Concordant hypermethylation of intergenic microRNA genes in human hepatocellular carcinoma as new diagnostic and prognostic marker

Abstract

Epigenetic inactivation by aberrant DNA methylation has been reported for many microRNA genes in various human malignancies. However, relatively little is known about microRNA gene methylation in hepatocellular carcinoma (HCC). Therefore, a systematic screen for identification of aberrantly hypermethylated microRNA genes in HCC was initiated. The methylation status of 39 intergenic CpG island associated microRNA genes was analyzed in HCC cell lines (n 5 7), immortalized hepatocytes (n 5 2) and normal liver samples (n 5 5). Subsequently, 13 differentially methylated microRNA genes were analyzed in primary human HCC samples (n 5 40), benign liver tumors (n 5 15) and the adjacent liver tissues employing pyrosequencing. Expression of microRNA genes was measured using quantitative real-time polymerase chain reaction (RT-PCR). In addition, DNA methylation and expression of microRNA genes were measured after DNMT1 knockdown or DNMT inhibition. Aberrant hypermethylation and concomitant reduction in expression of intergenic microRNA genes is a frequent event in human HCC: hsamir-9-2 (23%), hsa-mir-9-3 (50 %), hsa-mir-124-1 (20%), hsa-mir-124-2 (13%), hsa-mir-124-3 (43%), hsa-mir-129-2 (58%), hsa-mir-596 (28%) and hsa-mir-1247 (38%). Altogether, it affects 90% of the HCC specimens under study. MicroRNA gene methylation is not found in hepatocellular adenoma (n 5 10) and focal nodular hyperplasia (n 5 5). DNMT1 knockdown or DNMT inhibition reduced microRNA gene methylation and stimulated expression. In primary human HCC specimens hypermethylation and expression of microRNA genes showed an inverse correlation. Concordant hypermethylation of three or more microRNA genes is a highly specific marker for the detection of HCC and for poor prognosis.

Demethylation of miR-9-3 and miR-193a Genes Suppresses Proliferation and Promotes Apoptosis in Non-Small Cell Lung Cancer Cell Lines

“…We unraveled here that miR-9-3 overexpression of miR-9-3 either by demethylation or by forced expression suppressed proliferation and enhanced apoptosis in NSClCs. In addition to NSCLCs showed in this study and some previous studies [9,10,11,12,13,14,15], methylation of miR-9-3 has also been described in a number of cancer cells of other tissue origins, including gastric cancer cells [33,38], hepatocellular carcinoma [39], bladder cancer [40], oropharyngeal squamous cell carcinomas [41], and renal cell carcinoma [42]. Based on our findings in this study, it is not unreasonable to speculate that demethylation of miR-9-3 may also produce anti-cancer effects in other types of cancers.…”

miR-1247-5p functions as a tumor suppressor in human hepatocellular carcinoma by targeting Wnt3

“…Currently, one study reported that the miR-1247-5p gene was hypermethylated in clinical samples of patients with HCC (19). However, the expression level and functional effects of miR-1247-5p in HCC are largely unknown.…”

Contributions of polymorphisms in miR146a, miR196a, and miR499 to the development of hepatocellular carcinoma

“…Many studies on the association between microRNAs (miRNAs) and hepatocellular carcinoma have reported that miRNAs may play an important role in cancer initiation, progression, outgrowth, and drug resistance (Anwar et al, 2013;Wojcicka et al, 2014). miRNAs belongs to a class of non-coding small RNAs that comprise 18-23 nucleotides (Reddy, 2015).…”