Bioinformatic characterization and validation of miR-30a-5p and miR-139-5p as diagnostic and prognostic biomarkers of oral cancer

Abstract

Oral cancer is the most prevalent type of head and neck cancer, characterized by rising incidence, high relapse occurrence and low survival. There is a high demand for the identification of novel and sensitive diagnostic and prognostic molecular biomarkers of oral cancer. MicroRNAs (miRNAs) are considered as promising candidates. Previously conducted meta-analysis on high throughput miRNA profiling in oral cancer emphasized consistent down-regulation of miR-30a-5p and miR-139-5p. We aimed to validate these findings in oral cancer clinical samples and to test the diagnostic and prognostic potential of these miRNAs. Bioinformatical prediction of investigated miRNAs target genes was executed through the intersection of multiple miRNA target prediction algorithms. Enrichment analysis of miRNA target genes was performed using miRPathDB V2.0 software on GO:BP database. RNA was isolated from oral cancer and adjacent noncancerous tissue from 30 patients by miRVana kit. cDNA was synthesized by TaqMan microRNA reverse transcription kit. TaqMan gene expression kit was used for relative quantification of miRNAs normalized to RNU6B. Relative expression was calculated by comparative Ct method. The diagnostic potential was estimated by the ROC curve analysis. Expression of miR-30a-5p and miR-139-5p was dichotomized as high or low by the median. Association with clinicopathological features and miRNAs expression was calculated by χ2 test. Bioinformatical analysis demonstrated both enrichment of miR-30a-5p and miR-139-5p targets in biological processes associated with cancer pathology and complementarity in regulation of these processes thus ensuring non-redundant regulation. Both miRNA were significantly down-regulated in oral cancer compared to non-cancerous tissue (Wilcoxon test: p=0.0004, p=0.001, respectively). According to the ROC curve analysis, both miRNAs might be used as potential tools for discrimination between cancerous and non-cancerous tissue (miR-30a-5p AUC=0.677, p=0.019; miR-139-5p AUC=0.656, p=0.038). There was a significant correlation between the expression of miR-30a-5p and miR-139-5p in cancerous tissue (Spearman rank test r=0.901, p<0.0001). None of the analyzed miRNAs was associated with stage, nodal status, tumour size and overall survival (p>0.05) which indicates that miR-30a-5p and miR-139-5p can’t be used as prognostic biomarkers of oral cancer. Our results suggest that miR-30a-5p and miR-139-5p might be used as good diagnostic biomarkers for discrimination between oral cancer and non-cancerous tissue.