CRISPR knock out of programmed cell death protein 1 enhances anti-tumor activity of cytotoxic T lymphocytes

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// Zhilong Zhao 1 , Long Shi 2 , Wei Zhang 4 , Jinsheng Han 5 , Shaohui Zhang 6 , Zexian Fu 7 and Jianhui Cai 1, 3 1 Department of Surgery, Hebei Medical University, Shijiazhuang, Hebei, China 2 Department of Oncology, Hebei Medical University Second Affiliated Hospital, Shijiazhuang, Hebei, China 3 Department of Surgery & Oncology, Hebei General Hospital, Shijiazhuang, Hebei, China 4 Department of Surgery, Handan Central Hospital, Handan, Hebei, China 5 Department of Surgery, Cangzhou Hospital of Traditional Chinese Medicine and Western Medicine Integrated Hebei, Cangzhou, Hebei, China 6 Department of Oncology, Hebei Medical University Third Affiliated Hospital, Shijiazhuang, Hebei, China 7 Department of Oncology, Hebei University of Engineering Affiliated Hospital, Baoding, Hebei, China Correspondence to: Jianhui Cai, email: jianhuicai2002@163.com Keywords: anti-tumor immunity; CRISPR; cytotoxic T lymphocyte; immune checkpoint; PD-1 Received: July 18, 2017 Accepted: August 27, 2017 Published: December 27, 2017 ABSTRACT Programmed cell death protein 1 (PD-1) is an immune checkpoint receptor that functions to attenuate T cell activation. In this study, we knocked out (KO) PD-1 in cytotoxic T lymphocytes (CTLs) using CRISPR-Cas9 system to evaluate its effect on the anti-tumor activity of the CTLs against multiple myeloma (MM). Results show that PD-1 KO CTLs facilitate apoptosis and caspase activation of the co-cultured MM cells and enhanced MM cell death by 36% compared with the control. PD-1 KO also increased TNF-α and IFN-γ secretion of the CTLs by 2.4 and 1.9-fold respectively. The effectiveness of PD-1 KO in enhancing anti-tumor activity of the CTLs was verified in vivo using mouse xenograft model. The xenografted mice treated with PD-1 KO CTLs demonstrated repressed MM tumor growth and prolonged survival compared with the control group. We conclude that CRISPR-Cas9 is an efficient system to knock out PD-1 from CTLs and PD-1 KO could significantly enhance the anti-tumor activity of CTLs.