Activation of the NF-κB/ALDH1A1 signaling promotes non-mutational resistance to EGFR-TKIs in non-small cell lung cancer
Acquired resistance to tyrosine kinase inhibitors (TKIs) remains a major clinical challenge in the treatment of EGFR-mutant non-small cell lung cancer (NSCLC). This study established TKI-resistant variants by integrating cell lines, lung cancer organoids (LCOs), and in vivo models, revealing the pivotal role of the NF-κB/ALDH1A1 signaling in mediating non-mutational TKI resistance. Resistant cells exhibited elevated RELA phosphorylation, enhanced ALDH1A1 expression and enzymatic activity, and stem-like properties. Mechanistically, NF-κB activation occurred as an early response to TKI exposure and promoted ALDH1A1 transcription via RELA. In turn, ALDH1A1 contributed to the sustained activation of NF-κB signaling, forming a self-reinforcing positive feedback loop. Genetic ALDH1A1 or RELA silencing reversed the resistant phenotype. Pharmacologically, treatment with an EGFR-TKI and the ALDH1A1 inhibitor disulfiram or the NF-κB-targeting agent EGCG synergistically restored the antitumor efficacy of TKIs both in vitro and in vivo. These findings establish the NF-κB/ALDH1A1 signaling as a key non-genetic mechanism of acquired EGFR-TKI resistance and provide a rational combination strategy to overcome it. Catalysis-dependent NF-κB–ALDH1A1 feedback loop drives non-mutational EGFR-TKI resistance. EGFR-TKI treatment suppresses EGFR signaling but rapidly induces NF-κB activation as an early event under TKI stress in EGFR-mutant NSCLC cells. Activated NF-κB (p-RELA) drives ALDH1A1 transcription, while ALDH1A1 enzymatic activity reinforces NF-κB signaling via IKKβ phosphorylation, forming a catalysis-dependent positive feedback loop that sustains stemness and promotes non-mutational TKI resistance. Disruption of this axis by inhibiting ALDH1A1 (DSF) or NF-κB (EGCG) restores TKI sensitivity. Catalysis-dependent NF-κB–ALDH1A1 feedback loop drives non-mutational EGFR-TKI resistance. EGFR-TKI treatment suppresses EGFR signaling but rapidly induces NF-κB activation as an early event under TKI stress in EGFR-mutant NSCLC cells. Activated NF-κB (p-RELA) drives ALDH1A1 transcription, while ALDH1A1 enzymatic activity reinforces NF-κB signaling via IKKβ phosphorylation, forming a catalysis-dependent positive feedback loop that sustains stemness and promotes non-mutational TKI resistance. Disruption of this axis by inhibiting ALDH1A1 (DSF) or NF-κB (EGCG) restores TKI sensitivity.