CRISPR/Cas9 mediated deletion of the adenosine A2A receptor enhances CAR T cell efficacy

Adenosine is an immunosuppressive factor that limits anti-tumor immunity through the suppression of multiple immune subsets including T cells via activation of the adenosine A 2A receptor (A 2A R). Using both murine and human chimeric antigen receptor (CAR) T cells, here we show that targeting A 2A R with a clinically relevant CRISPR/Cas9 strategy significantly enhances their in vivo efficacy, leading to improved survival of mice. Effects evoked by CRISPR/Cas9 mediated gene deletion of A 2A R are superior to shRNA mediated knockdown or pharmacological blockade of A 2A R. Mechanistically, human A 2A R-edited CAR T cells are significantly resistant to adenosine-mediated transcriptional changes, resulting in enhanced production of cytokines including IFNγ and TNF, and increased expression of JAK-STAT signaling pathway associated genes. A 2A R deficient CAR T cells are well tolerated and do not induce overt pathologies in mice, supporting the use of CRISPR/Cas9 to target A 2A R for the improvement of CAR T cell function in the clinic. Activation of the adenosine receptor A2AR is associated with suppression of T cell function in the tumor microenvironment. To overcome immunosuppression, here the authors show that CRISPR/Cas9 mediated deletion of A2AR enhances CAR T cell effector functions without altering memory or persistence properties, improving CAR-T mediated tumor control in pre-clinical models.