Precise gene editing technologies are providing new opportunities to stably engineer host cells for recombinant production of therapeutic glycoproteins with different glycan structures. The glycosylation of recombinant therapeutics has long been a focus for both quality and consistency of products and for optimizing and improving pharmacokinetic properties as well as bioactivity. Structures of glycans on therapeutic glycoproteins are important for circulation, biodistribution and bioactivity. In particular, the latter has been demonstrated for therapeutic IgG1 antibodies where the core α1,6Fucose on the conserved N-glycan at Asn297 have remarkable dampening effects on antibody effector functions. We previously explored precise gene engineering and design options for N-glycosylation in CHO cells, and here we focus on engineering options possible for N-glycans on human IgG1. We demonstrate stable precise gene engineering of rather homogenous biantennary N-glycans with and without galactose (G0F, G2F) as well as the α2,6-linked monosialylated (G2FS1) glycoform. We were unable to introduce substantial disialylated glycoforms. Instead we engineered a novel monoantennary homogeneous N-glycan design with complete α2,6-linked sialic acid capping. All N-glycoforms may be engineered with and without core α1,6Fucose. The stably engineered design options enable production of human IgG antibodies with an array of distinct glycoforms for testing and selection of optimal design for different therapeutic applications.

中文翻译：

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使用精确基因编辑的CHO细胞中IgG1糖工程设计选项

精确的基因编辑技术为稳定改造重组具有不同聚糖结构的治疗性糖蛋白宿主细胞提供了新的机会。长期以来，重组治疗剂的糖基化一直是产品质量和一致性以及优化和改善药代动力学性质以及生物活性的焦点。治疗性糖蛋白上的聚糖结构对于循环，生物分布和生物活性很重要。特别是，后者已被证明可用于治疗性IgG1抗体，其中保守N上的核心α1,6FucoseAsn297的β-聚糖对抗体效应子功能具有显着的抑制作用。先前我们探索了CHO细胞中N-糖基化的精确基因工程和设计选项，在这里，我们重点研究人IgG1上N-聚糖的工程选项。我们展示了相当均一的双天线N聚糖（有和没有半乳糖（G0F，G2F）以及α2,6-连接的单唾液酸化（G2FS1）糖型）的稳定精确基因工程。我们无法引入大量的二唾液酸化糖型。取而代之的是，我们设计了一种新颖的单天线均质N-聚糖设计，该设计具有完整的α2,6-连接的唾液酸封端。全部N可以在有或没有核心α1,6Fucose的情况下改造β-糖型。稳定设计的设计选项使人IgG抗体具有一系列独特的糖型，可用于测试和选择针对不同治疗应用的最佳设计。