Targeting the TRIM14/USP14 axis enhances radiotherapy efficacy by inducing GPX4 degradation and disrupting ferroptotic defense in HCC.

Radiation resistance constitutes a formidable impediment in the treatment paradigm for hepatocellular carcinoma (HCC). Deubiquitinases (DUBs) exhibit notable efficacy in modulating cellular responses to stress and exogenous interventions, endowed with the critical trait of being targetable, thus facilitating the execution of precise therapeutic strategies. Here, we demonstrate that broad-spectrum inhibition of thiol hydrolase-type DUBs markedly augments radiotherapy sensitivity in HCC cells. Based on this, via CRISPR-based screening, we identified USP14 as the principal DUB orchestrating radioresistance. Ferroptosis emerged as a pivotal form of radiation-induced cell death, with our study singularly illustrating that USP14 is instrumental in directing cellular defenses against ferroptosis via the targeting and stabilization of glutathione peroxidase (GPX4). Mechanistically, we found that radiation triggers the assembly of Tripartite motif-containing protein 14 (TRIM14) at the GPX4 locus, subsequently recruiting USP14. The TRIM14/USP14 complex facilitates the excision of pronounced K48-linked polyubiquitination at lysine residues 48 or 118 on GPX4, thereby preserving GPX4's structural integrity and antioxidative function to counteract ferroptosis. Intriguingly, TRIM14-mediated GPX4 stabilization is further amplified in radioresistant HCC, and subsequent radiation enables USP14-dependent blockade of GPX4 degradation. Consequently, pharmacological inhibition of USP14 substantially increases the susceptibility of HCC cells, thereby sensitizing patient-derived xenograft (PDX) tumors to radiotherapy. Concurrently, we explored the abscopal effect of radiotherapy and revealed that targeting USP14-enhanced ferroptosis augments antitumor immune responses post-radiation, suggesting a strategy to sustain therapeutic efficacy. In conclusion, our study uncovers the TRIM14/USP14 axis as a critical suppressor of radiation-induced ferroptosis and an actionable target to overcome radioresistance in HCC. These findings provide mechanistic insights and a translational framework for improving radiotherapy outcomes.