Informace o publikaci
Low glucose depletes glycan precursors, reduces site occupancy and galactosylation of a monoclonal antibody in CHO cell culture
| Autoři | |
|---|---|
| Rok publikování | 2015 |
| Druh | Článek v odborném periodiku |
| Časopis / Zdroj | BIOTECHNOLOGY JOURNAL |
| Fakulta / Pracoviště MU | |
| Citace | |
| www | http://onlinelibrary.wiley.com/doi/10.1002/biot.201400662/abstract |
| Doi | http://dx.doi.org/10.1002/biot.201400662 |
| Obor | Imunologie |
| Klíčová slova | Adenylate energy charge; Chinese hamster ovary (CHO) cells; Glucose depletion; N-glycosylation; Nucleotide sugars |
| Popis | Controlled feeding of glucose has been employed previously to enhance the productivity of recombinant glycoproteins but there is a concern that low concentrations of glucose could limit the synthesis of precursors of glycosylation. Here we investigate the effect of glucose depletion on the metabolism, productivity and glycosylation of a chimeric human-llama monoclonal antibody secreted by CHO cells. The cells were inoculated into media containing varying concentrations of glucose. Glucose depletion occurred in cultures with an initial glucose 5.5 mM and seeded at low density (2.5 x 10(5) cells/mL) or at high cell inoculum (2.5 x 10(6) cells/mL) at higher glucose concentration (up to 25 mM). Glucose-depleted cultures produced non-glycosylated Mabs (up to 51%), lower galactosylation index (GI <0.43) and decreased sialylation (by 85%) as measured by mass spectrometry and HPLC. At low glucose a reduced intracellular pool of nucleotides (0.03-0.23 fmoles/cell) was measured as well as a low adenylate energy charge (<0.57). Low glucose also reduced GDP-sugars (by 77%) and UDP-hexosamines (by 90%). The data indicate that under glucose deprivation, low levels of intracellular nucleotides and nucleotide sugars reduced the availability of the immediate precursors of glycosylation. These results are important when applied to the design of fed-batch cultures. |