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CHO synthetic promoters improve expression and product quality of biotherapeutic proteins

机译:CHO合成启动子可提高生物治疗性蛋白的表达和产物质量

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摘要

Abstract When expressing complex biotherapeutic proteins, traditional expression plasmids and methods may not always yield sufficient levels of high‐quality product. High‐strength viral promoters commonly used for recombinant protein (rProtein) production in mammalian cells allow for maximal expression, but provide limited scope to alter their transcription dynamics. However, synthetic promoters designed to provide tunable transcriptional activity offer a plasmid engineering approach to more precisely regulate product quality, yield or to reduce product related contaminants. We substituted the viral promoter CMV with synthetic promoters that offer different transcriptional activities to express our gene of interest in Chinese hamster ovary (CHO) cells. Stable pools were established and the benefits of regulating transgene transcription on the quality of biotherapeutics were examined in stable pool fed‐batch overgrow experiments. Specific control of gene expression of the heavy chain (HC):light chain (LC) of a Fab, and the ratio between the two HCs in a Duet mAb reduced levels of aberrant protein contaminants; and the controlled expression of the helper gene XBP‐1s improved expression of a difficult‐to‐express mAb. This synthetic promoter technology benefits applications that require custom activity. Our work highlights the advantages of employing synthetic promoters for production of more complex rProteins.
机译:摘要 在表达复杂的生物治疗性蛋白时,传统的表达质粒和方法可能并不总是能产生足够水平的高质量产物。通常用于哺乳动物细胞中重组蛋白 (rProtein) 生产的高强度病毒启动子允许最大表达,但改变其转录动力学的范围有限。然而,旨在提供可调转录活性的合成启动子提供了一种质粒工程方法,可以更精确地调节产品质量、产量或减少产品相关污染物。我们用合成启动子代替病毒启动子CMV,这些启动子提供不同的转录活性,以表达我们在中国仓鼠卵巢(CHO)细胞中的目标基因。建立了稳定的池,并在稳定的池饲分批过度生长实验中检查了调节转基因转录对生物治疗药物质量的益处。对 Fab 的重链 (HC):轻链 (LC) 的基因表达进行特异性控制,以及 Duet mAb 中两种 HC 之间的比率可降低异常蛋白质污染物的水平;辅助基因 XBP-1 的受控表达改善了难表达单克隆抗体的表达。这种合成启动子技术有利于需要定制活性的应用。我们的工作突出了使用合成启动子生产更复杂的rProteins的优势。

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