163 Aberrant methylation of a panel of miRNA genes as a novel biomarker of bladder cancer

163 Aberrant methylation of a panel of miRNA genes as a novel biomarker of bladder cancer

163 - Aberrant methylation of a panel of miRNA genes as a novel biomarker of bladder c... Page 1 of 2 e163 Aberrant methylation of a panel of miRNA g...

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e163 Aberrant methylation of a panel of miRNA genes as a novel biomarker of bladder cancer Shimizu T.1, Suzuki H.2, Nojima M.3, Kitamura H.4, Yamamoto E.2, Imai K.5, Masumori N.4, Tsukamoto T.4, Toyota M.6 1Sapporo

Medical University School of Medicine, Dept. of Urology & Dept. of Molecular Biology, Sapporo, Japan, 2Sapporo Medical University School of Medicine, Dept. of Molecular Biology & First Dept. of Internal Medicine, Sapporo, Japan, 3Sapporo Medical University School of Medicine, Dept. of Public Health & First Dept. of Internal Medicine, Sapporo, Japan, 4Sapporo Medical University School of Medicine, Dept. of Urology, Sapporo, Japan, 5University of Tokyo, The Institute of Medical Science, Tokyo, Japan, 6Sapporo Medical University School of Medicine, Dept. of Molecular Biology, Sapporo, Japan INTRODUCTION & OBJECTIVES: Bladder cancer is the one of the most common malignancies in the urothelial tract. MicroRNAs (miRNAs) are small, non-coding RNAs that control gene expression post-transcriptionally, either by degradation of target mRNAs or by inhibition of protein translation. MiRNAs are involved in cancer development and progression, acting as tumor suppressors or oncogenes. Recent studies have demonstrated a role of DNA methylation in silencing miRNAs in bladder cancer. We aimed to explore the involvement of epigenetic alterations in the regulation of miRNAs in bladder cancer. MATERIAL & METHODS: Two bladder cancer cell lines (T-24 and UM-UC-3) were treated with or without a DNA demethylating agent 5-aza-2’deoxycytidine (5-aza-dC) and a histone deacetylase (HDAC) inhibitor 4-phenylbutyric acid (PBA). MiRNA expression signatures were analyzed using TaqMan Array. We used UCSC genome browser to identify CpG islands located in the proximal upstream of the miRNAs. Methylation of miRNA genes was measured in 5 cancer cell lines and 83 primary tumors by bisulfite-pyrosequencing. In addition, urine specimens of bladder cancer patients before (n = 120) and after surgical treatment (n = 47) were collected, and urinary methylation of miRNA genes was analyzed. Receiver operating characteristic (ROC) curve analysis was carried out to assess the diagnostic performance of the miRNA gene methylation in urine samples. RESULTS: Of 664 miRNAs examined, treatment with 5-aza-dC plus PBA induced significant upregulation (>5-fold) of 208 miRNAs in T24 cells, and that of 200 miRNAs in UM-UC-3 cells, respectively. Among the miRNAs, 146 were commonly up-regulated in both T-24 and UM-UC3 cells. CpG islands were identified in the proximal upstream of 23 miRNA genes, and 12 of them were hypermethylated in bladder cancer cell lines. Among them, miR-137, -124-2, -124-3 and -9-3 were frequently and tumor-specifically methylated in primary cancers (miR-137, 57/83, 68.7%; -124-2, 42/83, 50.6%; -124-3, 54/83, 65.1%; -9-3, 38/83, 45.8%). Methylation of the miRNA genes was also detectable in urine specimens from cancer patients, which was dramatically reduced after surgical resection (P < 0.001). ROC curve analysis revealed a high performance of the urinary DNA methylation to distinguish pre- and postoperative urine samples (sensitivity, 81%; specificity, 89%; area under the curve (AUC) of 0.916), suggesting its diagnostic usefulness as a tumor marker. CONCLUSIONS: Our results demonstrated that epigenetic silencing of miRNA genes may be involved in the development of bladder cancer, and that methylation of miRNA genes could be

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4/6/2012

163 - Aberrant methylation of a panel of miRNA genes as a novel biomarker of bladder c... Page 2 of 2 e163a a useful biomarker for the bladder cancer detection.

file://F:\RamShankar\April\04-05-12\Cip\Sour\163.html

4/6/2012