Activation of NF-κB/ALDH1A1 Signaling Promotes Non-Mutational Resistance to EGFR-TKIs in Non-Small Cell Lung Cancer

This study investigates the molecular mechanisms underlying acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in EGFR-mutant non-small cell lung cancer (NSCLC). First, we utilized NSCLC cells, lung cancer organoids (LCOs), xenograft tumor, and patient-derived orthotopic xenografts tumor models to establish TKI resistance. Their stem cell-like properties, drug sensitivity, tumor formation in vitro and in vivo and transcriptomic profiles were analyzed. The differentially expressed genes related to TKI resistance and associated pathways were identified. Functional experiments, including gain- and loss-of-function assays, dual-luciferase reporter analysis, chromatin immunoprecipitation, Western blot, and RT-qPCR, revealed the roles of NF- κ B/ALDH1A1 signaling in acquired resistance. Compared to the parental cells and LCOs, the TKI-resistant R-PC9 and LCO/R exhibited increased RELA phosphorylation, higher frequency of ALDH1 + cells, enhanced sphere formation. ALDH1A1 over-expression promoted sphere formation and enhanced resistance to TKIs while ALDH1A1 or RELA silencing had opposite effects. Furthermore, RELA activation enhanced ALDH1A1 activity, and pharmacological inhibition of either NF- κ B or ALDH1A1 activity enhanced the sensitivity to TKIs both in vitro and in vivo. These findings indicated that aberrant activation of the NF- κ B/ALDH1A1 signaling promoted non-mutational resistance to EGFR-TKIs and provided a therapeutic target for EGFR-resistant NSCLC.