Genetic Alterations and Transcriptional Expression of m(6)A RNA Methylation Regulators Drive a Malignant Phenotype and Have Clinical Prognostic Impact in Hepatocellular Carcinoma

Background: N6-methyladenosine (m(6)A) RNA methylation, associated with cancer initiation and progression, is dynamically regulated by the m(6)A RNA regulators. However, its role in liver carcinogenesis is poorly understood. Methods: Three hundred seventy-one hepatocellular carcinoma (HCC) patients from The Cancer Genome Atlas database with sequencing and copy number variations/mutations data were included. Survival analysis was performed using Cox regression model. We performed gene set enrichment analysis to explore the functions associated with different HCC groups. Finally, we used a machine-learning model on selected regulators for developing a risk signature (m(6)Ascore) The prognostic value of m(6)Ascore was finally validated in another two GEO datasets. Results: We demonstrated that 11 m(6)A RNA regulators are significantly differentially expressed among 371 HCC patients stratified by clinicopathological features (P<0.001). We then identified two distinct HCC clusters by applying consensus clustering to m(6)A RNA regulators. Compared with the cluster2 subgroup, the cluster1 subgroup correlates with poorer prognosis (P < 0.001). Moreover, the cell cycle, splicesome and notch signaling pathway are significantly enriched in the cluster1 subgroup. We further derived m(6)Ascore, using four m(6)A regulators, predicting HCC prognosis well at three (AUC = 0.7) or 5 years (AUC=0.7) in validation. The prognostic value of m(6)Ascore also was validated successfully in two GEO datasets (P < 0.05). Finally, we discovered that mutations and copy number variations of m(6)A regulators, conferring worse survival, are strongly associated with TP53 mutations in HCC. Conclusions: We find a significant relationship between the alterations and different expressions causing increased m(6)A level and worse survival, especially in TP53-mutated HCC patients. Genetic alterations of m(6)A genes might cooperate with TP53 and its regulator targets in the HCC pathogenesis. Our m(6)Ascore may be applied in the clinical trials for patient stratification in HCC.