Integrative analysis in head and neck cancer reveals distinct role of miRNome and methylome as tumour epigenetic drivers

BackgroundHead and neck cancer is the sixth most common malignancy worldwide, with the relatively low 5-year survival rate, mainly because it is diagnosed at a late stage. Infection with HPV is well known etiology, which affects nature of these cancers and patients survival. Besides, it is considered that the main driving force for this type of cancer could be epigenetics. In this study we aimed to find potential epigenetic biomarkers, by integrating miRNome, methylome, and transcriptome analyses. MethodsFrom the fresh head and neck cancer tissue samples, we chose a group for miRNome, methylome and transcriptome profiling, in comparison to adequate control samples. Bioinformatics analyses are performed in R v4.2.2. Count normalization and group differential expression for mRNA and the previously obtained miRNA count data was performed with DESeq2 v1.36. Gene set enrichment analysis was performed and visualized using gProfiler2 v0.2.1 Identification of miRNA targets was performed by querying in miRTarBase using multiMiR v1.18.0. Annotation of CpG sites merging into islands was obtained from RnBeads.hg19 v1.28.0. package. For the integrative analysis we performed kmeans clustering using stats v4.2.2 package, using 8-12 clusters and nstart 100. ResultsWe found that transcriptome analysis divides samples into cancers and controls clusters, with no relation to HPV status or cancer anatomical location. Differently expressed genes (n=2781) were predominantly associated with signaling pathways of tumour progression. We identified a cluster of genes under control of the transcription factor E2F, that are significantly underexpressed in cancer tissue, as well as T cell immunity genes and genes related to regulation of transcription. Among overexpressed genes in tumours we found those that belong to cell cycle regulation and vasculature. A small number of genes were found significantly differently expressed in HPV positive versus HPV negative tumours (for example NEFH, ZFR2, TAF7L, ZNF541, and TYMS). ConclusionsIn this comprehensive study on an overlapping set of samples where the integration of miRNome, methylome and transcriptome analysis were performed for head and neck cancer, we demonstrated that the majority of genes were associated exclusively with miRNome or methylome and, to a lesser extent, under control of both epigenetic mechanisms.