ARYL SULFOTRANSFERASE IN DRUG AND XENOBIOTIC METABOLISM

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Sulfation of arylhydroxamic acids, arylhydroxylamines, and benzylic alcohols is an essential step leading to their biotransformation into chemically reactive, electrophilic compounds which are capable of binding covalently to nucleophilic sites on cellular macromolecules. This covalent binding is the initial step leading to various cytotoxic, mutagenic, and/or carcinogenic responses. One of the major goals of the research described in this application is to provide an understanding of the mechanisms involved in the regulation of the catalytic activities and concentrations of the aryl sulfotransferse(s) that catalyze sulfation of molecules containing arylhydroxamic acid, arylhydroxylamine, and benzylic alcohol functional groups. The other goal of this research is to develop quantitative methods for assessing the reactivity of the sulfate esters of these functional groups. Our long-term objective is to be able to accurately predict sulfation, and subsequent reactivity of the sulfate ester, for both new and existing drugs as well as other xenobiotics that contain arylhydroxamic acid, arylhydroxylamine, and benzylic alcohol functional groups. The proposed research will utilize purified enzymes, isolated hepatocytes, and immunochemical techniques to accomplish these goals. Regulation of aryl sulfotransferase (AST) IV activity will be studied with the purified enzyme and with isolated rat hepatocytes. The relative participation of AST II and AST IV in sulfation reactions catalyzed by isolated rat hepatocytes will also be investigated. Purified AST IV will be used to develop a quantitative method for determining rate constants for reaction of electrophilic sulfate esters with model nucleophiles. Kinetic constants will be determined for the AST IV-catalyzed sulfation of N-hydroxy-2-acetylaminofluorene and N-hydroxy-2- aminofluorene, as well as for the reaction of the resulting sulfate esters with model nucleophiles. Structure-activity studies will be carried out for the AST IV-catalyzed sulfation of primary and secondary arylhydroxylamines and aromatic dihydrodiols. Immunochemical studies on AST will yield information on the localizations and concentrations of AST II and AST IV in hepatic, cutaneous, and respiratory tract tissues of untreated and xenobiotic-treated rats. Furthermore, the distribution of AST IV in liver will be studied immunohistochemically after chronic treatment of rats with 2-acetylaminofluorene.