Hepatic proton conductance as a bioenergetic vulnerability and therapeutic target in MASH-associated HCC

Metabolic dysfunction-associated steatohepatitis (MASH) is the fastest growing cause of hepatocellular carcinoma (HCC), yet the metabolic dependencies underlying MASH-HCC remain poorly defined. Here, we identify mitochondrial proton conductance as a central determinant of HCC progression and therapeutic vulnerability. Human HCC exhibited increased mitochondrial efficiency and glycolytic flux with reduced oxidative flux, consistent with large-scale HCC dependency datasets showing strong enrichment for downregulated mitochondrial pathways. To functionally interrogate this bioenergetic liability, we performed an unbiased PRISM barcoded-cell-line screen using the mitochondrial uncoupler BAM15. Lineage-level analysis revealed broad cytotoxicity with marked enrichment of sensitivity in HCC, which was validated in HepG2 cells, where BAM15 demonstrated greater potency than sorafenib, a frontline therapy for advanced HCC. We next evaluated mitochondrial uncoupling in vivo using two MASH-HCC models: (1) high-fat, fructose, and cholesterol diet with thermoneutral housing plus low-dose DEN, and (2) cocoa-butter diet plus DEN, each recapitulating metabolic injury and tumor development. In both models, BAM15 significantly reduced tumor burden and improved liver function. These effects persisted in PPARalpha-null mice, indicating that uncoupling exerts its efficacy through direct disruption of mitochondrial efficiency rather than secondary activation of fatty acid oxidation. To assess translational relevance, we tested the long-acting liver-targeted mitochondrial uncoupler TLC-1180, which similarly decreased tumor burden and improved hepatic function. Finally, liver-specific deletion of Ucp2 reduced hepatic proton conductance, increased tumorigenesis, and impaired liver function, establishing a mechanistic link between diminished uncoupling capacity and enhanced carcinogenic progression. Together, these data demonstrate that suppressed proton conductance is a fundamental metabolic adaptation that promotes HCC growth, and that enforced mitochondrial uncoupling directly counteracts this program. These findings position hepatic proton conductivity as a mechanistically grounded therapeutic target and support mitochondrial uncoupling, particularly liver-directed approaches, as a rational strategy for treating MASH-HCC.