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Enhanced Human Thrombopoietin Production by Sodium Butyrate Addition to Serum‐Free Suspension Culture of Bcl‐2‐Overexpressing CHO Cells

When sodium butyrate (NaBu) was added to serum‐free suspension culture of recombinant CHO (rCHO) cells for enhanced expression of human thrombopoietin (hTPO), apoptotic cell death of rCHO cells was induced in a dose‐dependent manner and hTPO quality was deteriorated in regard to sialic acid and acidic isoform contents. To overcome these problems, we overexpressed Bcl‐2 protein, an antiapoptotic protein, in rCHO cells producing hTPO. Compared to serum‐free suspension culture of control cells without Bcl‐2 overexpression (R‐neo cells) and NaBu addition, a more than 10‐fold increase in the maximum hTPO concentration was obtained in serum‐free suspension culture of cells with Bcl‐2 overexpression (R‐bcl2–14 cells) and 3 mM NaBu addition. Both the enhanced specific productivity endowed by NaBu and the extended culture longevity provided by the antiapoptotic effect of Bcl‐2 overexpression contributed to the enhancement of maximum hTPO concentration. The problem of quality reduction of hTPO induced by NaBu was not solved by Bcl‐2 overexpression, but it was not that significant. Compared to the culture in the absence of NaBu, the percentage of hTPO isoforms in pI 3–5 with high in vivo biological activity produced by R‐bcl2–14 cells was decreased by approximately 18% in the presence of 3 mM. As a result, a more than 6‐fold increase in the production of hTPO isoforms in pI 3–5 was achieved in R‐bcl2–14 cell culture with 3 mM NaBu addition. Taken together, the data obtained suggest that Bcl‐2 overexpression in rCHO cells and NaBu addition in serum‐free suspension culture can be an effective means to enhance the production of highly glycosylated protein such as hTPO.