Fig. 2: Inhibition of USP14 deubiquitinating activity sensitizes HCC cells and PDX models to radiation.

Colony formation assays were conducted on (A) USP14-KO Huh7 and (B) USP14-KO MHCC97H cells overexpressing either WT or C114A mutant USP14. Cells were subjected to 4 Gy IR or left untreated. Representative images and survival curves are provided. Experiments were conducted in triplicate, and data are expressed as mean ± SD. Statistical significance was assessed using one-way ANOVA for group comparisons. Colony formation assays in (C) Huh7 and (D) MHCC97H cells pre-treated with the USP14 inhibitor IU1-47 (5 μM; DMSO served as a vehicle control) for 12 h before IR exposure (0, 2, 4, 6, or 8 Gy). Representative images and survival curves are provided. Experiments were conducted in triplicate, data presented as mean ± SD. Statistical significance was evaluated by two-way ANOVA. The impact of IU1-47 on the proliferation of Huh7 (E) and MHCC97H (F) cells following IR treatment (0, 2, 4, 6, or 8 Gy). Experiments were carried out in triplicate and data are presented as mean ± SD. The IC₅₀ was calculated by normalizing cell viability to the radiation-only control group (without IU1-47), which was set as 100% survival. G–L Effect of IU1-47 combined with IR in PDX models. (G) PDX-1 and (J) PDX-3 models received IU1-47 (5 mg/kg/day for 10 sessions) and IR (2 Gy/day for 5 sessions every alternate day). G, J Representative tumor images. H, K Quantitative tumor volume analysis. I, L Kaplan–Meier survival analysis of mice bearing PDX (n = 6/group). Experiments were replicated in triplicate, and data are shown as mean ± SD. Statistical significance determined by two-way ANOVA. Results are exemplary of three independent studies.