Mounting evidence shows that genetic alterations in signaling pathways drive HCC tumorigenesis, highlighting the importance of molecular biomarkers in liver cancer detection. Although the mechanisms underlying HCC tumorigenesis are unclear, cellular and molecular changes are influential factors. Here, we first evaluated differentially expressed ceRNA and immune cells infiltration in normal vs tumor HCC tissues. We then developed nomograms for predicting HCC prognosis. The excellent concordance index and AUC value in the 2 nomograms may guide HCC survival prediction. K-M survival along with correlation analysis indicated that the ceRNA modulatory network of SNHG1 (lncRNA), HMMR (mRNA), hsa-miR-421 (miRNA), and Tregs may influence HCC progression.
It is estimated <2% of the human genome is protein-coding, suggesting that most human transcripts are non-coding . mRNAs, miRNAs, and lncRNAs are linked in an intricate crosstalk network by competing endogenous RNA . Interaction between miRNA, lncRNA, and mRNA, in ceRNA networks is extensively studied in various disorders, including lung, gastric, and gallbladder cancers [12,13]. However, the mechanisms of ceRNA networks in HCC oncogenesis are largely unclear. Here, we find that in a ceRNA network, SNHG1 (a lncRNA), may downregulate hsa-miR-421 and upregulate HMMR to promote HCC tumorigenesis. Hsa-miR-421 is reported to play a core role in malignant transformation, which is consistent with our findings [14,15]. HMMR is an important component of polo-like kinase 1 (PLK1)-dependent mitotic spindle localization cascade, which is essential for neurodevelopment, neonatal survival, and tumorigenesis . HMMR is upregulated in many cancers, such as lung cancer, glioblastoma, prostatic adenocarcinoma, and leukemia [17–20]. HMMR has been documented to enhance EMT (epithelial-mesenchymal transition) and carcinogenesis by activating TGF-β/Smad2 signaling in gastric cancer . Assessment of 1420 colorectal cancer tissues indicated that HMMR may be a better prognostic factor relative to tumor grade as well as vascular infiltration . How HMMR modulates cell cycle-linked gene expression has not been studied. HMMR antisense lncRNA HMMR-AS1 stabilizes HMMR mRNA and enhances the progress of lung adenocarcinoma, epithelial ovarian cancer, as well as glioblastoma [23–25]. HMMR negatively correlates with poor HCC prognosis, highlighting it as a potential indicator of HCC prognosis .
We also uncovered different proportions of immune cells in liver cancer. Naïve CD4 T-cells, resting mast cells, gamma delta T-cells, Tregs, and activated dendritic cells have been shown to be associated with HCC. A nomogram based on 5 immune cell types, built for overall survival prediction (consistency index = 0.66) may have high clinical value.
Tregs, which are CD25+, account for 5-10% of CD4+ T-cells and are one of the most widely studied immune cell types due to their inhibitory effects on tumor pathogenesis. Tregs repress the activation as well as the proliferation of CD4+ and CD8+ T-cells in vitro and in vivo using various mechanisms consisting of cell-cell contact along with secretion of immunosuppressive cytokines like TGF-β, IL-10, and IL-35 . Tregs are believed to be the most important determinant of hepatitis B prognosis . Infection with hepatitis B virus (HBV) is lonked to >60% of liver cancer cases in developing countries and <25% in developed countries. Our data show that Tregs are upregulated in HCC. It is suggested that Tregs depletion may slow HCC progression.
Levels of tumor infiltrating dendritic cells (DCs) correlate with clinical outcomes. CD14+CTLA4+ regulatory DCs have the same inhibitory effect as Tregs in HCC. These DCs inhibit T-cell response and upregulate PD-1 via IL-10 and CTLA4 .
High CD4+ T-cell levels also hinder liver cancer development. Anti-programmed death 1 antibody on the surface of CD4+ cells can demonstrate the clinical outcomes of HCC patients.
In the innate immune cells, total mast cells, monocytes, DCs, as well as neutrophils are considerably changed. Mast cells have key immune regulation functions, and the mechanism of cell inactivation in HCC remains unclear. IgE, a mast cell activator has been reported in HBV-linked HCC . Our data show a higher proportion of active mast cells in healthy livers and a higher proportion of inactive subtypes in advanced HCC. COX evaluation also established that the proportion of resting mast cells remarkably correlated with prognosis. Mast cells constitute the fastest immune cell responders that are particularly enriched in barrier sites. Their rapid release is mediated by factors like vasoactive amines, proteases, cytokines , and proteoglycans, from intracellular storage upon FCER1-bound IgE crosslinking . However, mast cell function in abnormal livers is still unclear and warrants further investigation.
Correlation analysis found that CD4 T-cells were linked to HMMR (R=-0.164, p=0.0016). CeRNA network correlation analysis and hypergeometry tests found that SNHG1 (a lncRNA), HMMR (a protein-coding RNA), and hsa-miR-421 (a miRNA) were remarkably associated (p=0.007847), suggesting that the interaction between hsa-miR-421, SNHG1, HMMR, and CD4 T-cells is associated with HCC development.
This study has several limitations. Firstly, multivariable survival assessment included only basic prognostic factors from TCGA and could not suggest other potential clinical factors like metastatic lesions status and patients’ performance status. Secondly, preponderant evidence suggests that HCC is a very heterogeneous tumor at genetic and molecular levels. Due to study limitations, gene expression data from GEO, SYMH, and ICGA cohorts was determined from one piece of HCC tissue from a single patient. Future studies should evaluate gene expression in several HCC specimens from a patient via single-cell whole-genome sequencing or RT-qPCR analysis to determine if the line map is a reliable and viable predictor of HCC overallsurvival markers. In addition, cooperation with other hospitals should be considered to recruit more patients and samples for gene model validation.