A Network pharmacology and experimental study of dihydroartemisinin in drug-resistant non-small cell lung cancer involving AURKA modulation

Abstract

Non-small cell lung cancer (NSCLC) remains the leading cause of cancer-related mortality globally, with acquired chemoresistance posing a significant therapeutic challenge. Artemisinin (DHA), a semisynthetic derivative of artemisinin, has been reported to exhibit antitumor activity; however, its molecular mechanism in drug-resistant NSCLC remains unclear. This study systematically examined whether DHA mitigates cisplatin resistance in NSCLC by modulating Aurora kinase A (AURKA) and subsequently regulating apoptosis and ferroptosis. An integrated network pharmacology analysis revealed 34 overlapping targets among DHA, NSCLC, ferroptosis, and apoptosis datasets, comprising 20 core targets, including STAT3, HIF1A, EGFR, and GSK3β. AURKA ranked 19th by degree centrality; it was prioritized for further investigation based on its established roles in chemoresistance and ferroptosis regulation. Molecular docking predicted a strong binding affinity between DHA and AURKA (binding energy: –8.1 kcal/mol), with hydrogen bond formation at the ALA-213 residue. Cellular thermal shift assay confirmed that DHA enhanced the thermal stability of AURKA, whereas 100-ns molecular dynamics simulations exhibited sustained conformational stability of the AURKA-DHA complex, characterized by persistent hydrogen bond interactions and favorable binding free energy calculations. Functionally, DHA inhibited A549/DDP cell proliferation in a concentration- and time-dependent manner (IC 50 : 53.14 μM at 24 h, 36.28 μM at 48 h, and 23.18 μM at 72 h), induced apoptosis as evidenced by BAX upregulation and BCL-2 downregulation, and suppressed migration and invasion. Simultaneously, DHA treatment triggered ferroptotic hallmarks, including intracellular reactive oxygen species accumulation, ferrous iron elevation, and downregulation of SLC7A11 and GPX4. Mechanistically, DHA diminished AURKA and NRF2 protein levels and decreased GSK3β phosphorylation at Ser9, indicating the involvement of the AURKA/GSK3β/NRF2 axis. Co-treatment with the AURKA inhibitor VX-680 demonstrated synergistic effects (combination index < 1), enhancing growth inhibition, apoptosis, and ferroptosis both in vitro and in A549/DDP xenograft models, accompanied by reduced AURKA and GPX4 expression in tumor tissues. Notably, no significant toxicity was detected in major organs or in liver/kidney function parameters after monotherapy or combination treatment. Collectively, these findings indicate that DHA may contribute to overcome cisplatin-resistant effects in NSCLC by modulating AURKA and the AURKA/GSK3β/NRF2 axis, concurrently inducing apoptosis and ferroptosis. The observed synergy between DHA and AURKA inhibition, together with a favorable safety profile, supports further investigation of this combination as a potential therapeutic strategy for chemoresistant NSCLC.