基因组规模的功能基因组学筛选强调了影响中国仓鼠卵巢细胞蛋白质岩藻糖基化的基因。

IF 2.7 4区 生物学 Q2 BIOCHEMICAL RESEARCH METHODS SLAS Discovery Pub Date : 2024-01-01 DOI:10.1016/j.slasd.2023.10.004
Kari Barlan , Gaurang P Bhide , Derek R White , Marc R Lake , Charles Lu , Stephanie E Rieder , Lianchun Fan , Chen-Lin Hsieh
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引用次数: 0

摘要

N-连接糖基化是一种常见的翻译后修饰,对多种类型的蛋白质具有不同的影响。由于糖基化会影响治疗性单克隆抗体(mAb)的疗效和安全性,因此N-连接糖蛋白的修饰程度和所涉及的聚糖物种的身份对生物制药行业非常感兴趣。例如,缺乏核心岩藻糖的单克隆抗体通过增加抗体依赖性细胞毒性而显示出增强的临床疗效。我们在中国仓鼠卵巢(CHO)细胞中进行了全基因组CRISPR敲除筛选,CHO细胞是用于工业生产单克隆抗体的主力细胞培养系统,旨在鉴定蛋白质岩藻糖基化的新调节因子。使用凝集素结合测定,我们鉴定了224个显著改变蛋白质岩藻糖基化的基因扰动,包括众所周知的糖基化基因。这一功能基因组学框架可以很容易地扩展和应用于研究涉及其他糖型调控的遗传途径。我们希望这一资源将为下一代CHO细胞系和mAb疗法的开发提供有用的指导。
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Genome-scale functional genomics screening highlights genes impacting protein fucosylation in Chinese hamster ovary cells

N-linked glycosylation is a common post-translational modification that has various effects on multiple types of proteins. The extent to which an N-linked glycoprotein is modified and the identity of glycans species involved is of great interest to the biopharmaceutical industry, since glycosylation can impact the efficacy and safety of therapeutic monoclonal antibodies (mAbs). mAbs lacking core fucose, for example, display enhanced clinical efficacy through increased antibody-dependent cellular cytotoxicity. We performed a genome-wide CRISPR knockout screen in Chinese hamster ovary (CHO) cells, the workhorse cell culture system for industrial production of mAbs, aimed at identifying novel regulators of protein fucosylation. Using a lectin binding assay, we identified 224 gene perturbations that significantly alter protein fucosylation, including well-known glycosylation genes. This functional genomics framework could readily be extended and applied to study the genetic pathways involved in regulation of other glycoforms. We hope this resource will provide useful guidance toward the development of next generation CHO cell lines and mAb therapeutics.

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来源期刊
SLAS Discovery
SLAS Discovery Chemistry-Analytical Chemistry
CiteScore
7.00
自引率
3.20%
发文量
58
审稿时长
39 days
期刊介绍: Advancing Life Sciences R&D: SLAS Discovery reports how scientists develop and utilize novel technologies and/or approaches to provide and characterize chemical and biological tools to understand and treat human disease. SLAS Discovery is a peer-reviewed journal that publishes scientific reports that enable and improve target validation, evaluate current drug discovery technologies, provide novel research tools, and incorporate research approaches that enhance depth of knowledge and drug discovery success. SLAS Discovery emphasizes scientific and technical advances in target identification/validation (including chemical probes, RNA silencing, gene editing technologies); biomarker discovery; assay development; virtual, medium- or high-throughput screening (biochemical and biological, biophysical, phenotypic, toxicological, ADME); lead generation/optimization; chemical biology; and informatics (data analysis, image analysis, statistics, bio- and chemo-informatics). Review articles on target biology, new paradigms in drug discovery and advances in drug discovery technologies. SLAS Discovery is of particular interest to those involved in analytical chemistry, applied microbiology, automation, biochemistry, bioengineering, biomedical optics, biotechnology, bioinformatics, cell biology, DNA science and technology, genetics, information technology, medicinal chemistry, molecular biology, natural products chemistry, organic chemistry, pharmacology, spectroscopy, and toxicology. SLAS Discovery is a member of the Committee on Publication Ethics (COPE) and was published previously (1996-2016) as the Journal of Biomolecular Screening (JBS).
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