HSPA6 Confers Lenvatinib Resistance in Hepatocellular Carcinoma by Promoting Phase Separation-Mediated TXNRD1 Stabilization to Suppress Ferroptosis

Abstract The development of therapy resistance compromises the long-term efficacy of lenvatinib in advanced hepatocellular carcinoma (HCC), highlighting the need to characterize the molecular drivers of resistance. Through integrated multi-omics analysis combining genome-wide CRISPR screening, transcriptomics, and proteomics, we identified HSPA6 as a critical driver of lenvatinib resistance. HSPA6 was consistently upregulated in resistant cell lines and patient tumors, and high expression correlated with poor treatment response and survival. HSPA6 recruited the deubiquitinase USP9X to stabilize the antioxidant enzyme TXNRD1, thereby suppressing lenvatinib-induced ferroptosis. Furthermore, HSPA6 underwent drug-enhanced liquid–liquid phase separation through its IDR1 domain, facilitating the assembly of biomolecular condensates that reinforce TXNRD1 stability and ferroptosis resistance. Leveraging these insights, canagliflozin, an FDA-approved SGLT2 inhibitor, was repurposed as a direct HSPA6-targeting compound that disrupted this resistance axis, restored ferroptosis sensitivity, and synergized with lenvatinib in patient-derived models. This work unveils a dynamic, condensate-driven mechanism of drug resistance and offers a readily translatable strategy to overcome lenvatinib resistance in HCC.