Promoter methylation profiling of 30 genes in human malignant melanoma

Abstract

Aberrant methylation and demethylation of promoter CpG islands lead to silencing of tumor-suppressor genes and abnormal expression of normally methylated genes, respectively. Here, we analyzed human melanomas for their methylation and demethylation profiles. Methylation status of core regions in promoter CpG islands was examined for 20 (candidate) tumor-suppressor genes, 4 genes that are not considered as tumor-suppressors, but are frequently silenced in human cancers, and 6 normally methylated melanoma antigen genes (MAGEs). Analysis of 13 melanoma cell lines and 2 cultured normal human epidermal melanocytes (HEMs) showed that 9 tumor-suppressor genes and all 4 non-tumor-suppressor genes were methylated in at least 1 cell line, but never in HEMs, and that all 6 MAGE genes were demethylated in 3 to 13 cell lines. Interestingly, we detected no methylation of MGMT, PTEN, MTAP and p27, which were previously reported as silenced in melanomas. Furthermore, 3 genes that were frequently methylated in the cell lines and 6 MAGE genes were analyzed in 25 surgical melanoma samples. RARB, RASSF1A and 3-OST-2 were methylated in 5 (20%), 9 (36%) and 14 (56%) samples, respectively. MAGE-A1, A2, A3, B2, C1 and C2 were demethylated in 9 (36%), 22 (88%), 20 (80%), 7 (28%), 21 (84%) and 16 (64%) samples, respectively. At least 1 gene was methylated in 18 (72%) samples and at least 1 was demethylated in 24 (96%) samples. No correlation between frequent methylation and frequent demethylation was observed. These profiles showed that both aberrant methylation and demethylation occur widely in human melanomas. (Cancer Sci 2004; 95: 962-968) ethylation of promoter CpG islands (CGIs) leads to transcriptional silencing of their downstream genes, and plays important roles in embryonic development and tissue differentiation.1) In tumors, silencing of tumor-suppressor genes, such as p16, CDH1 (E-Cadherin) and MLH1, by promoter methylation is one of the major mechanisms for their inactivation, along with mutations and loss of heterozygosity.2-5) At the same time, overall genome hypomethylation, which is mainly due to hypomethylation of repetitive sequences, 6, 7) is consistently observed in tumors.8, 9) Some tumors display aberrant demethylation of normally methylated CGIs and expression of their downstream product, such as melanoma antigen genes (MAGEs) and cancer-testis antigen genes.10-12) Frequent promoter methylation and frequent promoter demethylation are independent events, at least in human gastric cancers. 9)Malignant melanoma is one of major cause of cancer deaths and its incidence is increasing, especially in Western countries.13) Unexpectedly, in view of the deep involvement of ultraviolet radiation and mutations induced by it, aberrant methylation is observed for PTEN, p27, MTAP and RASSF1A in melanomas.14-17) Analysis of 15 tumor-suppressor genes in 18 melanoma cell lines showed methylation of TSP1, CDH1, RARB, GSTP1 and RASSF1A in melanoma cell lines.18) Recently, analysis of 7 tumor-suppressor genes in 15 melanoma cell line...

Identification of candidate tumor suppressor genes inactivated by promoter methylation in melanoma

“…It should be noted that similarly to the samples reported by Furuta et al, (Furuta et al, 2004), none of our cell lines showed PTEN promoter hypermethylation, which has been shown or suggested in some melanomas (Zhou et al, 2000;Mirmohammadsadegh et al, 2006). Similarly, using the criterion of a minimum 2-fold change after drug treatment, we have no evidence that the p14ARF promoter is methylated in any of the 7 cell lines tested in which the gene is not homozygously deleted.…”

Molecular mimicry of MAGE-A6 andMycoplasma penetransHF-2 epitopes in the induction of antitumor CD8+T-cell responses

“…as found in the pro-tumor niche), MAGE protein expression can be induced. 15 Expression of MAGE-A family proteins has been reported for more than half of all human cancers. 16 One of its members, MAGE-A6, is expressed in more than 60% of melanomas, 17 30% of renal cell carcinomas, 18 and by numerous other malignancies, such as breast, esophageal, head and neck, bladder, and lung carcinomas.…”

Silencing of tissue factor pathway inhibitor‐2 gene in malignant melanomas

“…These cell lines were selected because they had different origins and frequencies of methylation 19 : HMV-I, MeWo and WM-115 originated from a primary lesion of genital mucosa, a metastatic lesion of a lymph node, and a primary skin lesion, respectively, and exhibited frequent, infrequent and intermediately frequent gene silencing, respectively.…”