Generation of apoptosis‐resistant HEK293 cells with CRISPR/Cas mediated quadruple gene knockout for improved protein and virus production. Issue 11 (17th August 2017)

• Record Type: Journal Article
• Authors: Zhang, Weifeng; Xiao, Dan; Shan, Linlin; Zhao, Junli; Mao, Qinwen; Xia, Haibin
• Is Part Of: Biotechnology and bioengineering. Volume 114:Issue 11(2017)
• Page Start: 2539
• Page End: 2549
• Publication Date: 2017-08-17
• Subjects: adenovirus -- apoptosis -- CRISPR/Cas -- gene therapy -- genome editing; Biotechnology -- Periodicals; Bioengineering -- Periodicals; 660.6
• DOI: 10.1002/bit.26382
• Languages: English
• ISSNs: 0006-3592

Abstract

Abstract: Apoptosis has important functions during pathophysiologic processes. However, from a biopharmaceutical point of view, active apoptosis of host cells is undesirable during viral packaging or protein expression, because it decreases the efficiency of viral or protein production. Here we used the CRISPR/Cas technique to knock out four pro‐apoptotic genes, Caspase3, Caspase6, Caspase7 and AIF1, in HEK293 cells, and successfully produced an apoptosis‐resistant cell line. Furthermore, this cell line showed higher expression levels of pro‐apoptotic proteins and higher packaging efficiency for the virus carrying these proteins than control HEK293 cells. This study not only produced an apoptosis‐resistant cell line that is useful in producing apoptosis‐inducing proteins or viruses expressing these proteins, but also provides a methodology to build other apoptosis‐resistant cell lines. Abstract : The authors established an apoptosis‐resistant HEK293 cells with CRISPR/Cas mediated quadruple gene knockout to improve protein and virus production. Caspase3 gene was firstly knocked out in HEK293 cells followed by the simultaneous knockout of Caspase6, Caspase7 and AIF1 genes. The produced cell line is proven to be resistant to apoptosis triggers from both intracellular and extracellular environments, and shows increased capacity of protein expression and viral packaging.