Immune function assessing of TIM3/CD28‐modified CD19 CAR‐T cells and general CD19 CAR‐T cells through a high‐throughput single‐cell microarray platform

Abstract Chimeric antigen receptor (CAR) T cells are widely used to treat hematological tumors due to their powerful ability to target and kill cancer cells, of which accurate function evaluation at the single‐cell level is crucial to ensuring the efficacy of diagnosis and treatment. Currently, a universal platform to evaluate the efficacy of immune single cells rapidly, efficiently, and systematically is urgently needed. Here, the cytotoxicity, proliferative potential, and persistence of TIM3/CD28‐modified CD19 CAR‐T cells are evaluated in comparison with ordinary CD19 CAR‐T cells through high‐performance and throughput graphene oxide quantum dot (GOQD)‐based single‐cell microfluidic chips. Overall secretory factor expression, immune‐therapy effect of different effector‐target ratios, spatial immune‐therapy effects, and subgroup type profiling are demonstrated to explicit the immunotherapy effect of TIM3/CD28‐modified CD19 CAR‐T cells. TIM3/CD28‐modified CD19 CAR‐T cells show stronger anti‐tumor ability and maintain excellent immunotherapy effects even at low effector‐target ratios and remote distances. TIM3/CD28 also strengthens the local targeting ability of TIM3/CD28‐modified CD19 CAR‐T. Importantly, TIM3/CD28‐modified CD19 CAR‐T exhibits more distinct Th1/Th2 long‐term persistent and potent killer subgroups, which is very helpful for personalized therapy. Overall, this study provides a valuable approach that can be widely implemented to analyze current CAR‐T combinations and evaluate the function of innovative CAR treatments in the future.