Comparative metabolite analysis to understand lactate metabolism shift in Chinese hamster ovary cell culture process
Comparative metabolite analysis to understand lactate metabolism shift in Chinese hamster ovary cell culture process
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DOI：
10.1002/bit.23291
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发表时间：
2012-01-01
影响因子：
3.8
通讯作者：
Li, Feng
中科院分区：
文献类型：
--
作者：
Luo, Jun;Vijayasankaran, Natarajan;Li, Feng
在实施用于抗体生产的新化学成分确定的培养基（CDM）配方期间，在某些中国仓鼠卵巢（CHO）细胞系中观察到从乳酸盐产生（LP）到净乳酸盐消耗（LC）表型的代谢转变。此外，这种代谢转变通常导致细胞生长、生产率、工艺稳健性和可扩展性方面的工艺性能改善。在我们以前的研究中，观察到了关键培养基成分铜与乳酸代谢转变之间的相关性。为了进一步研究这一现象，进行了两项补充研究。在第一项研究中，在两种培养基中培养单个细胞系，这两种培养基仅在铜浓度上不同，但已知该细胞系产生LP或LC表型。在第二项研究中，两种不同的细胞系，这是已知具有固有的不同的乳酸代谢特性，培养在相同的培养基中具有高水平的铜;一个细胞系产生乳酸在整个培养过程中，和其他消耗乳酸后，LP的初始阶段。定期收集两项研究的细胞沉淀和上清液样品，并研究其代谢产物谱。代谢分析的主要发现是LP条件下的细胞表现出较低效率的能量代谢，葡萄糖主要转化为丙酮酸盐、山梨醇、乳酸盐和其他糖酵解中间体。这种能量效率的降低可能是由于丙酮酸和乙酰辅酶A不能进入TCA循环。在LP表型中缺乏进入TCA循环或溢流代谢导致细胞ATP供应不足。因此，糖酵解途径仍然是ATP的主要来源，这反过来又导致整个培养物中连续的LP。此外，还观察到游离脂肪酸的蓄积;这被认为是磷脂催化剂的结果，磷脂催化剂用于补充糖酵解产生的能量，以满足LP细胞的需求。对代谢谱的全面审查表明，乳酸代谢转变可能与细胞的氧化代谢能力有关。Biotechnol. Bioeng. 2012;109：146156. (c)2011 Wiley Periodicals，Inc.
A metabolic shift from lactate production (LP) to net lactate consumption (LC) phenotype was observed in certain Chinese hamster ovary (CHO) cell lines during the implementation of a new chemically defined medium (CDM) formulation for antibody production. In addition, this metabolic shift typically leads to process performance improvements in cell growth, productivity, process robustness, and scalability. In our previous studies, a correlation between a key media component, copper, and this lactate metabolism shift was observed. To further investigate this phenomenon, two complementary studies were conducted. In the first study, a single cell line was cultivated in two media that only differed in their copper concentrations, yet were known to generate an LP or LC phenotype with that cell line. In the second study, two different cell lines, which were known to possess inherently different lactate metabolic characteristics, were cultivated in the same medium with a high level of copper; one cell line produced lactate throughout the duration of the culture, and the other consumed lactate after an initial period of LP. Cell pellet and supernatant samples from both studies were collected at regular time intervals, and their metabolite profiles were investigated. The primary finding from the metabolic analysis was that the cells in LP conditions exhibited a less efficient energy metabolism, with glucose primarily being converted into pyruvate, sorbitol, lactate, and other glycolytic intermediates. This decrease in energy efficiency may be due to an inability of pyruvate and acetyl-CoA to progress into the TCA cycle. The lack of progression into the TCA cycle or overflow metabolism in the LP phenotype resulted in the inadequate supply of ATP for the cells. As a consequence, the glycolysis pathway remained the major source of ATP, which in turn, resulted in continuous LP throughout the culture. In addition, the accumulation of free fatty acids was observed; this was thought to be a result of phospholipid catabolism that was being used to supplement the energy produced through glycolysis in order to meet the needs of LP cells. A thorough review of the metabolic profiles indicated that the lactate metabolic shift could be related to the oxidative metabolic capacity of cells. Biotechnol. Bioeng. 2012;109: 146156. (c) 2011 Wiley Periodicals, Inc.