Efficient mAb production in CHO cells incorporating PEI-mediated transfection, mild hypothermia and the co-expression of XBP-1
Efficient mAb production in CHO cells incorporating PEI-mediated transfection, mild hypothermia and the co-expression of XBP-1
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DOI：
10.1002/jctb.2572
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发表时间：
2011-07-01
影响因子：
3.4
通讯作者：
Munro, Trent P.
中科院分区：
文献类型：
--
作者：
Codamo, Joe;Hou, Jeff Jia Cheng;Munro, Trent P.
背景：瞬时基因表达（TGE）为临床前评估提供了一种快速产生重组蛋白生物制剂的方法。人胚肾（HEK 293）细胞传统上用于TGE;然而，工业上需要利用中国仓鼠卵巢（CHO）细胞的高效、高产TGE系统。一个聚乙烯亚胺（PEI）为基础的TGE过程已经开发了CHO细胞使用附加型表达系统，以产生增强的重组蛋白titers.Results：一个五倍的单克隆抗体（mAb）的体积生产率的改善，通过检查关键参数，包括转染介质，细胞密度，转染试剂，DNA：试剂比，转移到亚低温和喂养策略转染后的时间。Epi-CHO系统允许转染后培养体积扩大六倍，而不会显着影响比生产率。当与非附加体系统相比时，该系统产生每μ g质粒DNA多400%的mAb。此外，使用X-box结合蛋白1，以提高分泌能力，并提供进一步的改进，在单克隆抗体生产与TGEweredinvestigated.CONCLUSION：通过优化的关键参数，我们的研究结果表明，开发一个低成本，高产量，游离型TGE系统，可通过在临床前生物药物开发。(C)2011年化学工业协会
BACKGROUND: Transient gene expression (TGE) provides a rapid way to generate recombinant protein biologics for pre-clinical assessment. Human embryonic kidney (HEK293) cells have traditionally been used for TGE; however, there is demand from industry for efficient, high-producing TGE systems that utilize Chinese hamster ovary (CHO) cells. A polyethyleneimine (PEI)-based TGE process has been developed for CHO cells using an episomal expression system to generate enhanced recombinant protein titers.RESULTS: A five-fold improvement in monoclonal antibody (mAb) volumetric productivity was achieved by examining key parameters including transfection medium, cell density, transfection reagent, DNA: reagent ratio, the time of transfer to mild hypothermia and feeding strategy post-transfection. The Epi-CHO system allowed for a six-fold expansion in culture volume post-transfection without significantly affecting specific productivity. This system generates 400% more mAb per mu g of plasmid DNA when compared with a non-episomal system. In addition, the use of X-box binding protein 1 to enhance secretion capacity and provide further improvements in mAb production with TGE was investigated.CONCLUSION: Through optimization of key parameters, our results demonstrate the development of a low-cost, high-yielding, episomal TGE system that may be adopted during pre-clinical biologic drug development. (C) 2011 Society of Chemical Industry