Abstract
BackgroundDNA methylation in miRNA genes has been reported as a mechanism that may cause dysregulation of mature miRNAs and consequently impact the gene expression. This mechanism is largely unstudied in papillary thyroid carcinomas (PTC).MethodsTo identify differentially methylated miRNA-encoding genes, we performed global methylation analysis (Illumina 450 K), integrative analysis (TCGA database), data confirmation (pyrosequencing and RT-qPCR), and functional assays.ResultsMethylation analysis revealed 27 differentially methylated miRNA genes. The integrative analyses pointed out miR-21 and miR-146b as potentially regulated by methylation (hypomethylation and increased expression). DNA methylation and expression patterns of miR-21 and miR-146b were confirmed as altered, as well as seven of 452 mRNAs targets were down-expressed. The combined methylation and expression levels of miR-21 and miR-146b showed potential to discriminate malignant from benign lesions (91–96% sensitivity and 96–97% specificity). An increased expression of miR-146b due to methylation loss was detected in the TPC1 cell line. The miRNA mimic transfection highlighted putative target mRNAs.ConclusionsThe increased expression of miR-21 and miR-146b due to loss of DNA methylation in PTC resulted in the disruption of the transcription machinery and biological pathways. These miRNAs are potential diagnostic biomarkers, and these findings provide support for future development of targeted therapies.
Highlights
DNA methylation in miRNA genes has been reported as a mechanism that may cause dysregulation of mature miRNAs and impact the gene expression
DNA methylation analysis comparing paired papillary thyroid carcinomas (PTC) (N = 50) and neoplastic thyroid tissue (NT) (N = 50) samples revealed 50 CpG probes (34 miRNA genes) differentially methylated, of which 86% (42 probes mapped in 27 miRNA genes) was confirmed using the The Cancer Genome Atlas (TCGA) database (Table 1)
A supervised hierarchical clustering analysis with all 42 probes revealed an enrichment of hypomethylation in both, our PTC cases and in the TCGA dataset (Additional file 1: Figure S1)
Summary
DNA methylation in miRNA genes has been reported as a mechanism that may cause dysregulation of mature miRNAs and impact the gene expression. This mechanism is largely unstudied in papillary thyroid carcinomas (PTC). Papillary thyroid carcinoma (PTC) is the most frequent thyroid malignant neoplasm and is responsible for the increased incidence of thyroid cancer worldwide [1, 2]. The main genetic alterations described in PTC are BRAF and RAS mutations and RET rearrangements [3]. A recent study conducted by our group identified DNA methylation alterations related to prognosis in well differentiated thyroid lesions [10]. This classifier was able to distinguish well-differentiated thyroid carcinomas of patients showing worse prognosis
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