Differential Proteomic Analysis of DEN-induced Hepatocellular Carcinoma in Male and Female Balb/c Mice Reveals Novel Sex-Specific Markers

BACKGROUND/AIM: Hepatocellular carcinoma (HCC) is one of the leading causes of hepatic malignancy with a higher prevalence in males compared to females; however, the distinct underlying mechanisms contributing to this disparity remain poorly understood. MATERIALS AND METHODS: In this study, we aimed to investigate comparative proteome profiling of a diethylnitrosamine (DEN) induced HCC model in male and female Balb/c mice. We extracted proteins from the liver tissue of DEN-treated male and female mice and their corresponding controls and subjected them to mass spectrometry and subsequent bioinformatics analyses. RESULTS: We identified 170 and 146 differentially expressed proteins (DEPs) in female and male mice, respectively. We identified chemical carcinogenesis, oxidative phosphorylation, actin binding and mitochondrial respiration as the shared pathways between the two groups. In addition, we identified distinct signaling pathways in DEN-treated male and female mice. Female mice showed enrichment in fatty acid biosynthesis, metabolism, degradation, and cytochrome P450 clusters. In contrast, in male mice, pathways were enriched in tryptophan metabolism, apoptotic execution phase and glutathione metabolism. Further, we identified the top ten genes ranked by highest maximal clique centrality, by protein-protein interaction analysis of differentially expressed proteins (DEPs) in both sexes. Of these hub genes, female mice showed up-regulation of previously unimplicated, NDUFA8 and ATP5H proteins which were associated with poor patient survival. On the other hand, in DEN-treated male mice up-regulation of RPS3 was associated with poor survival. CONCLUSION: In conclusion, our research provides sex-specific proteomic signatures in DEN-induced HCC. The identification of proteins associated with ribosomal subunits in males, and two previously unimplicated mitochondrial complex I proteins in females as prognostic markers suggests novel therapeutic targets that may inform sex-tailored treatment strategies for HCC.