Modulation of nutrient precursors for controlling metabolic inhibitors by flux balance analysis
Modulation of nutrient precursors for controlling metabolic inhibitors by flux balance analysis
复制标题
通过通量平衡分析调节营养前体以控制代谢抑制剂
DOI：
10.1002/btpr.3313
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发表时间：
2022
影响因子：
2.9
通讯作者：
Yoon, Seongkyu
中科院分区：
文献类型：
--
作者：
Hoang, Duc;Kuang, Bingyu;Liang, George;Wang, Zhao;Yoon, Seongkyu
治疗性蛋白质生产率和糖基化模式高度依赖于细胞代谢。细胞培养基组成和补料策略对调节细胞代谢至关重要。在这项研究中，探索了有毒代谢抑制剂及其营养前体之间的关系，以确定细胞生长和代谢副产物产生的关键培养基组分。定制通用CHO代谢模型，并将其与CHO补料分批代谢组学数据整合，以获得细胞系和工艺特定模型。对毒性代谢物胞苷一磷酸、鸟苷一磷酸和n-乙酰腐胺进行了通量平衡分析研究，所有这些代谢物都是先前报道的由内源性细胞代谢产生的，通过将它们映射到一个区室化的碳利用网络。使用这种方法，当比较三种工业相关的分批补料条件时，该研究预测了高水平的抑制性代谢物蓄积，并通过剂量依赖性氨基酸加标研究验证了结果。最后，采用培养基优化设计，以降低补充的营养素的量，其中实现了关键工艺性能的改善，在峰值活细胞密度（VCD）增加40%，积分VCD增加15%，和生长速率增加37%。还实现了对毒性副产物的严格控制，因为该研究测量了所有条件下抑制性代谢物蓄积的减少。总的来说，该研究成功地提出了一种数字孪生方法来研究补充培养基成分与宿主细胞代谢产生的下游毒性代谢产物之间的相互关系，进一步阐明了能够改善细胞表型和调节毒性抑制剂的不同控制策略。
Therapeutic protein productivity and glycosylation pattern highly rely on cell metabolism. Cell culture medium composition and feeding strategy are critical to regulate cell metabolism. In this study, the relationship between toxic metabolic inhibitors and their nutrient precursors was explored to identify the critical medium components toward cell growth and generation of metabolic by‐products. Generic CHO metabolic model was tailored and integrated with CHO fed‐batch metabolomic data to obtain a cell line‐ and process‐specific model. Flux balance analysis study was conducted on toxic metabolites cytidine monophosphate, guanosine monophosphate and n‐acetylputrescine—all of which were previously reported to generate from endogenous cell metabolism—by mapping them to a compartmentalized carbon utilization network. Using this approach, the study projected high level of inhibitory metabolites accumulation when comparing three industrially relevant fed‐batch feeding conditions one against another, from which the results were validated via a dose‐dependent amino acids spiking study. In the end, a medium optimization design was employed to lower the amount of supplemented nutrients, of which improvements in critical process performance were realized at 40% increase in peak viable cell density (VCD), 15% increase in integral VCD, and 37% increase in growth rate. Tight control of toxic by‐products was also achieved, as the study measured decreased inhibitory metabolites accumulation across all conditions. Overall, the study successfully presented a digital twin approach to investigate the intertwined relationship between supplemented medium constituents and downstream toxic metabolites generated through host cell metabolism, further elucidating different control strategies capable of improving cellular phenotypes and regulating toxic inhibitors.