Methylation of CpG island promoters in uveal melanoma

Abstract

Background: Inactivation of tumour-related genes by promoter hypermethylation is a common epigenetic event in the development of a variety of tumours. Aim: To investigate in primary uveal melanoma the status of promoter methylation of genes thought to be involved in tumour development: p16, TIMP3, RASSF1, RARB, FHIT, hTERT and APC. Methods: Gene promoter methylation was studied by methylation-sensitive single-strand conformation analysis and dot-blot assay in a series of 23 primary uveal melanomas. All DNA samples were obtained from paraffin-embedded formalin-fixed tissue blocks. Results: hTERT promoter methylation was found with a relatively high frequency (52%). Promoter methylation of p16, TIMP3, RASSF1, RARB, FHIT and APC was a rare event. For none of these genes did promoter methylation exceed 15% of tumour samples, and, for some genes (FHIT and APC), no methylation was found at all. Furthermore, promoter methylation was absent in 39% (9/ 23) of cases. In only 22% (5/23) of cases was hypermethylation of at least two promoters observed. Conclusions: Promoter methylation of hTERT is a regular event in uveal melanoma. Hypermethylation of the other genes studied does not seem to be an essential element in the development of this tumour. As promoter methylation of APC, RASSF1 and RARB is often observed in cutaneous melanoma, these results suggest that different epigenetic events occur in the development of cutaneous and uveal melanoma.Uveal melanoma is the most common primary intraocular tumour in adults, with a stable incidence of 4.3 new cases per million per year over the last 25 years.1 Despite new treatment modalities, mortality has not decreased, 2 mainly because of liver metastases. Although numerous studies have addressed the genetic events involved in the development of uveal melanoma, only a few have focused on the epigenetic events that may occur during tumorigenesis.CpG islands promoter methylation, associated with transcriptional gene silencing, has emerged as one of the most important epigenetic alterations in the development of human malignancies.3 Promoter methylation has been observed in many tumour types, 4 5 but there are few reports on uveal melanoma. p16 promoter has been found to be methylated in up to 32% of primary tumours and 50% of cell lines. [6][7][8] In derived cell lines, TIMP3 expression was found to be lower in liver metastatic cells than primary uveal tumour, and it was suggested that TIMP3 promoter methylation may be the cause of TIMP3 downregulation. 9 More recently, RASSF1 promoter methylation was identified in 50% of primary uveal melanoma, and a correlation was noticed between RASSF1 promoter methylation status and the development of metastasis. 10Although cutaneous and uveal melanoma share common morphological features, they differ substantially in their behaviour, metastatic spread and response to chemotherapy. There is increasing evidence that this is related to differences in their molecular phenotype. cDNA analysis has revealed that uveal and cutaneous melanoma...

Molecular pathology of uveal melanoma

“…12,14,15 ( Figure 1; Table 1) The role of CDKN2A promoter hypermethylation, as an additional mechanism of Rb inactivation, is controversial as the frequency of hypermethylation of this gene varies between 4 and 33% of UM. 12,[14][15][16]48,74 Increased cyclin D1 protein expression has been associated with larger tumour basal diameter, epithelioid cell type, and poor prognosis. 15 The p53 pathway is inhibited downstream to p53 in many UM, 30 and this may be a consequence of MDM2 overexpression, which is also common in UM and associated with a poor outcome.…”

Genetic and epigenetic insights into uveal melanoma

“…Moulin et al described promoter hypermethylation of hTERT as an essential event in UM. However, in the absence of any expression data and very limited studies on hTERT in UM, it is uncertain whether this altered promoter methylation or very rarely occurring mutational events in TERT gene contributes to metastasis . Hypermethylation of the Tumor necrosis factor–related apoptosis‐inducing ligand (TRAIL) receptors DcR1 and DcR2 was detected not only in UM but also in cutaneous melanoma .…”

Methylation in uveal melanoma

“…This observation is not disputed, but controversy arises when primary tumours are analysed, and estimates are given for the extend of CDKN2A methylation in uveal melanoma 2 – 4. The study by Moulin et al is an example of a study that displays some remarkable deviations from results we and others have achieved previously 5 6. We would like to comment on some of the technical and biological factors that may contribute to the lack of consistency in the results that are achieved in the field of uveal melanoma epigenetics.…”