In vitro metabolism of acrylonitrile to 2-cyanoethylene oxide, reaction with glutathione, and irreversible binding to proteins and nucleic acids
Guengerich, F.P.; Geiger, L.E.; Hogy, L.L.; Wright, P.L.
Cancer Research 41(12 Pt 1): 4925-4933
1981
ISSN/ISBN: 0008-5472 PMID: 6272984 Document Number: 172067
The metabolism of the suspected carcinogen acrylonitrile was studied using subcellular fractions isolated from rats and humans. Irreversible binding of radioactive label from [1-14C]- or [2,3-14C]acrylonitrile to protein and DNA was enhanced by reduced NADP in the presence of rat liver microsomes or a reconstituted cytochrome P-450 system. During the reduced NADP-dependent reaction, HCN was produced and the heme of cytochrome P-450 was destroyed. Rat brain microsomes did not produce detectable levels of metabolites. Conclusive evidence of metabolically mediated binding of acrylonitrile to protein and DNA in human systems was not found. Metabolism was induced in rats by pretreatment of animals with either phenobarbital or 5,6-benzoflavone. Labeled 2-cyanoethylene oxide was found to bind irreversibly to calf thymus DNA and microsomal protein. The extent of binding was greater in the case of 2,3-14C-labeled than 1-14C-labeled material. The relative 1st-order rate of acrylonitrile binding to calf thymus DNA in rat liver microsomal systems was 1-2 orders of magnitude less than that for 1,1,2-trichloroethylene and 3 orders of magnitude less than that for vinyl chloride or vinyl bromide. Rat liver microsomes or a reconstituted cytochrome P-450 system catalyzed the mixed-function oxidation of acrylonitrile to 2-cyanoethylene oxide. 2-Cyanoethylene oxide has a half-life of about 2 h in neutral buffer at 37.degree. This epoxide was found to serve as a substrate for microsomal epoxide hydrolase. HCN was released during hydrolysis of 2-cyanoethylene oxide or reaction of the epoxide with reduced glutathione; HCN release in either case was not stoichiometric with epoxide disappearance. 2-Cyanoethylene oxide reacted less rapidly with reduced glutathione than did acrylonitrile. Rat liver cytosol preparations contained glutathione S-transferase activity towards acrylonitrile and greater activity with 2-cyanoethylene oxide as a substrate. Cytosol preparations from rat brain and human liver had no detectable glutathione S-transferase activity towards acrylonitrile but did exhibit some activity towards 2-cyanoethylene oxide. These studies establish the basic pathways involved in the metabolism of acrylonitrile and should provide a basis for examination of the relevance of these individual steps and their roles in bioactivation and detoxication under more physiological situations.