Strategies for enhancing therapeutic protein production in recombinant CHO cells under hyperosmotic condition

Abstract

During recombinant Chinese hamster ovary (rCHO) cell culture, various events, such as feeding with concentrated nutrient solutions or the addition of base to maintain an optimal pH, increase the osmolality of the medium. To examine the effect of hyperosmolality on therapeutic protein production, an interferon-β (IFN-β)-producing cell line was cultivated in hyperosmotic condition. For efficient production of native IFN-β in rCHO cell culture, the IFN-β molecular aggregation that occurs during culture needs to be minimized. To do so, we investigated the effect of hyperosmolality and hypothermia on IFN-β production and molecular ag-gregation. Both hyperosmolality (470 mOsm/kg) and hypothermia (32℃) increased specific native INF-β productivity (qIFN-β). Furthermore, they decreased the IFN-β molecular aggregation, although severe IFN-β molecular aggregation could not be avoided in the later phase of culture. To overcome growth suppression at hyperosmolality and hypothermia, cells were cultivated in a biphasic mode. Cells were first cultivated at 310 mOsm/kg and 37℃ for 2 days to rapidly obtain a reasonably high cell concentration. The temperature and osmolality were then shifted to 32℃ and 470 mOsm/kg, respectively, to achieve high qIFN-β and reduced IFN-β molecular aggregation. Due to the enhanced qIFN-β and delayed molecular aggregation, the highest native IFN-β concentration achieved on day 6 was 18.03 ± 0.61 mg/L, which was 5.30-fold higher than that in a control batch culture (310 mOsm/kg and 37℃). Taken together, a combination of hyperosmolality and hypothermia in a biphasic culture is a useful strategy for improved native IFN-β production from rCHO cells. The hyperosmotic condition has detrimental effect on cell growth despite its merit in $q_p$. To determine the effect of hyperosmotic stress on apoptosis, type Ⅰ programmed cell death (PCD), and autophagy, which can be type Ⅱ PCD or a survival mechanism, of rCHO cells, two rCHO cell lines, producing antibody ...