Formation of Fused-Ring 2′-Deoxycytidine Adducts from 1-Chloro-3-buten-2-one an in Vitro 13-Butadiene Metabolite under in Vitro Physiological Conditions

摘要

1-Chloro-3-buten-2-one (CBO) is a potential metabolite of 1,3-butadiene (BD), a carcinogenic air pollutant. CBO is a bifunctional alkylating agent that readily reacts with glutathione (GSH) to form mono-GSH and di-GSH adducts. Recently, CBO and its precursor 1-chloro-2-hydroxy-3-butene (CHB) were found to be cytotoxic and genotoxic in human liver cells in culture with CBO being approximately 100-fold more potent than CHB. In the present study, CBO was shown to react readily with 2′-deoxycytidine (dC) under in vitro physiological conditions (pH 7.4, 37 °C) to form four dC adducts with the CBO moieties forming fused rings with the N3 and N⁴ atoms of dC. The four products were structurally characterized as 2-hydroxy-2-hydroxymethyl-7-(2-deoxy-β-D-erythro-pentofuranosyl)-1,2,3,4-tetrahy dro-6-oxo-6H,7H-pyrimido[1,6-a]pyrimidin-5-ium (>dC-1 and >dC-2, a pair of diastereomers), 4-chloromethyl-4-hydroxy-7-(2-deoxy-β-D-erythro-pentofuranosyl)-1,2,3,4-tetrahydr o-6-oxo-6H,7H-pyrimido[1,6-a]pyrimidin-5-ium (>dC-3), and 2-chloromethyl-2-hydroxy-7-(2-deoxy-β-D-erythro-pentofuranosyl)-1,2,3,4-tetrahydr o-6-oxo-6H,7H-pyrimido[1,6-a]pyrimidin-5-ium (>dC-4). Interestingly, >dC-1 and >dC-2 were stable under our experimental conditions (pH 7.4, 37 °C, 6 h) and existed in equilibrium as indicated by HPLC analysis, whereas >dC-3 and >dC-4 were labile with the half-lives being 3.0 ± 0.36 and 1.7 ± 0.06 h, respectively. Decomposition of >dC-4 produced both >dC-1 and >dC-2, whereas acid hydrolysis of >dC-1/>dC-2 and >dC-4 in 1 M HCl at 100 °C for 30 min yielded the deribosylated adducts >dC-1H/dC-2H and >dC-4H, respectively. Because fused-ring dC adducts of other chemicals are mutagenic, the characterized CBO-dC adducts could be mutagenic and play a role in the cytotoxicity and genotoxicity of CBO and its precursors, CHB and BD. The CBO-dC adducts may also be used as standards to characterize CBO-DNA adducts and to develop potential biomarkers for CBO formation in vivo.