Restoration of LAT activity improves CAR T-cell sensitivity and persistence in response to antigen-low acute lymphoblastic leukemia

Chimeric antigen receptor (CAR) T-cells induce responses in patients with relapsed/refractory leukemia; however, long-term efficacy is frequently limited by post-CAR relapses. The inability to target antigen-low cells is an intrinsic vulnerability of second-generation CAR T-cells and underlies the majority of relapses following CD22BBz CAR T-cell therapy. We interrogated CD22BBz CAR signaling in response to low antigen and found inefficient phosphorylation of LAT, limiting downstream signaling. To overcome this, we designed the Adjunctive LAT-Activating CAR T-cell (ALA-CART) platform, pairing a second-generation CAR with a LAT-CAR incorporating the intracellular domain of LAT. ALA-CART cells demonstrated reduced differentiation during manufacturing and increased LAT phosphorylation, MAPK signaling and AP-1 activity. Consequently, ALA-CART cells showed improved cytotoxicity, proliferation, persistence and efficacy against antigen-low leukemias that were refractory to clinically-active CD22BBz CAR T-cells. These data suggest restoration of LAT signaling through the ALA-CART platform represents a promising strategy for overcoming multiple mechanisms of CAR T-cell failure.