NR2F6 deletion revives CAR-T cell function and induces antigen-agnostic immune memory in solid tumors

CAR-T cell therapy is effective in hematologic malignancies but remains challenging in solid tumors owing to antigen heterogeneity and tumor microenvironment-induced exhaustion. Here, gene editing of the nuclear receptor NR2F6 restores CAR-T cell functionality, sustaining a TCF1? progenitor-exhausted T cell phenotype, enhancing metabolic fitness, and preserving cytotoxic potency under chronic antigen exposure. In immunocompetent models, Nr2f6-deficient CAR-T cells suppress solid tumor growth and induce robust, polyclonal host antitumor responses, as shown by protection in tumor re-challenge experiments. Although infused CAR-T cells disappear within 2 weeks, durable tumor control coincides with epitope spreading and secondary immune responses, likely via dendritic and other antigen-presenting cell reactivation. Protection against antigen-negative tumors and transferable immunity reveal a dual mode of direct cytotoxicity followed by durable immune reprogramming. This broadened host immunity may offset immune escape driven by antigen loss and tumor heterogeneity, establishing NR2F6 inhibition as a promising CAR-T engineering strategy for durable, antigen-agnostic solid-tumor immunotherapy. Overall design: Bulk RNA sequencing: Transcriptional profiling of wildtype and Nr2f6 knockout CAR-T cells in co-culture with Panc02-EpCAM tumor cells at day 0, day 4 and day 8. Single cell RNA sequencing: single cell transcriptomic profiling and V(D)J analysis of either naive or Nr2f6 knock-out CAR-T treated survivor mice challenged with Panc02-EpCAM negative tumors.