Effect of culture pH on erythropoietin production by Chinese hamster ovary cells grown in suspension at 32.5 and 37.0°C

Yoon, Seong Jun; Choi, Sang Lim; Song, Ji Yong; Lee, Gyun Min

doi:10.1002/bit.20353

Cited by 140 publications

(114 citation statements)

References 31 publications

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“…negatively growth-associated) in the CHO1-15 500 cell line. This finding is in agreement with several investigators who have correlated reduced specific growth rate to increased recombinant protein productivity for CHO cell lines utilizing the DHFR amplification system (Fox et al 2005;Fussenegger et al 1997;Hayduk and Lee 2005;Hendrick et al 2001;Kaufmann et al 1999;Yoon et al 2003Yoon et al , 2005. Although not conclusive, this negative growth-association suggests that the protein may be preferentially or exclusively produced during the G1 cell cycle phase.…”

Section: Specific Productivitysupporting

confidence: 91%

Cell cycle phase dependent productivity of a recombinant Chinese hamster ovary cell line

Dutton¹,

Scharer

Moo-Young

2007

Cytotechnology

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A Chinese Hamster Ovary cell line, CHO1-15 500 , producing recombinant human tissue type plasminogen activator (tPA) via the dihydrofolate reductase (DHFR) amplification system, was studied in batch culture. In this system both DHFR and tPA are under the control of the strong constitutive viral SV40 early promoter. Employing the cumulative viable cell-hour approach, the specific productivity of tPA had maxima in the lag (0.065 pg cell -1 h -1 ) and early decline (0.040 pg cell -1 h -1 ) population growth phases. The viable population was assigned into four subpopulations (G1, S, G2/M phase, and Apoptotic cells) using flow cytometric analysis. As expected, intracellular DHFR was maximally expressed during the S cell cycle phase. The production of tPA, however, was found to be a direct linear function of the G1 phase, with a subpopulation specific productivity of 0.080 pg c-h -1. Productivity maxima in the lag and early decline corroborate the flow cytometric data, indicative that this recombinant tPA production occurs primarily in the G1 cell cycle phase, not the S phase. This suggests that endogenous regulatory mechanisms are important controlling influences on the production of recombinant tPA in this ovarian cell line. Productivity from recombinant cell lines cannot be inferred from either the plasmid construct or the host cell alone. Elucidation of the relationship between expression of recombinant protein and the cell cycle phases of the host cell is a major component of the characterization of the animal cell production system. This information facilitates rational process design, including operating mode, modelling and control, and medium formulation.Keywords Cell cycle associated protein productivity Á Chinese hamster ovary (CHO) Á DHFR Á Regulation of expression Á Specific productivity Á Tissue plasminogen activator (tPA) Abbreviation CH vol volumetric cell-hours

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Section: Specific Productivitysupporting

confidence: 91%

Cell cycle phase dependent productivity of a recombinant Chinese hamster ovary cell line

Dutton¹,

Scharer

Moo-Young

2007

Cytotechnology

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“…Hypothermic cultivation of mammalian cells is widely used in the biotechnology industry (11)(12)(13)(14)(15). However, to date no substantial difference was detected in the glycosylation of recombinant proteins expressed under conditions of mild hypothermia from recombinant cell lines or transiently transfected cells as compared to production of the same antibody at 37°C (12,18,43).…”

Section: Discussionmentioning

confidence: 99%

“…In standard bioprocesses, mammalian cells are cultivated at 37°C, but it is well-known that mild hypothermic conditions (27-34°C) can induce an increase in protein production in some recombinant cell lines (11)(12)(13)(14)(15)(16). More recently, hypothermic conditions have been shown to enhance TGE in CHO cells (17,18).…”

Section: Introductionmentioning

confidence: 99%

Mild Hypothermia Improves Transient Gene Expression Yields Several Fold in Chinese Hamster Ovary Cells

Wulhfard

Tissot

Bouchet

et al. 2008

Biotechnology Progress

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Large-scale transient gene expression (TGE) in mammalian cells is a rapid method to generate recombinant proteins, but the volumetric productivity for secreted proteins is still more than an order of magnitude lower than the yields typically achieved with recombinant cell lines. Here transient recombinant protein production in Chinese hamster ovary cells transfected with linear 25 kDa polyethylenimine was significantly enhanced by incubation of the cells at temperatures ranging from 29 to 33°C after DNA delivery. With this approach, transient recombinant antibody yields of 60-80 mg/L were achieved within 6 days of transfection. The increase in TGE correlated with the accumulation of cells in the G1 phase of the cell cycle, increased cell size, higher cell viability, higher steady-state levels of transgene mRNA, reduced consumption of nutrients, and decreased accumulation of waste products. The enhancement of TGE was not vector-dependent, but the presence of the woodchuck hepatitis virus post-transcriptional regulatory element in the 3′ untranslated region of the transgene mRNA increased transient recombinant antibody expression more than 3-fold at 31°C as compared to expression at 37°C. The yields achieved by the low-temperature enhancement of TGE in CHO cells makes this technology feasible for the rapid production of gram amounts of secreted recombinant proteins at large scale (up to 100 L).

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“…It can be noticed that the pH decrease has been already shown to decrease glucose uptake and to promote the cell oxidative metabolism (Yoon et al 2005;Kuwae et al 2005;Burky et al 2007). The pH decrease occurring after 45 h in Erlenmeyer flask culture could have induced this metabolism shift.…”

Section: Glycolysismentioning

confidence: 99%

Combination of yeast hydrolysates to improve CHO cell growth and IgG production

et al. 2012

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Many studies underlined the great benefits of hydrolysates used as additives in animal free media on cell culture performances. However, to precisely define hydrolysate supplementation strategies, a deeper understanding of their effect on cell growth and protein production is required. In the present study, the effect of addition of one yeast extract (YE) and two yeast peptones (named YP.A and YP.B) in a chemically defined medium was first assessed on cell culture performances. Interestingly, specific effects were found depending on the degree of degradation of yeast hydrolysates. The YE at 1 g L -1 increased the maximal cell density by 70 %, while a mixture of YE (1 g L -1 ) and YP.A (4 g L -1 ) increased IgG production by 180 %. These conditions were then evaluated on the CHO cell kinetics all over cultures. Hydrolysates extended the cell growth phase in Erlenmeyer flask and increased the maximal growth rate in bioreactor up to 20 %. Cell growth stimulation induced by hydrolysates addition was linked with energetic metabolism improvement suggesting that they promote oxidative pathway. Furthermore, hydrolysates provided an additional source of substrate that supported cell growth despite glutamine limitation.

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Effect of culture pH on erythropoietin production by Chinese hamster ovary cells grown in suspension at 32.5 and 37.0°C

Cited by 140 publications

References 31 publications

Cell cycle phase dependent productivity of a recombinant Chinese hamster ovary cell line

Cell cycle phase dependent productivity of a recombinant Chinese hamster ovary cell line

Mild Hypothermia Improves Transient Gene Expression Yields Several Fold in Chinese Hamster Ovary Cells

Combination of yeast hydrolysates to improve CHO cell growth and IgG production

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