DNA and protein adducts as indicators of in vivo methylation by nitrosatable drugs.

Abstract

Exposure to methylating carcinogens may be monitored by measuring both the formation of S-methylcysteine in haemoglobin and the urinary excretion of N-7-methylguanine (7-MeG), which is derived in part from methylated nucleic acids. Female rats were exposed to methylmethanesulphonate, methylnitrosourea and to three drugs, aminopyrine, cimetidine and pyrilamine, which are potential methylating agents if nitrosation occurs in vivo. Because S-methylcysteine in haemoglobin and urinary 7-MeG occur naturally, the experiments were carried out with stable isotope-labelled analogues which contained trideutero (d3)-methyl groups. Gas chromatography-mass spectrometry was used for the quantitative determination of d3-labelled adducts after their separation from the biological matrix and chemical derivatization. Transfer of the intact d3-methyl group to cysteine and guanine was detected after intragastric administration of d3-methyl-methanesulphonate (50 mg/kg), d3-N-methyl-N-nitrosourea (50 mg/kg), and d6-aminopyrine (AP) and sodium nitrite (both 100 mg/kg). AP alone gave no detectable d3-methyl adducts. Co-administration of nitrite and d6-pyrilamine or d3-cimetidine yielded no d3-7-MeG, although N-nitroso-d3-cimetidine alkylated DNA in vitro in a dose-dependent fashion. For AP and nitrite combinations urinary excretion of d3-7-MeG was linearly related to the dose of nitrite and was essentially complete within 5 days. For d3-methylmethane-sulphonate (50 mg/kg) the ratio of haemoglobin d3-S-methylcysteine to urinary d3-7-MeG was considerably (greater than 9-fold) higher than for either d3-N-methyl-N-nitrosourea or AP/nitrite (100 mg/kg) mixture. This is in accord with the SN2 nature of the weak carcinogen methylmethanesulphonate compared with the SN1 nature of the reactive methylating agent derived from either one of the N-methyl-N-nitroso compounds.