Bicarbonate concentration and osmolality are key determinants in the inhibition of CHO cell polysialylation under elevated pCO(2) or pH

Accumulation of CO2 in ani mal cell cu Itu res can be a significant problem during scale-up and production of recombinant glycoprotein biopharmaceutic als. By examining the cell-surface polysialic acid (PSA) content, we show t hat elevated CO2 partial pressure (pCO(2)) can alter protein glycosylation. PSA is a high-moiecular-weight polymer attached to several complex N-linke d oligosaccharides on the neural cell adhesion molecule (NCAM), so that sma ll changes in either core glycosylation or in polysialylation are amplified and easily measured. Flowcytometric analysis revealed that PSA levels on C hinese hamster ovary (CHO) cells decrease with increasing pCO(2) in a dose- dependent manner, independent of any change in NCAM content. The results ar e highly pH-dependent, with a greater decrease in PSA at higher pH. By mani pulating medium pH and pCO(2), we showed that decreases in PSA correlate we ll with bicarbonate concentration ([HCO3-]). In fact, it was possible to of fset a 60% decrease in PSA content at 120 mm Hg pCO(2) by decreasing the pH from 7.3 to 6.9, such that [HCO3-] was lowered to that of control (38 mm H g pCO(2)). When the increase in osmolality associated with elevated [HCO3-] was offset by decreasing the basal medium [NaCl], elevated [HCO3-] still c aused a decrease in PSA, although less extensive than without osmolality co ntrol. By increasing [NaCl], we show that hyperosmolality alone decreases P SA content, but to a lesser extent than for the same osmolality increase du e to elevated [NaHCO3]. In conclusion, we demonstrate the importance of pH and pCO(2) interactions, and show that [HCO3-] and osmolality can account f or the observed changes in PSA content over a wide range of pH and pCO(2) v alues. (C) 1999 John Wiley & Sons, Inc.