PD-1 silencing impairs the anti-tumor function of chimeric antigen receptor modified T cells by inhibiting proliferation activity

Jianshu Wei  ¹ , Can Luo  ¹ , Yao Wang  ¹ , Yelei Guo  ¹ , Hanren Dai  ¹ , Chuan Tong  ¹ , Dongdong Ti  ¹ , Zhiqiang Wu  ² , Weidong Han  ³

• ¹ Department of Bio-therapeutic, Department of Molecular & Immunology, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China.
• ² Department of Bio-therapeutic, Department of Molecular & Immunology, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. wuzhiqiang1006@163.com.
• ³ Department of Bio-therapeutic, Department of Molecular & Immunology, Chinese PLA General Hospital, No. 28 Fuxing Road, Beijing, 100853, China. hanwdrsw69@yahoo.com.

Abstract

Background: Blocking programmed death-1 (PD-1) is considered to be a promising strategy to improve T cell function, and this is being explored in many ongoing clinical trials. In fact, our knowledge about PD-1 is primarily based on the results of short-term experiments or observations, but how long-lasting PD-1 blockade can affect T cell function remains unclear.

Methods: We planned to use shRNA-based gene knockdown technology to mimic long-lasting PD-1 blockade. We constructed PD-1 steadily blocked chimeric antigen receptor modified T (CAR-T) cells, and with these cells we can clearly study the effects of PD-1 knockdown on T cell function. The anti-tumor function, proliferation ability and differentiation status of PD-1 silenced CAR-T cells were studied by in vitro and animal experiments.

Results: According to short-term in vitro results, it was reconfirmed that the resistance to programmed death-ligand 1 (PD-L1)-mediated immunosuppression could be enhanced by PD-1 blockade. However, better anti-tumor function was not presented by PD-1 blocked CAR-T cells in vitro or in vivo experiments. It was found that PD-1 knockdownmight impair the anti-tumor potential of CAR-T cells because it inhibited T cells’ proliferation activity. In addition, we observed that PD-1 blockade would accelerate T cells’ early differentiation and prevent effector T cells from differentiating into effect memory T cells, and this might be the reason for the limited proliferation of PD-1 silenced CAR-T cells.

Conclusion: These results suggest that PD-1 might play an important role in maintaining the proper proliferation and differentiation of T cells, and PD-1 silencing would impair T cells’ anti-tumor function by inhibiting their proliferation activity.

Keywords: Chimeric antigen receptor modified T cells; PD-1 blockade; Persistence; T cell differentiation; T cell proliferation.