Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Ibaraki, Japan.
Biotechnol Prog. 2019 Jul;35(4):e2820. doi: 10.1002/btpr.2820. Epub 2019 Apr 22.
Chinese hamster ovary (CHO) cells have been used as host cells in the production of a range of recombinant therapeutic proteins, including monoclonal antibodies and Fc-fusion proteins. Host cell proteins (HCP) represent impurities that must be removed from therapeutic formulations because of their potential risks for immunogenicity. While the majority of HCP impurities are effectively removed in typical downstream purification processes, clearance of a small population of HCP remains challenging. In this study, we knocked out the Anxa2 and Ctsd genes to assess the feasibility of knockout approaches for diminishing the risk of contamination with HCP. Using the CRISPR/Cas9 system, Anxa2-, and Ctsd-knockout CHO cell lines were successfully established, and we confirmed the complete elimination of the corresponding HCP in cell lysates. Importantly, all knockout cell lines showed similar growth and viability to those of the wild-type control during 8 days of cultivation. Thus, knockout of unrequired genes can reduce contamination with HCP in the production of recombinant therapeutic proteins.
中国仓鼠卵巢（CHO）细胞已被用作生产一系列重组治疗蛋白的宿主细胞，包括单克隆抗体和 Fc 融合蛋白。宿主细胞蛋白（HCP）是杂质，由于其潜在的免疫原性风险，必须从治疗制剂中去除。虽然大多数 HCP 杂质在典型的下游纯化过程中被有效去除，但清除一小部分 HCP 仍然具有挑战性。在这项研究中，我们敲除了 Anxa2 和 Ctsd 基因，以评估敲除方法降低 HCP 污染风险的可行性。使用 CRISPR/Cas9 系统，成功建立了 Anxa2 和 Ctsd 基因敲除 CHO 细胞系，并证实细胞裂解物中相应的 HCP 完全消除。重要的是，在 8 天的培养过程中，所有敲除细胞系的生长和活力与野生型对照相似。因此，敲除不需要的基因可以减少重组治疗蛋白生产中 HCP 的污染。
