Expression profiles and prognostic significance of RNA N6-methyladenosine-related genes in patients with hepatocellular carcinoma: evidence from independent datasets

Yu Zhou,1 Zi Yin,1 Baohua Hou,1 Min Yu,1 Ruiwan Chen,2 Haosheng Jin,1 Zhixiang Jian11Department of General Surgery, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong, People’s Republic of China; 2Department of Radiation Oncology, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong, People’s Republic of ChinaBackground: N6-methyladenosine (m6A) is the most prevalent modification of mammalian RNA. Emerging evidence suggest that m6A has critical roles in multiple biological activities, but little is known about its roles in cancer pathogenesis. Herein, we report the expression profiles and prognostic relevance of twelve m6A-related genes in hepatocellular carcinoma (HCC) by analyzing four independent datasets.Materials and methods: RNA levels of twelve m6A-related genes were detected in samples of 162 HCC patients who underwent curative resection (the Guangdong General Hospital dataset). We additionally analyzed the expression profiles of m6A-related genes in The Cancer Genome Atlas liver HCC dataset and two Gene Expression Omnibus datasets (GSE14520, GSE63898). Prognostic value of genes was evaluated by Kaplan–Meier curves of overall survival (OS) with the log-rank test and multivariate Cox regression analysis. Gene set enrichment analysis (GSEA) was conducted to identify associated KEGG pathways.Results: Five genes (METTL3, YTHDF1, YTHDF2, YTHDF3, and EIF3) showed consistent upregulation in all four datasets. Abnormal expressions of either METTL3 or YTHDF1 but not the other ten genes were associated with OS. Protein expression of METTL3 and YTHDF1 were confirmed in HCC tissues by immunohistochemical staining. Multivariate Cox regression analysis confirmed the independent predictive value of both METTL3 and YTHDF1 on OS. We further divided patients into three groups based on the median expression values of METTL3 and YTHDF1. In all datasets, the low METTL3/low YTHDF1 group showed a consistent better prognosis than other groups. GSEA revealed that both METTL3 and YTHDF1 regulate HCC cell cycle, RNA splicing, DNA replication, base excision repair, and RNA degradation.Conclusion: Both METTL3 and YTHDF1 were upregulated in HCC, and they were independent poor prognostic factors. Combination of METTL3 and YTHDF1 can be regarded as the biological marker that reflect malignant degree and evaluate prognosis in HCC.Keywords: hepatocellular carcinoma, N6-methyladenosine, m6A, prognosi