{"title":"The new frontier in CHO cell line development: From random to targeted transgene integration technologies","authors":"","doi":"10.1016/j.biotechadv.2024.108402","DOIUrl":null,"url":null,"abstract":"<div><p>Cell line development represents a crucial step in the development process of a therapeutic glycoprotein. Chinese hamster ovary (CHO) cells are the most frequently employed mammalian host cell system for the industrial manufacturing of biologics. The predominant application of CHO cells for heterologous recombinant protein expression lies in the relative simplicity of stably introducing ectopic DNA into the CHO host cell genome. Since CHO cells were first used as expression host for the industrial production of biologics in the late 1980s, stable genomic transgene integration has been achieved almost exclusively by random integration. Since then, random transgene integration had become the gold standard for generating stable CHO production cell lines due to a lack of viable alternatives. However, it was eventually demonstrated that this approach poses significant challenges on the cell line development process such as an increased risk of inducing cell line instability. In recent years, significant discoveries of new and highly potent (semi)-targeted transgene integration systems have paved the way for a technological revolution in the cell line development sector. These advanced methodologies comprise the application of transposase-, recombinase- or Cas9 nuclease-mediated site-specific genomic integration techniques, which enable a scarless transfer of the transgene expression cassette into transcriptionally active loci within the host cell genome. This review summarizes recent advancements in the field of transgene integration technologies for CHO cell line development and compare them to the established random integration approach. Moreover, advantages and limitations of (semi)-targeted integration techniques are discussed, and benefits and opportunities for the biopharmaceutical industry are outlined.</p></div>","PeriodicalId":8946,"journal":{"name":"Biotechnology advances","volume":null,"pages":null},"PeriodicalIF":12.1000,"publicationDate":"2024-06-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S073497502400096X/pdfft?md5=11fd21ba09a2621d0568d8fa03e17d05&pid=1-s2.0-S073497502400096X-main.pdf","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biotechnology advances","FirstCategoryId":"5","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S073497502400096X","RegionNum":1,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Cell line development represents a crucial step in the development process of a therapeutic glycoprotein. Chinese hamster ovary (CHO) cells are the most frequently employed mammalian host cell system for the industrial manufacturing of biologics. The predominant application of CHO cells for heterologous recombinant protein expression lies in the relative simplicity of stably introducing ectopic DNA into the CHO host cell genome. Since CHO cells were first used as expression host for the industrial production of biologics in the late 1980s, stable genomic transgene integration has been achieved almost exclusively by random integration. Since then, random transgene integration had become the gold standard for generating stable CHO production cell lines due to a lack of viable alternatives. However, it was eventually demonstrated that this approach poses significant challenges on the cell line development process such as an increased risk of inducing cell line instability. In recent years, significant discoveries of new and highly potent (semi)-targeted transgene integration systems have paved the way for a technological revolution in the cell line development sector. These advanced methodologies comprise the application of transposase-, recombinase- or Cas9 nuclease-mediated site-specific genomic integration techniques, which enable a scarless transfer of the transgene expression cassette into transcriptionally active loci within the host cell genome. This review summarizes recent advancements in the field of transgene integration technologies for CHO cell line development and compare them to the established random integration approach. Moreover, advantages and limitations of (semi)-targeted integration techniques are discussed, and benefits and opportunities for the biopharmaceutical industry are outlined.
细胞系开发是治疗性糖蛋白开发过程中的关键一步。中国仓鼠卵巢(CHO)细胞是工业化生产生物制剂最常用的哺乳动物宿主细胞系统。CHO 细胞在异源重组蛋白表达方面的主要应用在于,将异位 DNA 稳定导入 CHO 宿主细胞基因组相对简单。自 20 世纪 80 年代末首次将 CHO 细胞作为表达宿主用于生物制剂的工业化生产以来,稳定的基因组转基因整合几乎完全是通过随机整合实现的。从那时起,由于缺乏可行的替代方法,随机转基因整合已成为产生稳定的 CHO 生产细胞系的黄金标准。然而,最终的事实证明,这种方法给细胞系的开发过程带来了巨大的挑战,如增加了诱导细胞系不稳定性的风险。近年来,新型高效(半)靶向转基因整合系统的重大发现为细胞系开发领域的技术革命铺平了道路。这些先进的方法包括应用转座酶、重组酶或 Cas9 核酸酶介导的位点特异性基因组整合技术,这些技术能将转基因表达盒无疤痕地转移到宿主细胞基因组内的转录活性位点。本综述总结了用于 CHO 细胞系开发的转基因整合技术领域的最新进展,并将其与既有的随机整合方法进行了比较。此外,还讨论了(半)靶向整合技术的优势和局限性,并概述了生物制药行业的优势和机遇。
期刊介绍:
Biotechnology Advances is a comprehensive review journal that covers all aspects of the multidisciplinary field of biotechnology. The journal focuses on biotechnology principles and their applications in various industries, agriculture, medicine, environmental concerns, and regulatory issues. It publishes authoritative articles that highlight current developments and future trends in the field of biotechnology. The journal invites submissions of manuscripts that are relevant and appropriate. It targets a wide audience, including scientists, engineers, students, instructors, researchers, practitioners, managers, governments, and other stakeholders in the field. Additionally, special issues are published based on selected presentations from recent relevant conferences in collaboration with the organizations hosting those conferences.