Xu Sen, Jiang Rubin, Mueller Roland, Hoesli Nadja, Kretz Thomas, Bowers John, Chen Hao
Upstream Process Development and Engineering, Biologics Process Development & Clinical Manufacturing, Merck & Co., Inc., Kenilworth, NJ.
Biologics Process Development & Clinical Manufacturing, MSD Werthenstein BioPharma GmbH, Schachen, Switzerland.
Biotechnol Prog. 2018 May;34(3):756-766. doi: 10.1002/btpr.2620. Epub 2018 Mar 6.
Lactate metabolism variations are frequently encountered in mammalian cell culture processes, especially during process scale-up. In this study, we took a multipronged approach to investigate the impact of pH, pCO , osmolality, base addition, and mixing conditions on the observed lactate variations in a Chinese Hamster Ovary (CHO) fed-batch process at 2,000 L scale. Two cultivating methods, CO -controlled and pH-controlled, were used to decouple the individual and synergistic effects from those factors. The individual effects from pH, pCO , and osmolality on lactate consumption/reproduction in the stationary phase were insignificant in the ranges studied though the initial lactate production rates varied. In contrast, lactate metabolism was found to be impacted by an interaction between mixing conditions and CO accumulation. High CO accumulation and poor mixing led to lactate reproduction, whereas either low CO or improved mixing were sufficient to result in lactate consumption. Base addition was not required for pH control in the low CO conditions, and therefore lactate reproduction was correlated with base addition under poor mixing conditions. Under good mixing conditions, CO -triggered base addition did not significantly impact lactate reproduction. It is reasonable to postulate that increased mixing times further promoted lactate production during base addition. As lactate reproduction results in more base addition to maintain pH, a cycle could be formed between lactate production and base addition. As a remediation, we showed that such lactate reproduction could be eliminated by improving CO removal at 2,000 L scale. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:756-766, 2018.
乳酸代谢变化在哺乳动物细胞培养过程中经常出现，尤其是在工艺放大期间。在本研究中，我们采用了多管齐下的方法来研究pH值、二氧化碳分压、渗透压、碱添加量和混合条件对2000升规模的中国仓鼠卵巢（CHO）补料分批培养过程中观察到的乳酸变化的影响。采用了两种培养方法，即二氧化碳控制法和pH值控制法，以区分这些因素的单独作用和协同作用。尽管初始乳酸产生速率有所不同，但在所研究的范围内，pH值、二氧化碳分压和渗透压对稳定期乳酸消耗/产生的单独影响并不显著。相比之下，发现乳酸代谢受到混合条件和二氧化碳积累之间相互作用的影响。高二氧化碳积累和不良混合导致乳酸产生，而低二氧化碳或改善混合足以导致乳酸消耗。在低二氧化碳条件下，pH值控制不需要添加碱，因此在混合不良条件下，乳酸产生与碱添加相关。在良好的混合条件下，二氧化碳触发的碱添加对乳酸产生没有显著影响。可以合理推测，增加混合时间会在添加碱期间进一步促进乳酸产生。由于乳酸产生会导致添加更多碱来维持pH值，因此在乳酸产生和碱添加之间可能会形成一个循环。作为一种补救措施，我们表明，在2000升规模下，通过改善二氧化碳去除可以消除这种乳酸产生。©2018美国化学工程师学会生物技术进展，34:756 - 766，2018。
