Increasing the Productivity of TNFR-Fc in GS-CHO Cells at Reduced Culture Temperatures

다국어 초록 (Multilingual Abstract)

In an effort to improve TNFR-Fc production in GS-CHO cells, batch cultures were performed to investigate the effects of low culture temperature on TNFR-Fc production. It was observed that low culture temperatures resulted in cell cycle arrest in the G1 phase, led to suppressed cell growth, and prolonged the culture period.
Although the highest TNFR-Fc concentration was achieved with a culture temperature of 32°C at 247.4 mg/L, the maximum q_(TNFR-Fc) of 15.7 pg/cells/day was achieved at 30°C. Because the inhibition effect on cell growth at 30°C compromised its beneficial effects specifically to TNFR-Fc productivity, TNFR-Fc concentration at this temperature was not significantly increased. Furthermore, the increase in productivity of specific TNFR-Fc at low culture temperatures was also found to be due to an increase in the transcriptional level of the TNFR-Fc gene, determined by RT-PCR analysis. In addition, low culture temperatures had no significant effect on the degree of sialylation of TNFRFc.
Taken together; a biphasic cultivation process developed in a fed-batch mode with a low temperature-production phase enhanced TNFR-Fc production by GS-CHO cells and therefore offers major potential for bioprocess optimization.

참고문헌 (Reference)

1 Fogolín, M. B., "of temperature reduction and expression of yeast pyruvate carboxylase on hGM-CSF-producing CHO cells" 109 : 179-191, 2004

2 Roessler, B., "Temperature: A simple parameter for process optimization in fed-batch cultures of recombinant Chinese hamster ovary cells" 18 : 423-427, 1996

3 Jenkins, N., "Temperature control of growth and productivity in mutant Chinese hamster ovary cells synthesizing a recombinant protein" 42 : 1029-1036, 1993

4 Mohler, K. M., "Soluble tumor necrosis factor (TNF) receptors are effective therapeutic agents in lethal endotoxemia and function simultaneously as both TNF carriers and TNF antagonists" 151 : 1548-1561, 1993

5 Kaufmann, H., "Influence of low temperature on productivity, proteome and protein phosphorylation of CHO cells" 63 : 573-582, 1999

6 Mazur, X., "Higher productivity of growth-arrested Chinese hamster ovary cells expressing the cyclin-dependent kinase inhibitor p27" 14 : 705-713, 1998

7 Chuppa, S., "Fermentor temperature as a tool for control of high-density perfusion cultures of mammalian cells" 55 : 328-338, 1997

8 Reuveny, S., "Factors affecting cell growth and monoclonal antibody production in stirred reactors" 86 : 53-59, 1986

9 Renard, J. M., "Evidence that monoclonal antibody production kinetics is related to the integral of viable cells in batch systems" 10 : 91-96, 1988

10 Ducharme, E., "Etanercept" 8 : 491-502, 2008

11 Müthing, J., "Effects of buffering conditions and culture pH on production rates and glycosylation of clinical phase I anti-melanoma mouse IgG3 monoclonal antibody R24" 83 : 321-334, 2003

12 Ting Chen, "Effects of Low Temperature and Lactate on Osteogenic Differentiation of Human Amniotic Mesenchymal Stem Cells" 한국생물공학회 14 (14): 708-715, 2009

13 Yoon, S. K., "Effect of low culture temperature on specific productivity and transcription level of anti-4-1BB antibody in recombinant Chinese hamster ovary cells" AMER CHEMICAL SOC 19 : 1383-1386, 2003

14 Ahn, W. S., "Effect of culture temperature on erythropoietin production and glycosylation in a perfusion culture of recombinant CHO cells" 101 : 1234-1244, 2008

15 Furukawa, K., "Effect of culture temperature on a recombinant CHO cell line producing a C-terminal aamidating enzyme" 26 : 153-164, 1998

16 Borys, M. C., "Culture pH affects expression rates and glycosylation of recombinant mouse placental lactogen proteins by Chinese hamster ovary (CHO) cells" 11 : 720-724, 1993

17 Kubbies, M., "Cell cycle-dependent DHFR and t-PA production in cotransfected, MTX-amplified CHO cells revealed by dual-laser flow cytometry" 188 : 267-271, 1990

18 Gawlitzek, M., "Ammonium alters N-glycan structures of recombinant TNFRIgG: Degradative versus biosynthetic mechanisms" 68 : 637-646, 2000

19 Fussenegger, M., "A novel cytostatic process enhances the productivity of Chinese hamster ovary cells" 55 : 927-939, 1997

20 Masuda, S., "A new biological strategy for high productivity of recombinant proteins in animal cells by the use of hypoxia-response enhancer" 67 : 157-164, 1999