Synergistic Promotion of Ferroptosis in Cardiomyocytes by Tumor Cell Secretory Factors and PD‐1 Blockade

Immune checkpoint inhibitors (ICIs) targeting PD-1 frequently cause severe cardiotoxicity, yet the mechanisms involving tumor-host crosstalk remain poorly defined. Emerging evidence implicates ferroptosis, an iron-dependent regulated cell death, in cardiac injury during cancer immunotherapy. Primary neonatal murine ventricular cardiomyocytes (NMVCs) were exposed to lung cancer-derived tumor-conditioned media (TCM; tumor-conditioned media) enriched with exosomes, anti-PD-1 antibody, or both. Ferroptotic markers, mitochondrial bioenergetics, inflammatory signaling, and exosomal microRNA cargo were systematically evaluated using standardized biochemical assays, quantitative PCR, and targeted pharmacological rescue experiments. TCM primed cardiomyocytes for ferroptosis by downregulating the GPX4/SLC7A11/Nrf2 antioxidant axis, depleting glutathione, and elevating labile iron and lipid peroxidation. Anti-PD-1 treatment alone induced minimal oxidative stress but synergized with TCM to amplify ferroptosis > 4.7-fold, triggering profound mitochondrial depolarization and upregulating pro-ferroptotic enzymes ACSL4/LPCAT3. Tumor-derived exosomal miR-214-3p and miR-23a-3p directly suppressed GPX4 and Nrf2, mediating cellular vulnerability. The combined insult hyperactivated NF-κB and STING pathways in a mitochondria-dependent manner, markedly increasing pro-inflammatory cytokine secretion. Co-treatment with ferrostatin-1, deferoxamine, or Nrf2 activators effectively reversed ferroptotic markers and restored mitochondrial function. Tumor-derived exosomal factors and PD-1 blockade synergistically drive ferroptotic cardiomyocyte death by crippling antioxidant defenses and amplifying inflammatory signaling. Targeting ferroptosis or its upstream regulators presents a promising cardioprotective strategy for patients receiving cancer immunotherapy.