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Cas9-cleavage sequences in minimal plasmids enhance non-viral genome targeting of CARs in primary human T cells

By Ruirui Jing, Peng Jiao, Jiangqing Chen, Xianhui Meng, Xiaoyan Wu, Yanting Duan, Kai Shang, Liling Qian, Yanjie Huang, Junwei Liu, Tao Huang, Jin Jin, Wei Chen, Xun Zeng, Weiwei Yin, Xiaofei Gao, Chun Zhou, Michel Sadelain, Jie Sun

Posted 02 Jan 2021
bioRxiv DOI: 10.1101/2020.12.31.424920

T cell genome editing holds great promise to advance a range of immunotherapies but is encumbered by the dependence on difficult-to-produce and expensive viral vectors. Here we have designed small double-stranded plasmid DNA modified to mediate high-efficiency homologous recombination. The resulting chimeric antigen receptor (CAR)-T cells display a similar phenotype, transcriptional profile and in vivo potency as CAR-T cells generated using adeno-associated viral (AAV) vector. This method should simplify and accelerate the use of precision engineering to produce edited T cells for research and clinical purposes.

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