Magnolin overcomes EGFR TKI resistance in NSCLC by modulation of NDRG1-NRG2-HECW1 pathway

Resistance of non-small cell lung cancer (NSCLC) to EGFR tyrosine kinase inhibitors (TKIs) limits the efficacy and leads to disease progression with mechanisms such as activation of autophagy in tumor cells, but the current therapeutic strategies are unable to intervene in this mechanism. Magnolin (Mag), a naturally derived compound, has garnered significant interest due to its potential antitumor properties. Through virtual screening methods, Mag was identified as a compound with potential to regulate molecular pathways closely related to drug resistance mechanisms. In this study, we investigated the ability of Mag to enhance EGFR TKI efficacy in resistant NSCLC. Afatinib-resistant cell line (HCC827AR) was established by continuously exposing HCC827 cells to afatinib (4 µM) for 6 months. Medium containing 4 µM afatinib was refreshed every 48 h. By conducting RNA sequencing (RNA-seq) and exome sequencing on HCC827AR cells, NRG2 was identified as a core-enriched gene. We demonstrated that Mag directly bound to the TYR112 residue of NDRG1, stabilizing its expression and preventing its degradation. This interaction upregulated NDRG1, which in turn promoted its interaction with the E3 ubiquitin ligase HECW1, facilitating the ubiquitination and degradation of NRG2 at lysine 223 (K223). By targeting the NDRG1-NRG2-HECW1 pathway, Mag uniquely inhibited autophagy and restored the sensitivity of HCC827AR cells to EGFR TKIs, thereby reversing resistance. In vivo, the combined treatment with Mag (30 mg· kg⁻¹ ·d⁻¹, i.g.) and Afa (10 mg· kg⁻¹ ·d⁻¹, i.g.) significantly reduced tumor growth in patient-derived xenografts without inducing major toxicity. This study unravels the intricate role of NDRG1 in modulating NRG2 via HECW1. The results not only illuminate Mag’s promising potential as an adjunctive therapy to surmount EGFR TKI resistance, but also underscore the significant therapeutic potential of targeting the NDRG1-NRG2-HECW1 pathway as a novel strategy to reverse EGFR TKI resistance in NSCLC.