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R. B. Avraham, E. Merhavi, Y. Cohen, S. Frenkel, J. Pe'er, N. Goldenberg-Cohen; DNA Hypermethylation Status of Multiple Genes in Retinoblastoma. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1602. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Retinoblastoma, a rare pediatric eye tumor, is the most common primary intraocular cancer in children. The retinoblastoma protein, Rb, functions as a tumor suppressor by controlling cell cycle progression through complex interaction of multiple kinases and their inhibitors that together form the Rb pathway. The pathogenesis of retinoblastoma is known to involve inactivation of both copies of the RB1 gene and other genetic or epigenetic alterations in independent molecular pathways. Epigenetic silencing of tumor suppression genes (TSG) by methylation of discrete regions of the CpG Island is a major mechanism for tumorigenesis. To further explore the role of epigenetic genes inactivation in RB, we investigated the methylation phenotype commonly inactivated in other human cancer. Methylation of at least 3 of 5 specific genes may represent a distinct trait referred to CpG island methylator phenotype (CIMP). Positive CIMP tumors are associated with positive BRAF mutations. In this study, we will investigate oncogenic BRAF mutations, the role of epigenetic silencing of multiple TSG in retinoblastoma, and the methylation phenotype.
The promoter methylation status of the genes RAS-associated domain family 1A (RASSF1A), death associated protein kinase (DAPK), retinoic acid receptor beta (RAR-beta2) and O6-methyl-guanine-DNA methyltransferase (MGMT), which are important in cell growth regulation, apoptosis, and DNA repair, was determined in DNA extracted from RB tumor samples of 19 patients using methylation-specific polymerase chain reaction. CIMP status was determined for other genes: SOCS-1, IGF-2, RUNX3, NEUROG1 and CACNA1G. BRAF mutations in RB samples were analyzed using PCR and direct sequencing.
No BRAF mutations were found. The frequency of cancer-related gene methylation was: RASSF1A 89%, NEUROG1 50%, DAP-kinase 5%, RUNX3 5%, CACNA1G 5% , and none in MGMT , RAR-ß2, SOCS-1 and IGF-2.
High rate of hypermethylated RASSF1A in RB has been previously reported. The absence of MGMT hypermethylation might be associated with low tumor stage. We found negative CIMP in RB, as well as lack of BRAF mutations. The frequent methylation status found in NEUROG1 may provide an alternative pathway in the development and progression of RB, but further study is needed.
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