〔Abstract〕 Objective To investigate the inhibitory effect of Elesclomol(ES)+Cu2+on the proliferation of human hepatoma cell lines PLC/PRF/5 and Huh-7 and its potential to induce Cuproptosis. Methods Human hepatoma cell lines PLC/PRF/5 and Huh-7 cells were Culturedin vitro. ES solution, Cu2+solution and copper chelating agent ammonium tetrathiomolybdate VI(ATTM) solution was treated separately or in combination. The effect of ES+Cu2+on the survival rate of human hepatoma cell lines PLC/PRF/5 and Huh-7 cells and the effect of ES+Cu2+on the survival rate after pretreatment with copper chelating agent ATTM were evaluated using CCK-8 kit. The cell death induced by ES+Cu2+was detected by flow cytometry and the changes of ES+Cu2+after pretreatment with copper chelating agent ATTM. The expression of Cuproptosis related proteins ATPase copper transporting beta(ATP7B),ferredoxin 1(FDX1), dihydrolipoamide s-acetyltransferase(DLAT) and superoxide dismutase 1(SOD1)were detected by Western blot. The effect of ES+Cu2+on cell proliferation and the reverse effect after ATTM pretreatment was detected by cell scratch assay. Results The toxicity of ES+Cu2+to human hepatocellular carcinoma cell lines PLC/PRF/5 and Huh-7 was significantly dose-dependent(P<0.05). Compared with the control group, the combined application of ES and Cu2+had a more significant inhibitory effect on hepatocellular carcinoma cells than ES or Cu2+alone(P<0.05), and copper chelating agent ATTM could reverse the inhibitory effect of ES+Cu2+on hepatocellular carcinoma cells(P<0.05). Flow cytometry results showed that compared with the control group, the proportion of cell death in PLC/PRF/5 and Huh-7 cells treated with ES+Cu2+increased, while the proportion of cell death decreased after ATTM intervention(P<0.05). The results of cell scratch test showed that the migration ability of PLC/PRF/5 and Huh-7 cells was decreased after ES+Cu2+treatment, however, the addition of ATTM reversed the inhibitory effect of ES+Cu2+on cell migration(P<0.05). Compared with the control group, the expression levels of copper death related proteins ATP7B, FDX1, DLAT and SOD1 decreased after ES+Cu2+treatment, but the addition of ATTM reversed the expression trend of these proteins(P<0.05). Conclusion The combination of ES and Cu2+can effectively inhibit the proliferation and migration of PLC/PRF/5 and Huh-7 of hepatocellular carcinoma cells, and induce Cuproptosis, which provides a new strategy for the treatment of hepatocellular carcinoma.