CHO DUKX cell lineages preadapted to growth in serum-free suspension culture enable rapid development of cell culture processes for the manufacture of recombinant proteins

Sinacore, Martin S.; Charlebois, Timothy S.; Harrison, Scott H.; Brennan, Stephanie; Richards, Thomas J.; Hamilton, Maureen A.; Scott, S.; Brodeur, Scott R.; Oakes, Patrick W.; Leonard, Mark; Switzer, Mary E.; Anagnostopoulos, A.; Foster, Bradley A.; Harris, Andrew L.; Jankowski, Maurycy; Bond, M.; Martin, Sara; Adamson, Sheena

doi:10.1002/(sici)1097-0290(19961120)52:4<518::aid-bit7>3.0.co;2-s

Cited by 49 publications

(21 citation statements)

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“…1 samples for cell counting were withdrawn. We observed a generation time of ≈25 h in CHO-T1-SF medium, a value that is comparable to other results published for CHO cells grown in suspension (Gasser et al, 1985;Lee et al, 1996;Renner et al, 1995;Sharif and Stevenson, 1993;Sinacore et al, 1996).…”

Section: Growth Characteristics In Serum-free Medium Cho-t1-sfsupporting

confidence: 91%

“…This is the optimum for rodent cells in vitro (Parchment and Natarajan, 1992). For other serum-free media generation times of ≈24 h were reported (Gasser et al, 1985;Lee et al, 1996;Renner et al, 1995;Sharif and Stevenson, 1993;Sinacore et al, 1996). Only for medium UC203 generation times similar to those observed in medium CHO-T1-SF were reported (Ii et al, 1995).…”

Section: Growth Characteristics In Serum-free Medium Cho-t1-sfmentioning

confidence: 93%

“…However, in CHO DUKXB11 cells only one allele carries a deletion, whereas the other carries a point mutation . This allele can revert, and result in overgrowth of the culture by dhfr + cells during adaptation to serum-free conditions Sinacore et al, 1996). For this reason we tested if reversion to dhfr + and overgrowth of dhfr -cells by dhfr + cells occurred during the adaptation process to serum-free conditions.…”

Section: Colony Forming Efficiencies In Medium Cho-t1-sfmentioning

confidence: 99%

“…A protein-free medium for CHO SSF3 cells was described (Zang et al, 1995). In addition, a cell line preadapted to serum-free conditions, derived from DUKXB11 cells was described (Sinacore et al, 1996). However, the serum-free medium formulation used in this study was not published.…”

Section: Introductionmentioning

confidence: 99%

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Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11

Schröder

Matischak

Friedl

2004

Journal of Biotechnology

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Citation for published item:hr¤ oderD wF nd w tis h kD uF nd priedlD F @PHHRA 9 erumE nd proteinEfree medi formul tions for the ghinese h mster ov ry ell line h u fIIF9D tourn l of iote hnologyFD IHV @QAF ppF PUWEPWPF Further information on publisher's website: httpXGGdxFdoiForgGIHFIHITGjFj iote FPHHQFIPFHHS Publisher's copyright statement:Additional information: Use policyThe full-text may be used and/or reproduced, and given to third parties in any format or medium, without prior permission or charge, for personal research or study, educational, or not-for-pro t purposes provided that:• a full bibliographic reference is made to the original source • a link is made to the metadata record in DRO • the full-text is not changed in any way The full-text must not be sold in any format or medium without the formal permission of the copyright holders.Please consult the full DRO policy for further details. AbstractThe production of therapeutic proteins in mammalian cell lines is of outstanding importance. The maintenance of most mammalian cell lines in culture requires the addition of serum to the culture medium.The elimination of serum from mammalian cell culture is desirable since serum is expensive and a source of contaminants, e.g. viruses, mycoplasma or prions. Here we describe the composition of serum-and protein-free media for the Chinese hamster ovary (CHO) cell line DUKXB11. The serum-free formulation supports excellent growth of CHO DUKXB11 cells at low (23 cells/cm 2 ) and high (2·10 4 cells/cm 2 ) seeding densities characterized by a generation time of 10 -12 h, and, after addition of 0.2% pluronic F-68, the growth of a recombinant suspension cell line derived from DUKXB11. In addition, this formulation also allowed us to adapt recombinant cell lines expressing various amounts of human antithrombin ATIII (ATIII)to serum-free conditions. Secretion of ATIII was readily observed in the serum-free medium. Minor changes to the serum-free formulation resulted in a protein free formulation that supported growth of CHO DUKXB11 cells, growth of recombinant CHO cells expressing ATIII, and production of ATIII. Key wordsChinese hamster ovary cells, serum-free medium, protein-free medium, recombinant protein production,

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Section: Growth Characteristics In Serum-free Medium Cho-t1-sfsupporting

confidence: 91%

Section: Growth Characteristics In Serum-free Medium Cho-t1-sfmentioning

confidence: 93%

Section: Colony Forming Efficiencies In Medium Cho-t1-sfmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11

Schröder

Matischak

Friedl

2004

Journal of Biotechnology

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“…NS0 cells are capable of cholesterol-free growth and can be isolated by selecting mutant clones (Birch et al 1994). Adaptation generally involves passaging cells over time during which the cholesterol concentration is gradually reduced (Sinacore et al 1996). In this study by Seth et al (2005), proteomic tools were used to compare the protein profiles of regular 'wild-type' NS0 cells and a 'revertant' cell line that can grow without cholesterol.…”

Section: Media Supplementationmentioning

confidence: 99%

Proteomic profiling of recombinant cells from large-scale mammalian cell culture processes

Meleady

2007

Cytotechnology

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Global expression profiling of mammalian cells used for the production of biopharmaceuticals will allow greater insights into the molecular mechanisms that result in a high producing cellular phenotype. These studies may give insights for genetic intervention to possibly create better host cell lines or even to provide clues to more rational strategies for cell line and process development. In this review I will focus on the contribution of proteomic technologies to a greater understanding of the biology of Chinese hamster ovary cells and other producing cell lines such as NS0 mouse cells.

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Cytogenetic analysis of chimeric antibody-producing CHO cells in the course of dihydrofolate reductase-mediated gene amplification and their stability in the absence of selective pressure

Kim

Lee

1999

Biotechnol. Bioeng.

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Previously, the highest producing (HP) recombinant CHO subclones isolated at various methotrexate (MTX) levels showed different antibody production stability during long‐term culture, although they were clonally derived from CS13 transformant. In this study, genetic basis for their difference in antibody production stability was investigated using southern blot hybridization and fluorescence in situ hybridization (FISH) techniques. Southern analysis of HP subclones revealed that light‐chain (LC) and heavy‐chain (HC) cDNAs were located closely within 23 kb on an amplification unit, and the configuration of LC and HC cDNAs within this amplification unit was not disrupted during long‐term culture in the absence of MTX. However, when LC and HC genes were localized on the metaphase chromosomes of HP subclones using FISH, the amplified sequences were present as an extended array on diverse marker chromosomes. HP subclones selected at higher MTX level had more kinds of marker chromosomes. CS13*‐002 isolated at 0.02 μM MTX had only one marker chromosome (m002), whereas CS13*‐1.0 isolated at 1 μM MTX had five different ones (m10A, m10B, m10C, m10D, and m10E). Each marker chromosome showed different fate during long‐term culture of HP subclones in the absence of MTX, resulting in different degrees of stability among the HP subclones. The m10A and m10B remained unchanged, whereas the others disappeared or evolved to variants with shortened amplified arrays. The cells containing stable marker chromosomes constituted dominant subpopulations in CS13*‐1.0, and thereby CS13*‐1.0 became most stable in regard to antibody production during long‐term culture. Furthermore, our dual‐color FISH showed that the telomeric ends of amplified arrays on the stable marker chromosomes were always surrounded by (TTAGGG)n sequences, indicating that (TTAGGG)n sequences are closely related to the stability and evolution of amplified sequences. Taken together, our data show that the assessment of genotypic stability of amplified CHO cells is a prerequisite for understanding their production stability during long‐term culture in the absence of selection pressure. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 64: 741–749, 1999.

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CHO DUKX cell lineages preadapted to growth in serum-free suspension culture enable rapid development of cell culture processes for the manufacture of recombinant proteins

Cited by 49 publications

References 33 publications

Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11

Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11

Proteomic profiling of recombinant cells from large-scale mammalian cell culture processes

Cytogenetic analysis of chimeric antibody-producing CHO cells in the course of dihydrofolate reductase-mediated gene amplification and their stability in the absence of selective pressure

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