Hemopexin mitigates doxorubicin cardiotoxicity by preserving cardiac metabolism while maintaining antitumor efficacy in a mouse model of breast cancer

Introduction Doxorubicin (DOX) is a cornerstone of anticancer therapy, but its use is limited by dose-dependent cardiotoxicity associated with mitochondrial dysfunction and oxidative stress. Hemopexin (HPX) has recently been identified as a modulator of anthracycline-induced cardiac toxicity in patients and preclinical models (Liu et al., 2022). Using a murine model of DOX-treated breast cancer, we investigated whether HPX modulates mitochondrial metabolism and function in both the heart and the tumor during chemotherapy. Methods Ten-week-old female C57BL/6 wild-type (WT) mice were subcutaneously injected with 2×10⁵ EO771 cells. After 10 days of tumor growth, mice received i.p. doxorubicin (DOX; 3 mg/kg every 48 h for 14 days), with or without i.p. hemopexin supplementation (700 µg every 48 h for 14 days). Tumor growth was monitored every two days and cardiac function was assessed by echocardiography at baseline and 4 weeks after treatment initiation. Cardiac RNA sequencing was performed at 4 weeks in WT mice (DOX or DOX+HPX groups) and in HPX-deficient (HPX KO) mice following saline or DOX. Fatty acid oxidation, glycolytic enzyme activity, electron transport chain function, ALAS1 activity and lipid peroxidation were assessed in cardiac and tumor tissues at the end of the experiment. Mitochondrial respiration was evaluated in neonatal mouse ventricular cardiomyocytes (NMVMs) using Seahorse analysis following 3 h of DOX exposure. Results Unexpectedly, HPX supplementation enhanced DOX-induced tumor cell death in tumor-bearing mice while concomitantly improving cardiac function and attenuating chemotherapy-induced cardiotoxicity. Cardiac transcriptomic analysis revealed that HPX supplementation upregulated genes involved in substrate metabolism and mitochondrial function while downregulating pro-inflammatory pathways, compared with DOX alone. Conversely, DOX-treated HPX-deficient mice displayed a more pronounced suppression of mitochondrial and metabolic gene programs, corroborated by reduced levels of key mitophagy and mitochondrial proteins. Consistently, HPX supplementation improved mitochondrial respiration in NMVMs under metabolic stress and rescued DOX-induced metabolic dysfunction. In vivo, HPX restored fatty acid utilization, reduced glycolytic reliance and improved electron transport chain activity and ATP production in chronically DOX-treated hearts, potentially through the rescue of ALAS1 activity and heme biosynthesis. Notably, HPX supplementation also normalized tumor mitochondrial metabolism and reduced aberrant glycolysis, providing a potential mechanistic basis for its anti-tumor effects. Conclusions Hemopexin supplementation restores mitochondrial metabolism and cardiac function during doxorubicin treatment while preserving, and enhancing, antitumor efficacy. Therefore, these data identify HPX as a promising cardioprotective strategy to improve the therapeutic index of anthracycline chemotherapy.