Adaptation process of engineered cell line FCHO/IL-24 stably secreted rhIL-24 in serum-free suspension culture.

Jilei Hua, Yuexian Wei, Yao Zhang, Hanli Xu, Jianlin Ge, Mengzhe Liu, Yuqi Wang, Yinan Shi, Lingling Hou, Hong Jiang

Protein expression and purification 2022 Nov

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Interleukin-24 (IL-24) displays tumor cell-specific proliferation inhibition in vitro and in vivo. Recombinant human IL-24 (rhIL-24) has significantly higher activity, yet significantly lower expression level in mammalian cells than in bacteria. To further realize therapeutic potential of IL-24, we enhanced rhIL-24 expression in mammalian cell systems by adapting engineered Flp-InTMCHO/IL-24 (FCHO/IL-24) cells (adherent cultured in Ham's F12 medium with 10% serum) to serum-free suspension culture. First, MTT assay showed that among four different media (F12, DMEM/F12, 1640 and DMEM), DMEM/F12 medium was the most suitable media for lower-serum adherent culture. Then, cells were adherently cultured in DMEM/F12 with serum concentration reduced from 10% to 0.5% in a gradient manner. Compared to cells in 10% serum, cells in 0.5% serum displayed significantly lower relative cell viability by 40%, increased G0/G1 phase arrest (8.5 ± 2.4%, p < 0.05), decreased supernatant rhIL-24 concentration by 73%, and altered metabolite profiles, such as glucose, lactate and ammonia concentration. Next, the cells were directly adapted to 0.5% serum suspension culture in 125 mL shake flask at 119 rpm with the optimal cell seeding density of 5 × 105 cells/mL (3.3 times higher than that of adherent culture), under which the concentration of rhIL-24 in culture medium was stable at 3.5 ng/mL. Finally, cells adapted to 0.5% serum proliferated better in serum-free medium Eden™-B300S with higher rhIL-24 expression level compared to CDM4CHO. The successful adaptation of engineered cells FCHO/IL-24 laid foundation for adapting cells from adherent culture to suspension serum-free culture to mass produce rhIL-24 protein for therapeutic purposes. Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.

Citation

Jilei Hua, Yuexian Wei, Yao Zhang, Hanli Xu, Jianlin Ge, Mengzhe Liu, Yuqi Wang, Yinan Shi, Lingling Hou, Hong Jiang. Adaptation process of engineered cell line FCHO/IL-24 stably secreted rhIL-24 in serum-free suspension culture. Protein expression and purification. 2022 Nov;199:106154