Maryland Hall, Baltimore, Maryland 21218-2694, USA.
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MeSH Terms
Descriptor/Qualifier:
Animals
Apoptosis
CHO Cells
Cell Survival*
Cricetinae
Cricetulus
Gene Expression*
Recombinant Fusion Proteins / biosynthesis*
Transfection
bcl-X Protein / genetics*
Chemical
Reg. No./Substance:
0/Recombinant Fusion Proteins; 0/bcl-X Protein

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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