Metal-dependent cell death in doxorubicin-induced cardiotoxicity and mitigation by luteolin: Network pharmacology and experimental verification

Doxorubicin (DOX) is a highly effective anthracycline chemotherapeutic agent whose clinical utility is limited by dose-dependent cardiotoxicity. Although oxidative stress and mitochondrial injury are established mechanisms, the contribution of regulated metal-dependent cell death (MCD) pathways remains incompletely defined. This study investigated the ferroptosis and cuproptosis role in doxorubicin-induced cardiotoxicity (DIC) and evaluated the protective effects of luteolin (LUT) and nanoliposomal LUT using an integrated network pharmacology and experimental validation approach in rats. Network pharmacology identified multitarget interactions linking LUT with oxidative stress, metal homeostasis, and cell death signaling pathways. In vivo, DOX administration induced marked cardiac dysfunction, elevated serum cardiac injury biomarkers, myocardial histopathological damage, and ultrastructural abnormalities. These changes were accompanied by significant cardiac iron and copper accumulation, increased malondialdehyde (MDA), depleted superoxide dismutase (SOD) and glutathione (GSH), reduced glutathione peroxidase-4 (GPX4) activity, and increased tumor protein p53 (TP53) expression. At the molecular level, DOX downregulated the ferroptosis-protective genes, SLC7A11 and SLC3A2, while upregulating the iron transport genes, transferrin receptor 1 (TFR1) and SLC39A14, together with the cuproptosis-related genes ferredoxin-1 (FDX1) and SLC31A1, while suppressing ATP7A. These changes were further supported by altered protein expression of SLC7A11, ATP7A, and TP53. LUT significantly ameliorated these functional, biochemical, molecular, and structural alterations, indicating suppression of both ferroptotic and cuproptotic signaling. Nanoliposomal LUT showed superior efficacy to free LUT across most assessed parameters. Collectively, these findings support the involvement of ferroptosis and cuproptosis related signaling in DIC and highlight LUT, particularly in nanoliposomal form, as a promising cardioprotective strategy against anthracycline toxicity.