Novel miRNA genes deregulated by aberrant methylation in ovarian carcinoma are involved in metastasis.

Methylation of promoter CpG islands may suppress the function of miRNAs by inhibiting their expression. Our work analyzes the role of promoter methylation in altering the expression of 12 miRNAs associated with epithelial ovarian cancer (EOC): miR-124-3p, -125b-5p, -127-5p, -129-5p, -132-3p, -137, -148a-3p, -191-5p, -193a-5p, -203a, -339-3p, and -375. The role of methylation in the deregulation of these miRNAs has not been previously assessed in a representative set of EOC samples. Using 76 paired (tumor/matched normal) ovarian samples and methylation-specific PCR, we demonstrated significant aberrations in the methylation patterns of 11 miRNA genes and identified 8 novel hypermethylated miRNA genes (MIR-124-1, -124-2, -124-3, -127, -132, -137, -193A, and -339) as well as one hypomethylated miRNA gene (MIR-191). Quantitative PCR on a subset of 29 paired EOC samples allowed us to establish a strong correlation between methylation status and alterations in expression levels for all 12 miRNAs studied. These findings demonstrate the functional role of aberrant methylation of examined miRNA genes in EOC. Moreover, we showed a significant association of hypermethylation of 10 miRNA genes (MIR-124-2, -124-3, -125B-1, -127, -129-2, -137, -193A, -203A, -339, -375) with EOC metastasis to lymph nodes, peritoneum, and distant organs. Interestingly, MIR-203A and MIR-375 were hypermethylated only in disseminated ovarian tumors, implying that non-suppressor miR-203a and miR-375 have anti-metastatic properties. Hypermethylation of 10 miRNA genes in EOC metastases was validated using an additional sample set of 13 primary tumors and matched peritoneal metastases. Together, these results show the impact of aberrant methylation on deregulation of 12 miRNAs in EOC, the involvement of 10 hypermethylated miRNA genes in metastasis (including peritoneal macro-metastases), and suggest novel potential biomarkers.