Journal Article

Multiple markers for melanoma progression regulated by DNA methylation: insights from transcriptomic studies

William M. Gallagher, Orla E. Bergin, Mairin Rafferty, Zoë D. Kelly, Ilse-Maria Nolan, Edward J.P. Fox, Aedin C. Culhane, Linda McArdle, Mario F. Fraga, Linda Hughes, Caroline A. Currid, Fiona O'Mahony, Aileen Byrne, Alison A. Murphy, Catherine Moss, Susan McDonnell, Raymond L. Stallings, Jane A. Plumb, Manel Esteller, Robert Brown, Peter A. Dervan and David J. Easty

in Carcinogenesis

Volume 26, issue 11, pages 1856-1867
Published in print November 2005 | ISSN: 0143-3334
Published online June 2005 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/bgi152
Multiple markers for melanoma progression regulated by DNA methylation: insights from transcriptomic studies

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The incidence of melanoma is increasing rapidly, with advanced lesions generally failing to respond to conventional chemotherapy. Here, we utilized DNA microarray-based gene expression profiling techniques to identify molecular determinants of melanoma progression within a unique panel of isogenic human melanoma cell lines. When a poorly tumorigenic cell line, derived from an early melanoma, was compared with two increasingly aggressive derivative cell lines, the expression of 66 genes was significantly changed. A similar pattern of differential gene expression was found with an independently derived metastatic cell line. We further examined these melanoma progression-associated genes via use of a tailored TaqMan Low Density Array (LDA), representing the majority of genes within our cohort of interest. Considerable concordance was seen between the transcriptomic profiles determined by DNA microarray and TaqMan LDA approaches. A range of novel markers were identified that correlated here with melanoma progression. Most notable was TSPY, a Y chromosome-specific gene that displayed extensive down-regulation in expression between the parental and derivative cell lines. Examination of a putative CpG island within the TSPY gene demonstrated that this region was hypermethylated in the derivative cell lines, as well as metastatic melanomas from male patients. Moreover, treatment of the derivative cell lines with the DNA methyltransferase inhibitor, 2′-deoxy-5-azacytidine (DAC), restored expression of the TSPY gene to levels comparable with that found in the parental cells. Additional DNA microarray studies uncovered a subset of 13 genes from the above-mentioned 66 gene cohort that displayed re-activation of expression following DAC treatment, including TSPY, CYBA and MT2A. DAC suppressed tumor cell growth in vitro. Moreover, systemic treatment of mice with DAC attenuated growth of melanoma xenografts, with consequent re-expression of TSPY mRNA. Overall, our data support the hypothesis that multiple genes are targeted, either directly or indirectly, by DNA hypermethylation during melanoma progression.

Keywords: 5mC, 5-methylcytosine; CGH, comparative genomic hybridization; COBRA, combined restriction bisulfite restriction analysis; DAC, 2′-deoxy-5-azacytidine; FISH, fluorescence in situ hybridization; LDA, low density array; RGP, radial growth phase; VGP, vertical growth phase

Journal Article.  8372 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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