The Activity of EGFR CAR NK and CAR T Cells against EGFR Inhibitor-Resistant NSCLC and Drug-Tolerant Persister Cells.

PURPOSE: Patients with non-small cell lung cancer harboring EGFR mutations typically have significant clinical benefits from EGFR tyrosine kinase inhibitors (TKI) such as osimertinib. However, a residual population of drug-tolerant persister cells (DTPC) inevitably remains, which ultimately gives rise to fully drug-resistant cells (DRC). This study evaluates the activity of EGFR chimeric antigen receptor (CAR)-based therapies in this context. EXPERIMENTAL DESIGN: We developed EGFR CAR T and CAR NK cells and evaluated their antitumor activity against parental cells, DTPC, and DRC in vitro and in vivo. We investigated the mechanisms regulating the sensitivity of DTPC and DRC to CAR T or CAR NK cells, including NK-activating ligands, TGF-β signaling, and EGFR surface levels. Additionally, we developed strategies that included galunisertib treatment and the expression of a dominant-negative TGF-β receptor II in CAR NK cells. RESULTS: DTPC demonstrated increased sensitivity to both EGFR CAR T and CAR NK cells. DRC were relatively resistant to CAR T cells but more sensitive to CAR NK cells. DRC and DTPC had higher levels of natural cytotoxicity triggering receptor-3 and NKG2D ligands, which enhance the effectiveness of CAR NK cells. Elevated TGF-β levels in DRC impaired CAR function, but this was reversed by coexpression of galunisertib or dominant-negative TGF-β receptor II in CAR NK cells. Continued TKI treatment increased EGFR expression on DRC, possibly contributing to the improved killing activity seen with TKI/CAR combinations compared with CAR alone in TKI-resistant cells. CONCLUSIONS: EGFR-directed cellular therapies, particularly EGFR CAR NK cells, demonstrate activity against EGFR-mutant DTPC and DRC in vitro and in vivo, with enhanced activity observed when combined with EGFR TKI or TGF-β pathway blockade.