Identification of hub methylated-CpG sites and associated genes in oral squamous cell carcinoma

Abstract

To improve personalized diagnosis and prognosis for oral squamous cell carcinoma (OSCC) by identification of hub methylated-CpG sites and associated genes, weighted gene comethylation network analysis (WGCNA) was performed to examine and identify hub modules and CpG sites correlated with OSCC. Here, WGCNA modeling yielded blue and brown comethylation modules that were significantly associated with OSCC status. Following screening of the differentially expressed genes (DEGs) from gene expression microarrays and differentially methylated-CpG sites (DCGs), integrated multiomics analysis of the DEGs, DCGs, and hub CpG sites from the modules was performed to investigate their correlations. Expression levels of 16 CpG sites-associated genes were negatively correlated with methylation patterns of promoter. Moreover, Kaplan-Meier survival analysis of the hub CpG sites and associated genes was carried out using 2 public databases, MethSurv and GEPIA. Only 5 genes, ACTA1, ACTN2, OSR1, SYNGR1, and ZNF677, had significant overall survival using GEPIA. Hypermethylated-CpG sites ACTN2-cg21376883 and OSR1-cg06509239 were found to be associated with poor survival by MethSurv. Methylation status of specific site and expression levels of associated genes were determined using clinical samples by quantitative methylation-specific PCR and real-time PCR. Pearson's correlation analysis showed that methylation levels of cg06509239 and cg18335068 were negatively related to OSR1 and ZNF677 expression levels, respectively. Our classification schema using multiomics analysis represents a screening framework for identification of hub CpG sites and associated genes.