Demethylation of miR-495 inhibits cell proliferation, migration and promotes apoptosis by targeting STAT-3 in breast cancer

Oncol Rep. 2017 Jun;37(6):3581-3589. doi: 10.3892/or.2017.5621. Epub 2017 May 3.


In breast cancer (BC), silencing of miRNA genes due to miRNA gene promoter methylation are the important mechanisms directly contributing to tumorigenesis and tumor progression. miRNA-495 (miR-495) has been reported to be a tumor suppressor gene in various cancers, but its role and regulation in BC remains unclear. In the present study, the level of miR-495 was inversely correlated with the expression of STAT-3 in BC tissues and cell lines. miR-495 can directly target 3'-UTR of STAT-3 mRNA and thereby decrease the expression of STAT-3 in MCF-7 and HCC1973 cells by Targetscan and Dual-luciferase assay. We further analyzed miR-495 promoter methylation by sodium bisulfite sequencing method (BSP), and found DNA methyltransferase inhibitor, 5-AzaC concomitantly upregulated expression of miR-495 and downregulated its target gene STAT-3 and its downstream target VEGF. Furthermore, we further observed that 5-AzaC treatment, miR-495 mimics and STAT-3 knockdown significantly inhibited cell function in breast cancer by Transwell assay, EdU flow cytometry, Annexin V-FITC/PI combined with flow cytometry and Hoechst staining. Taken together, our data are first to demonstrate that the miR-495 is silenced due to promoter methylation in breast cancer. DNA methyltransferase inhibitor 5-AzaC could reverse miR‑495 (suppressor gene) and STAT-3 (oncogene). The anticancer properties of 5-AzaC were preliminarily confirmed in breast cancer.

MeSH terms

  • Apoptosis / genetics
  • Azacitidine / administration & dosage
  • Breast Neoplasms / genetics*
  • Breast Neoplasms / pathology
  • Cell Movement / genetics
  • Cell Proliferation / genetics
  • DNA Methylation / genetics
  • Demethylation / drug effects
  • Female
  • Gene Expression Regulation, Neoplastic
  • Humans
  • MCF-7 Cells
  • MicroRNAs / genetics*
  • Promoter Regions, Genetic
  • STAT3 Transcription Factor / genetics*
  • Vascular Endothelial Growth Factor A / genetics*


  • MIRN495 microRNA, human
  • MicroRNAs
  • STAT3 Transcription Factor
  • STAT3 protein, human
  • VEGFA protein, human
  • Vascular Endothelial Growth Factor A
  • Azacitidine