Gymconopin C exhibits anti-non-small cell lung cancer effect by regulating miR-6777-5p/ADRB2 pathway to promote mitophagy

• Firstly identifies Gymconopin C-a compound isolated from Bletilla striata-as a potent anti-NSCLC agent suppressing cancer development. • First evidence that miR-6777-5p/ADRB2 axis regulates PINK/Parkin-mediated mitophagy in NSCLC pathogenesis. • Elucidates the mechnistic cascade: Gymconopin C → miR-6777-5p ↓ →ADRB2 ↑ →PINK/Parkin ↑ →triggers mitophagy → induces cancer growth arrest. • Validates therapeutic efficacy across in vitro, zebrafish xenograft and murine ectopic tumor models. • Proposes a novel therapeutic strategy targeting mitophagy via miRNA/mRNA modulation for NSCLC treatment. Non-small cell lung cancer (NSCLC) remains the leading cause of morbidity and mortality from malignant tumors in China with therapeutic limitations. Developing novel therapeutic agents and innovative treatment strategies is critical for advancing NSCLC management. Gymconopin C, a compound isolated from Bletilla striata, has demonstrated potential anti-NSCLC activity; However, the underlying mechanism remains elusive. This study aimed to decipher the anti-NSCLC mechanism of Gymconopin C by demonstrating its induction of PINK1/Parkin-mediated mitophagy via the miR-6777-5p/ADRB2 pathway, and to establish the fundamental regulatory role of miRNA/mRNA axis in NSCLC suppression. This study establishes a novel therapeutically targetable pathway and offers a mechanistically grounded candidate for NSCLC therapy. Integrated in vitro and in vivo strategies were used to elucidate the anti-NSCLC mechanism of Gymconopin C. Inhibitory effects on two NSCLC cell lines were assessed using CCK-8 proliferation, transwell migration and invasion assays. Subsequently, transmission electron microscopy and mitochondrial functional analyses were conducted to assess mitophagy. Transcriptomic profiling revealed dysregulation of the miR-6777-5p/ADRB2 axis, and gain/loss-of-function experiments were performed to investigate its functional role in mitophagy. Finally, the therapeutic efficacy and pathway modulation were evaluated in vivo using zebrafish xenograft and murine ectopic tumor models. Gymconopin C significantly inhibited the proliferation, migration and invasion of NSCLC cells; induced cell cycle arrest and enhanced apoptosis of A549 cells. Mechanistically, it modulated the miR-6777/ADRB2 axis to promote PINK/Parkin-mediated mitophagy, ultimately leading to NSCLC growth arrest in both in vivo and in vitro experiments. Gymconopin C exerts tumor-suppressive effects by activating PINK1/Parkin-mediated mitophagy via the miR-6777-5p/ADRB2 axis, highlighting its potential as a therapeutic agent for NSCLC.