Global alterations of DNA methylation in cholangiocarcinoma target the Wnt signaling pathway

Abstract

The molecular mechanisms underlying the genesis of cholangiocarcinomas (CCs) are poorly understood. Epigenetic changes such as aberrant hypermethylation and subsequent atypical gene expression are characteristic features of most human cancers. In CC, data regarding global methylation changes are lacking so far. We performed a genome-wide analysis for aberrant promoter methylation in human CCs. We profiled 10 intrahepatic and 8 extrahepatic CCs in comparison to non-neoplastic biliary tissue specimens, using methyl-CpG immunoprecipitation (MCIp) combined with whole-genome CpG island arrays. DNA methylation was confirmed by quantitative mass spectrometric analysis and functional relevance of promoter hypermethylation was shown in demethylation experiments of two CC cell lines using 5-aza-2 0 deoxycytidine (DAC) treatment. Immunohistochemical staining of tissue microarrays (TMAs) from 223 biliary tract cancers (BTCs) was used to analyze candidate gene expression at the protein level. Differentially methylated, promoter-associated regions were nonrandomly distributed and enriched for genes involved in cancer-related pathways including Wnt, transforming growth factor beta (TGF-b), and PI3K signaling pathways. In CC cell lines, silencing of genes involved in Wnt signaling, such as SOX17, WNT3A, DKK2, SFRP1, SFRP2, and SFRP4 was reversed after DAC administration. Candidate protein SFRP2 was substantially down-regulated in neoplastic tissues of all BTC subtypes as compared to normal tissues. A significant inverse correlation of SFRP2 protein expression and pT status was found in BTC patients. Conclusion: We provide a comprehensive analysis to define the genome-wide methylation landscape of human CC. Several candidate genes of cancer-relevant signaling pathways were identified, and closer analysis of selected Wnt pathway genes confirmed the relevance of this pathway in CC. The presented global methylation data are the basis for future studies on epigenetic changes in cholangiocarcinogenesis. (HEPATOLOGY 2014;59:544-554)

Cholangiocarcinoma: current knowledge and future perspectives consensus statement from the European Network for the Study of Cholangiocarcinoma (ENS-CCA)

“…A retrospective multicentre study published in 2013, including close to 200 patients with iCCA, concluded that intraarterial therapy was safe and this approach resulted in stable disease (62%) or partial to complete response (26%) 253 . The majority of patients were treated with conventional TACE or 90 Y-TARE, and there was no statistically significant difference in overall survival (median 13.2 months) between these two groups. In addition, 90 Y-TARE has provided partial response (27%) or stable disease (68%) in patients with iCCA 254 .…”

“…The majority of patients were treated with conventional TACE or 90 Y-TARE, and there was no statistically significant difference in overall survival (median 13.2 months) between these two groups. In addition, 90 Y-TARE has provided partial response (27%) or stable disease (68%) in patients with iCCA 254 . Hepatic arterial infusion includes a catheter-based intra-arterial infusion of chemotherapeutic agents, without embolization 253 .…”

“…Marked reduction in DNA hydroxymethylation character izes CCA tissue when compared with non-neoplastic tissue and a DNA mCyt content of ≥5.59% in peripheral blood mononuclear cells relates to a favourable outcome in primary liver cancers 81 . On the other hand, several promoters of genes involved in Wnt signalling are hypermethylated in CCA tissue 90 . A number of publications strongly suggest that epi genetic changes are early events in the malignant process, linking the tumour epigenetics to the microenvironment 82 and opening up opportunities for early detection of CCA 83 .…”

Molecular Pathogenesis and Targeted Therapies for Intrahepatic Cholangiocarcinoma

“…2) WNT signaling. The WNT pathway is highly activated in the tumor epithelium of human CCAs and is often characterized by overexpression of the ligands WNT7B and WNT10A along with several target genes (69,70). It has been demonstrated that inflammatory macrophages in the stroma surrounding the tumor are required for the maintenance of this highly activated WNT signaling status (69,71).…”

Molecular Pathogenesis and Current Therapy in Intrahepatic Cholangiocarcinoma

“…Intriguingly, the etiological origins of CCA have been shown to impact on the associated mutational landscapes, both in comparisons of liver fluke-associated and non-associated tumors [34] , as well as hepatitis-positive versus -negative tumors [35] . Epigenetic lesions, such as focal promoter-associated hypermethylation [36,37] , global loss of 5-hydroxymethylcytosine [38] and aberrant non-coding RNA expression [39,40] , also contribute to iCCA pathogenesis.…”