Modulating AP-1 enables CAR-T cells to establish an intratumoral stem-like reservoir and overcomes resistance to PD-1 blockade
CAR-T therapy has shown limited synergy with PD-1/PD-L1 blockade, but the mechanisms underlying resistance remain unclear. PD-1?TCF1? stem-like T cells mediate responses to PD-1/PD-L1 blockade and are maintained by MHC-dependent interactions with DCs in tumor-draining lymph nodes (dLN). Because CAR-Ts recognize intact antigen, not peptide-MHC, their activation is confined to tumors, potentially preventing maintenance of this critical subset. In ROR1? lung cancer models, ROR1-targeting CAR-Ts rapidly down-regulated TCF1, became exhausted, and were not enhanced by PD-L1 blockade. Overexpression of the AP-1 transcription factor c-Jun, but not BATF, enabled formation of PD-1?TCF1? reservoirs in tumors, not dLN, but did not prevent exhaustion, as PD-1 induced post-transcriptional c-Jun down-regulation. Remarkably, PD-L1 blockade restored c-Jun levels, dramatically increased CAR-Ts, and enabled near-complete ROR1+ tumor clearance. These findings identify PD-1 as a suppressor of c-Jun and reveal that MHC-independent CAR-Ts can be engineered to establish intratumoral stem-like reservoirs that overcome resistance to PD-1 blockade. Overall design: Bulk RNA-seq profiling of ROR1 CAR-T and cJun.ROR1 CAR-T cells sorted 5 days after infusion from KP-ROR1 lung tumor–bearing mice treated with either vehicle (PBS) or anti–PD-L1 checkpoint blockade immunotherapy.