The combined effect of sodium butyrate and low culture temperature on the production, sialylation, and biological activity of an antibody produced in CHO cells

Abstract

Cell cultures containing 0 ~ 5 mM sodium butyrate (NaBu) and grown at 30 and 37°C were conducted to investigate the combined effect of NaBu and low temperature on the quantity and quality of an antibody production in CHO cells. Although NaBu addition decreased cell viability by apoptosis in a dose-dependent manner at both 30 and 37°C, the onset of significant apoptosis induced by NaBu was delayed by lowering culture temperature. The highest specific antibody productivity (q p ) of 23.26 pg/cell/day was obtained in the culture containing 2 mM NaBu at 30°C; however, the highest antibody concentration of 167.84 mg/L was achieved in the culture containing 1 mM NaBu at 30°C, as the detrimental effect of further NaBu addition on cell growth compromised its beneficial effect on q p . Moreover, protein quality with respect to the total sialic acid content and Nglycolylneuraminic acid (Neu5Gc) level was evaluated. There were no apparent changes regarding the total sialic acid content of the antibody, but manipulation of cultures with NaBu treatment or (and) low culture temperature did decrease Neu5Gc levels by 5 ~ 10%. Biological activity of the antibody was also assessed, and no obvious changes were observed. Collectively, the simultaneous application of NaBu and low culture temperature was an effective way to extend culture period and enhance final antibody concentration, without compromising the sialic acid content or biological activity.

Enhanced recombinant factor VII expression in Chinese hamster ovary cells by optimizing signal peptides and fed-batch medium

“…16,24 Several tested chemicals had toxic effect on the cell growth and led to the decline of viable cell density. [19][20][21] In this study, high percentage of G0/G1 phase cells was accompanied by the reduced cell density in all tested feed media. However, the decreased of cell density after adding 3 feed media was different and F4 had a slower cell density decline.…”

“…Some cell cycle arresting chemicals had toxic effect on cell growth and thus cell density reduced after adding this chemicals. [19][20][21] In this study, cell density also reduced after day 6 and this may be caused by cell cycle arresting component in feed media. However, the cell density decline in M4CF4 was slower than other 2 fed-batch culture (M4CF2 and M4CF3) (Fig.…”

“…2,4,5,23 Moreover, it has been reported that the cell cycles phase, specifically the percentage of cells in the G0/G1 phase, significantly influences the specific productivity and production of recombinant proteins in CHO cells. [16][17][18][19] Many researchers have focused on screening for chemicals that pause the cells in the G0/ G1 phase to enhance protein specific productivity in CHO cells. 16,24 Several tested chemicals had toxic effect on the cell growth and led to the decline of viable cell density.…”

4.1 Control of Biotheraputics Glycosylation

“…Humans produce Neu5Ac sialic acid, whereas CHO cells produce both, Neu5Gc and Neu5Ac. Sodium butyrate supplementation increased the ratio of Neu5Ac to Neu5Gc, which may be implicated in increasing the circulatory half-life of the recombinant protein therapeutic [79,80]. The decreased synthesis of Neu5Gc may relate to decreased CMPNeu5Ac hydroxylase activity or NADH supply.…”

“…Butyrate was also investigated for its effect on glycosylation and was found to increase site occupancy of N-linked glycoforms [61,78]. The increase in site occupancy was linearly correlated with an increase in sialic acid content to suggest that butyrate may increase sialyltransferase activity [61,78,79]. Humans produce Neu5Ac sialic acid, whereas CHO cells produce both, Neu5Gc and Neu5Ac.…”