Biologics represent the fastest growing group of therapeutics, but many advanced recombinant protein moieties remain difficult to produce. Here, we identify metabolic engineering targets limiting expression of recombinant human proteins through a systems biology analysis of the transcriptomes of CHO and HEK293 during recombinant expression. In an expression comparison of 24 difficult to express proteins, one third of the challenging human proteins displayed improved secretion upon host cell swapping from CHO to HEK293. Guided by a comprehensive transcriptomics comparison between cell lines, especially highlighting differences in secretory pathway utilization, a co-expression screening of 21 secretory pathway components validated ATF4, SRP9, JUN, PDIA3 and HSPA8 as productivity boosters in CHO. Moreover, more heavily glycosylated products benefitted more from the elevated activities of the N- and O-glycosyltransferases found in HEK293. Collectively, our results demonstrate the utilization of HEK293 for expression rescue of human proteins and suggest a methodology for identification of secretory pathway components for metabolic engineering of HEK293 and CHO.

生物制剂代表了增长最快的疗法组，但许多先进的重组蛋白部分仍然难以生产。在这里，我们通过对重组表达过程中 CHO 和 HEK293 转录组的系统生物学分析，确定了限制重组人类蛋白质表达的代谢工程靶点。在 24 种难以表达的蛋白质的表达比较中，三分之一的具有挑战性的人类蛋白质在宿主细胞从 CHO 交换到 HEK293 后表现出改善的分泌。在细胞系之间的全面转录组学比较的指导下，特别是突出了分泌途径利用的差异，对 21 种分泌途径成分的共表达筛选验证了 ATF4、SRP9、JUN、PDIA3 和 HSPA8 作为 CHO 的生产力促进剂。而且，HEK293 中发现的 N- 和 O- 糖基转移酶活性升高，使糖基化程度更高的产品受益更多。总的来说，我们的结果证明了 HEK293 用于人类蛋白质的表达拯救，并提出了一种用于识别 HEK293 和 CHO 代谢工程的分泌途径成分的方法。