Aberrant metabolic sialylation of recombinant proteins expressed in Chinese hamster ovary cells in high productivity cultures

The incorporation of sialic acid into therapeutic recombinant glycoprotein expressed in Chinese hamster ovary (CHO) cells during growth in large biore actors (10 l) has been monitored under high productivity conditions induced by the presence of sodium butyrate. Samples of the bioreactor culture (sim ilar to 4 x 10(6) cells) were labeled with H-3-N-acetylmannosamine, a metab olic precursor of sialic acid. After 24 h, the recombinant glycoprotein, an immunoadhesion chimeric molecule, was purified and the amount of sialic ac id incorporated was determined as radioactive counts. The labeling profile of the protein over the course of the culture was compared with the sialic acid content of the molecule as determined by direct chemical analysis. Ear ly in the culture, the two methods of analysis gave a similar sialylation p rofile. However, after sodium butyrate was included in the culture, the met abolically incorporated sialic acid rapidly and dramatically decreased to n ear undetectable levels. In contrast, sialic acid content of the protein, a s determined by chemical analysis, decreased only moderately and gradually over the culture period, from a maximum of 6.1 to about 5.0 mol sialic acid /mole of protein after 10 days in culture. These results suggest that butyr ate may enhance reutilization of existing glycoproteins in the culture, gen erating sialic acid for biosynthesis through lysosomal degradation and ther eby bypassing de novo biosynthesis. (C) 1999 Academic Press.