Background: Sorafenib exerts its anti-tumor effects partly by inducing ferroptosis. However, the upstream regulatory mechanisms governing this process, particularly the role of eukaryotic translation initiation factor 2α (eIF2α) phosphorylation, remain largely unclear in renal cell carcinoma (RCC). Therefore, this study aimed to investigate the role and regulatory mechanism of eIF2α phosphorylation in sorafenib-induced ferroptosis in RCC.

Methods: To determine the optimal concentration, RCC cells were treated with sorafenib at doses of 0, 2.5, 5, 10, and 20 μM. Subsequently, sorafenib (10 μM) was administered to RCC cells either alone or in combination with ferrostatin-1 (Fer-1, 5 μM) or Isrib (200 nM). Lipid peroxidation, cell viability, apoptosis, and intracellular Fe²⁺ levels were assessed. In vivo BALB/c nude mice bearing orthotopic RCC tumors received intraperitoneal injections of sorafenib (20 mg/kg) or Isrib (2.5 mg/kg). Tumor volume and survival were recorded, while ferroptosis-related proteins, p-EIF2α/EIF2α ratios, and tumor lipid peroxidation (4-hydroxynonenal, 4-HNE) were evaluated.

Results: Sorafenib significantly reduced cell viability, increased apoptosis and lipid peroxidation, downregulated glutathione peroxidase 4 (GPX4), and elevated Fe²⁺ levels and acyl-CoA synthetase long-chain family member 4 (ACSL4) expression (p < 0.05). Fer-1 reduced the effects of sorafenib (p < 0.05), whereas Isrib enhanced them (p < 0.05). Fer-1 further promoted sorafenib-induced phosphorylation of eIF2α (p < 0.01), while Isrib inhibited this regulation (p < 0.001). In vitro experiments also confirmed that inhibition of eIF2α phosphorylation enhanced the anti-tumor effect of sorafenib (p < 0.05).

Conclusion: This study reveals eIF2α phosphorylation as a pivotal regulator of sorafenib-induced ferroptosis in RCC. Specifically, inhibition of eIF2α phosphorylation enhances sorafenib-mediated ferroptosis by modulating key ferroptosis-associated proteins and lipid peroxidation, thereby improving sorafenib sensitivity and anti-tumor efficacy in RCC. Targeting eIF2α phosphorylation thus represents a potential strategy to optimize sorafenib-based RCC therapy, based on these new insights into the drug's molecular mechanism of action.