methyl-radical and cumene-hydroperoxide

During carcinogenesis, methylation of the C-5 position of cytosines in the promoter region of tumor suppressor genes is often observed. Enzymatic DNA methylation is a widely accepted mechanism for this phenomenon. It is interesting to propose a free radical mechanism for 5-methyldeoxycytidine (m(5)dC) production, because the C-5 position of cytosine is an active site for free radical reactions. When deoxycytidine (dC) and cumene hydroperoxide (CuOOH), a tumor promoter and a methyl radical producer, were reacted in the presence of ferrous ion at pH 7.4, the formation of m(5)dC was observed. The same reaction also proceeded with t-butyl hydroperoxide (BuOOH). The formation of m(5)dC was also observed in DNA by the CuOOH treatment. This is the first report of chemical DNA methylation at cytosine C-5 by environmental tumor promoters. We propose here that this reaction is one of the important mechanisms of de novo DNA methylation during carcinogenesis, because methyl radicals are produced by the biotransformation of various endogenous and exogenous compounds.

Topics: Benzene Derivatives; Carcinogens; Chromatography, Liquid; Cytosine; DNA; DNA Methylation; Environmental Pollutants; Epigenesis, Genetic; Mass Spectrometry; Methane; tert-Butylhydroperoxide

The mechanism for the reaction of cytochrome c with t-butyl hydroperoxide and cumene hydroperoxide was investigated. ESR spin trapping studies using 5,5-dimethyl-1-pyrroline N-oxide were performed to demonstrate the presence of hydroperoxide-derived peroxyl, alkoxyl, and methyl radicals. Computer simulation of the experimental data obtained at various 5,5-dimethyl-1-pyrroline N-oxide concentrations was used to determine the relative contributions of each radical adduct to each composite ESR spectrum. From these analyses, it was concluded that the alkoxyl radical of the hydroperoxide was the initial radical produced, presumably by homolytic scission of the O-O bond by ferric cytochrome c. This was in contrast to a previous ESR study that proposed a heterolytic peroxidase-type mechanism for the reaction of cytochrome c with organic hydroperoxides. Methyl radicals were produced from the beta-scission of the alkoxyl radical. The peroxyl radicals are shown to be secondary products formed from the reaction of oxygen with the methyl radical to produce the methyl peroxyl radical. In separate experiments, visible absorption spectroscopy revealed that the heme center was destroyed during the reaction. Both the heme destruction and production of radical adducts were inhibited by cyanide, presumably due to the formation of a cyanoheme complex.

Topics: Alcohols; Benzene Derivatives; Computer Simulation; Cyclic N-Oxides; Cytochrome c Group; Deferoxamine; Electron Spin Resonance Spectroscopy; Free Radicals; Methane; Models, Chemical; Oxygen; Peroxides; Reactive Oxygen Species; Spectrophotometry; Spin Labels; tert-Butylhydroperoxide