Overexpression of cold-inducible RNA-binding protein increases interferon-γ production in Chinese-hamster ovary cells

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Abstract

Culturing recombinant CHO (Chinese-hamster ovary) cells at low temperatures (30–33 °C) increases specific recombinant protein productivity by 2–5-fold. However, even though the specific productivity is increased, cell growth is decreased in low-temperature culture such that the final recombinant protein titre remains unchanged or is even diminished, owing to the lower cell density. Exposing mammalian cells to low temperatures results in a change in the expression of many ‘cold-stress’ genes. CIRP (cold-inducible RNA-binding protein) is a cold-stress protein that is highly expressed at 32 °C, but not at 37 °C. In the present study we demonstrated that overexpression of CIRP at 37 °C can increase the recombinant-protein titre in CHO cells. Stable overexpression of CIRP at 37 °C improved the final titre of CHO IFN-γ, a recombinant CHO cell line producing human IFN-γ (interferon-γ), by 25% in adherent culture and up to 40% in suspension culture. Real-time PCR analysis showed that the increase in the recombinant IFN-γ titre could be attributed to increased recombinant IFN-γ mRNA levels, while growth data showed that CIRP overexpression did not result in growth arrest in CHO IFN-γ cells. Glycan analysis showed that the increase in IFN-γ titre as a result of CIRP overexpression did not affect the site occupancy, glycan structures or sialic acid content of IFN-γ. Using this strategy, the final IFN-γ titre was increased by 40% compared with current temperature-based strategies. Furthermore, there is no decrease in cell growth or recombinant-protein glycosylation quality, as previously observed in low-temperature culture.

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