In vitro reactions of 2-cyanoethylene oxide with calf thymus DNA

Abstract

Acrylonitrile oxide (CEO) is a direct-acting mutagen and the postulated proximate carcinogenic form of acrylonitrile (AN). We have studied the reactions of CEO with 2′-deoxyribonucleosides and in vitro with calf thymus DNA at pH 7.0–7.5 and 37°C for 3 h. Reaction of CEO with dAdo gave 2 adducts, N 6-(2-hydroxy-2-carboxyethyl)-dAdo ( N 6-HOCE-dAdo) (2% yield) and 1, N 6-etheno-dAdo (ε-dAdo) (11%); reaction with dCyd resulted in the isolation of 3-HOCE-dUrd (22%); reaction with dGuo gave 7-(2-oxoethyl)-Gua (7-OXE-Gua) (31%) and reaction with dThd yielded 3-OXE-dThd (3%). Structural elucidation of adducts was accomplished by ultraviolet spectroscopy, high-field proton NMR spectroscopy and mass spectrometry. Structural confirmation was provided by an accurate mass measurement technique where diagnostic ions in the electron impact mass spectra of trimethylsilyl derivatives were measured to within 0.0007 atomic mass units. The facile Dimroth rearrangement of 1-HOCE-dAdo to N 6-HOCE-dAdo and hydrolytic deamination of a dCyd adduct to 3-HOCE-dUrd is postulated to be catalyzed by the hydroxyl group on the 3-carbon side chain of the adduct. Reaction of CEO with calf thymus DNA yielded (nmol/mg DNA) N 6-HOCE-dAdo (2); ε-dAdo (11); 3-HOCE-dUrd (80); 7-OXE-Gua (110) and 3-OXE-dThd (1). Thus CEO, like its metabolic precursor AN, directly alkylates DNA in vitro but at a much more rapid rate.