IL-1α-driven TNF-α/NF-κB activation confers osimertinib resistance in EGFR-mutant NSCLC cells

Non-small-cell lung cancer (NSCLC) remains the leading cause of cancer-related deaths globally. While osimertinib, a third-generation EGFR tyrosine kinase inhibitor (TKI), has significantly improved outcomes for patients with EGFR-mutant NSCLC, its efficacy is frequently limited by both primary and acquired drug resistance. A key challenge lies in understanding the mechanisms of early resistance, particularly the role of cycling cancer persister cells (CPCs), a transient subpopulation that can evade targeted therapies. The specific biological characteristics and signaling pathways driving osimertinib resistance in these CPCs have been poorly understood, representing a critical gap in current treatment strategies.

Researchers established osimertinib-resistant cycling cancer persister cells (CPCs) from two distinct EGFR-mutant NSCLC cell lines, PC9 and HCC827. To identify the underlying molecular mechanisms, RNA sequencing (RNA-seq) was performed on these PC9-CPCs to map their unique transcriptional programs. Subsequent experiments involved treating PC9 parental cells with IL-1α to observe its effects on TNF-α and NF-κB p65 expression. Conversely, IL-1α or TNF-α signaling was inhibited in PC9-CPCs to assess the restoration of osimertinib sensitivity. Immunohistochemical analysis was also conducted on patient tumor specimens to correlate NF-κB expression with osimertinib resistance development.

RNA sequencing analyses revealed a significant upregulation of the tumor necrosis factor alpha (TNF-α)/nuclear factor-κB (NF-κB) signaling pathway in osimertinib-resistant PC9-CPCs. Among the genes analyzed, interleukin-1 alpha (IL-1α) was identified as the most significantly overexpressed gene within this pathway in the resistant cells. Experimental treatment of PC9 parental cells with IL-1α directly led to increased expression of both TNF-α and NF-κB p65. Conversely, targeted inhibition of IL-1α in PC9-CPCs resulted in a decrease in TNF-α expression.

Crucially, the inhibition of either IL-1α or TNF-α signaling successfully restored the sensitivity of these EGFR-mutant NSCLC cells to osimertinib, indicating a direct role for this axis in resistance. Immunohistochemical analysis further corroborated these in-vitro findings, showing that NF-κB expression was notably higher in tumor specimens from patients who had developed osimertinib resistance compared to their pre-treatment samples.