CRISPRa screen unveils noncanonical NF-κB to boost the antitumor activity of TCR-based CAR-T cells.

Engineering autologous T cells for adoptive therapies has revolutionized cancer treatment, yet limited persistence remains a major barrier to robust clinical responses and durable disease control. Although costimulation with 4-1BB markedly improves CAR-T-cell persistence, no comparably effective approach has been identified to enhance persistence in T-cell therapies that rely on native TCR signaling, including conventional TCR-T cells and synthetic T-cell receptor and antigen receptor (STAR) T cells. To identify intrinsic persistence regulators in a TCR-based context, we performed an in vivo CRISPR activation screen in STAR-T cells and identified the noncanonical NF-κB axis, with RELB as the top hit. Direct RELB overexpression, however, exhibited dose-dependent cytotoxicity, motivating a translational alternative: overexpression of p52, the ncNF-κB transcription partner of RELB. Under prolonged antigen exposure, p52 overexpression sustained proliferation, preserved effector functions, and increased the intratumoral trafficking potential, particularly in CD8⁺ T cells, thereby yielding superior tumor control. Mechanistically, transcriptomic profiling revealed coordinated upregulation of cell cycle programs and chemokine receptors, along with downregulation of apoptosis and exhaustion-associated modules. Moreover, p52 overexpression restored RELB nuclear translocation in the context of PD-1/PD-L1 engagement, indicating partial resistance to checkpoint-mediated suppression and sustained ncNF-κB activity in PD-L1hi immunosuppressive environments. Collectively, our findings identify p52 as an antigen-dependent enhancer of ncNF-κB, conferring durable persistence and checkpoint resilience on TCR-based T cells and providing a feasible strategy to improve therapeutic effects in solid tumors.