EGCG improves recombinant protein productivity in Chinese hamster ovary cell cultures via cell proliferation control

Abstract

Chinese hamster ovary cell lines are good manufacturing practice-certified host cells and are widely used in the field of biotechnology to produce therapeutic antibodies. Recombinant protein productivity in cells is strongly associated with cell growth. To control cell proliferation, many approaches have previously been tested including: genetic engineering, chemical additives such as cell cycle inhibitors, and temperature shift of the culture. To be widely adopted in the biopharmaceutical industry, the culture methods should be simple, uniform and safe. To this end, we examined the use a natural compound to improve the production capacity. In this study, we focused on the antioxidants, catechin polyphenols, which are found in green tea, for cell proliferation control strategies. (-)-Epigallocatechin-3-gallate (EGCG), the major catechin that induces G0/G1 cell cycle arrest, was investigated for its effect on recombinant protein production. Adding EGCG to the cell culture media resulted in slower cellular growth and longer cell longevity, which improved the specific productivity and total yield of recombinant IgG1 in batch cultures by almost 50% for an extra 2 or 3 days of culture. A lower L-glutamine consumption rate was observed in cells cultured in EGCG-containing media, which may be suggesting that there was less stress in the culture environment. Additionally, EGCG did not affect the N-glycan quality of IgG1. Our results indicated that adding EGCG only on the first day of the culture enhanced the specific productivity and total amount of recombinant protein production in batch cultures. This approach may prove to be useful for biopharmaceutical production.

Establishment of fast-growing serum-free immortalised cells from Chinese hamster lung tissues for biopharmaceutical production

“…For example, the temperature shift strategy is a famous way to suppress cell proliferation in CHO and CHL cell cultures 14 , 24 . Additionally, we have previously found that epigallocatechin-3-gallate (EGCG), which induces G0/G1 cell cycle arrest, improves recombinant IgG1 productivity in CHO cells 25 . This strategy could be applied to control the proliferation of CHL-YN cells, although conditions such as EGCG concentration may differ.…”

“…Finally, the cells were acclimated to chemical differential medium, EX-CELL CD CHO Fusion (#14365C, Sigma-Aldrich) medium containing 6 mM l -glutamine (Wako Pure Chemical Industries, Ltd.). After adaptation, the cells were cultured at 37 °C in 125-mL Erlenmeyer flasks (Corning Inc., Corning, NY, USA) with shaking at 80 rpm, 5% CO 2 , and 80% humidity in an orbital Climo Shaker ISF1-X (Kuhner Shaker, Inc., Basel, Switzerland) as described in a previous report 25 . An automated cell analyser (Vi-cell XR, Beckman Coulter, Inc., Brea, CA, USA) was used to analyse total/viable cell concentrations 25 .…”

“…After adaptation, the cells were cultured at 37 °C in 125-mL Erlenmeyer flasks (Corning Inc., Corning, NY, USA) with shaking at 80 rpm, 5% CO 2 , and 80% humidity in an orbital Climo Shaker ISF1-X (Kuhner Shaker, Inc., Basel, Switzerland) as described in a previous report 25 . An automated cell analyser (Vi-cell XR, Beckman Coulter, Inc., Brea, CA, USA) was used to analyse total/viable cell concentrations 25 . CHO-K1 (ATCC CCL-61) cells (American Type Culture Collection, Manassas, VA, USA) adapted to EX-CELL CD CHO Fusion medium in the same manner as CHL-YN cells were used for comparison.…”

“…It was performed as described in a previous report 25 . IgG1 purification from the cell culture supernatants and labelling of released N-glycans with 2-aminobenzamide were performed by EZGlyco (Sumitomo Bakelite Co. Ltd., Tokyo, Japan).…”

Serum-Free Medium for Recombinant Protein Expression in Chinese Hamster Ovary Cells

“…Rather, valeric acid improved galactosylation of the mAb ( Backliwal et al, 2008 ; Park et al, 2016b ). Catechins induced G0/G1 cell cycle arrest to affect the recombinant protein production; glucosylation of catechin required access to a panel of catechin derivatives protected on all hydroxyl groups, except the one to be glucosylated ( Yamano and Omasa, 2018 ). Meanwhile, VPA inhibited both class I and II histone deacetylases (HDACs), with high potency for class I HDACs, and induced cell cycle arrest at the G1 phase.…”