Capella Roca Berta, Alarcón Miguez Antonio, Keenan Joanne, Suda Srinivas, Barron Niall, O'Gorman Donal, Doolan Padraig, Clynes Martin
National Institute for Cellular Biotechnology, Dublin City University, Dublin 9, Ireland.
SSPC-SFI, Centre for Pharmaceuticals, Dublin City University, Dublin 9, Ireland.
Cytotechnology. 2019 Oct;71(5):915-924. doi: 10.1007/s10616-019-00334-1. Epub 2019 Aug 8.
In order to study the impact of zinc and copper on the titer levels of mAb and recombinant protein in CHO cells, the IgG-expressing (DP12) and EPO-expressing (SK15) cell lines were cultured in chemically defined media with increasing concentrations of either metal. Supplementation with 25 mg/l in CDM media resulted in a significant increase in EPO (1.7-fold) and IgG (2.6-fold) titers compared to control (no added zinc). Titers at this Zn concentration in CDM containing the insulin replacing agent aurintricarboxylic acid (ATA) (CDM + A) showed a 1.8-fold (EPO) and 1.2-fold (IgG) titers increase compared to control. ATA appeared to also reduce the specific productivity (Qp) enhancement induced by Zn-25, with up to 4.9-fold (DP12) and 1.9-fold (SK15) Qp increase in CDM compared to the 1.6-fold (DP12) and 1.5-fold (SK15) Qp increase observed in CDM + A. A 31% reduced Viable Cell Density (VCD) in DP12 was observed in both Zn-supplemented media (3 × 10 cells/ml vs 4.2 × 10 cells/ml, day 5), whereas SK15 Zn-25 cultures displayed a 24% lower peak only in CDM + A (2.2 × 10 cells/ml vs 3.2 × 10 cells/ml, day 5). Supplementation with copper at 13.7-20 mg/l resulted in less significant cell line/product-type dependent effects on titer, VCD and Viability. Analysis of the energetic phenotype of both cell lines in 25 mg/l Zn-supplemented CDM media revealed a twofold increase in the oxygen consumption rate (OCR) compared to non-supplemented cells. Together, these data suggest that high zinc supplementation may induce an increase in oxidative respiration metabolism that results in increased Qp and titers in suspension CHO cultures.
为了研究锌和铜对CHO细胞中mAb和重组蛋白滴度水平的影响，将表达IgG的（DP12）和表达EPO的（SK15）细胞系在含有浓度不断增加的两种金属之一的化学成分确定的培养基中培养。与对照（未添加锌）相比，在CDM培养基中添加25mg/l锌导致EPO（1.7倍）和IgG（2.6倍）滴度显著增加。在含有胰岛素替代剂金精三羧酸（ATA）的CDM（CDM + A）中，该锌浓度下的滴度与对照相比，EPO增加了1.8倍，IgG增加了1.2倍。ATA似乎还降低了由25mg/l锌诱导的比生产率（Qp）提高，与在CDM + A中观察到的1.6倍（DP12）和1.5倍（SK15）的Qp增加相比，CDM中的Qp增加高达4.9倍（DP12）和1.9倍（SK15）。在两种添加锌的培养基中均观察到DP12的活细胞密度（VCD）降低了31%（第5天，3×10⁶个细胞/ml对4.2×10⁶个细胞/ml），而SK15 25mg/l锌培养物仅在CDM + A中显示峰值降低了24%（第5天，2.2×10⁶个细胞/ml对3.2×10⁶个细胞/ml）。添加13.7 - 20mg/l铜对滴度、VCD和活力的细胞系/产品类型依赖性影响较小。对添加25mg/l锌的CDM培养基中两种细胞系的能量表型分析显示，与未添加锌的细胞相比，耗氧率（OCR）增加了两倍。总之，这些数据表明，高锌添加可能会诱导氧化呼吸代谢增加，从而导致悬浮CHO培养物中的Qp和滴度增加。
