Effect of MitoCur-1 on development of hepatocellular carcinoma by targeting inhibition of ubiquitin-specific protease 14

Objective To investigate the effects of MitoCur-1 on the proliferation and migration of hepatocellular carcinoma through the inhibition of ubiquitin-specific protease 14 (USP14), providing a theoretical basis for the antitumor activity of MitoCur-1.

Methods HepG2 cells were cultured in vitro, and CCK-8 assays and colony formation assays were used to assess the effects of MitoCur-1 on cell proliferation. The scratch assay was employed to evaluate the impact of MitoCur-1 on cell migration. real-time quantitative PCR (RT-qPCR) and Western blot were performed to detect changes in both USP14 mRNA and protein expression levels in HepG2 cells after treatment with MitoCur-1. The CHX assay was conducted to examine the effect of MitoCur-1 on protein half-life. An animal model was established, and hematoxylin and eosin (HE) staining along with immunohistochemistry (IHC) were used to analyze the in vivo antitumor effects of MitoCur-1.

Results The cell viability of HepG2 cells decreased with increasing concentrations of MitoCur-1, with an IC₅₀ of 3.62 μmol/L. Compared to the control group, MitoCur-1 at concentrations of 0.2, 0.4, and 0.8 μmol/L significantly reduced the colony formation ability of HepG2 cells (all P < 0.05). After 48 hours of treatment with 1, 2, and 4 μmol/L MitoCur-1, the migratory ability of HepG2 cells was diminished (all P < 0.05). Additionally, treatment with 1, 2, and 4 μmol/L MitoCur-1 for 24 hours led to decreased mRNA and protein expression levels of USP14 in HepG2 cells compared to the control group (all P < 0.05). CHX assay results indicated that MitoCur-1 reduced the protein half-life of USP14 (P < 0.05). The experiments in vivo showed that the growth rate of tumor volume was slowed in the 15 mg/kg (in body weight) MitoCur-1 group of mice, resulting in a significantly lower tumor volume than that of the control group (P < 0.05). However, there was no statistically significant difference in the overall weight gain of mice between the treated and control groups (P > 0.05).

Conclusions MitoCur-1 treatments inhibited the expression of USP14, reduced the proliferation and migratory capabilities of HepG2 cells, induced apoptosis, promoted autophagy, and suppressed the development of hepatocellular carcinoma.