Adaptive targeting of lung cancer using the anti-P329G adapter CAR T-cell platform.

Abstract

e14514 Background: Chimeric antigen receptor (CAR) T cell therapy has transformed the treatment of hematologic malignancies but remains less effective in solid tumors such as lung cancer due to antigen heterogeneity, on-target/off-tumor toxicities, and immunosuppressive tumor microenvironments (TME). Adapter CAR systems offer a strategy to improve flexibility and toxicity control by separating antigen recognition from CAR signaling. Anti-P329G adapter CAR T cells recognize a P329G point mutation in the Fc region of engineered human IgG1 antibodies, permitting precise and reversible retargeting through P329G-Fc–mutated antibodies against multiple tumor antigens. This work investigates the anti-P329G CAR platform in lung cancer, emphasizing its performance against the clinically relevant and therapeutically validated target EGFR. Methods: Anti-P329G CAR T cells demonstrated strong, antibody-dependent in vitro activation and effector functions against EGFR-positive lung cancer cells and recombinant EGFR protein. CAR T cell activity required presence of the P329G-Fc–engineered antibody, confirming reversible and modular antigen control. To further explore controllability, antibody displacement experiments with mock binders were conducted to assess reversibility. In vivo, EGFR-redirected anti-P329G CAR T cells induced significant tumor regression and prolonged survival in lung cancer–bearing mice without systemic toxicity. In both TCS-PCLS and TD-PCLS, EGFR-directed anti-P329G CAR T cells effectively reduced EdU+ tumor proliferation and enhanced TUNEL+ apoptosis, while sparing adjacent nonmalignant tissue despite EGFR co-expression. Results: Anti-P329G CAR T cells showed strong, antibody-dependent in vitro activation and effector functions against EGFR-positive tumor cells and recombinant EGFR protein. Activity required the P329G-Fc–engineered antibody, confirming modular and reversible antigen control. To explore controllability, antibody displacement experiments with mock binders were performed. In vivo, EGFR-redirected anti-P329G CAR T cells induced significant tumor regression and prolonged survival in lung cancer–bearing mice without systemic toxicity. In both TCS-PCLS and TD-PCLS, EGFR-directed anti-P329G CAR T cells reduced EdU+ tumor proliferation and increased TUNEL+ apoptosis, while sparing nonmalignant lung tissue despite EGFR co-expression. Conclusions: The anti-P329G adapter CAR platform provides a tunable, specific, and reversible approach for EGFR-targeted immunotherapy in lung cancer. Demonstrated efficacy across in vitro, in vivo, and ex vivo models, together with feasibility testing of antibody displacement, highlights its translational potential to overcome key barriers of CAR T therapy in solid tumors while maintaining improved safety and controllability.