MECHANISMS OF RADIOPROTECTION & TOXICITY OF SH COMPOUNDS

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This project is designed to elucidate the molecular mechanisms of radiation-induced damage and repair as modified by exogenous thiols. The specific aims involve two interrelated areas: radioprotection and thiol toxicity. The radioprotection research will investigate (1) the importance of thiol and oxygen concentrations in determining the degree of radioprotection by thiols, thus testing the degree of applicability of the damage repair-fixation competition model and relate models to radioprotection; (2) the generality of mechanisms of radioprotection in thiols with varying chemicals and biochemical reactivities with 02, OH, organic radicals, H202, metals, etc.; (3) the relationship between radiation-induced cell killing and production of DNA damage, especially double strand breaks, and modification of that relationship by thiols; and (4) the involvement of thiols in enzymatic repair processes in irradiated cells. The toxicity studies will establish conditions for control of thiol-generated peroxide in order that the thiols are used properly in the radioprotection experiments or as a means to generate reactive oxygen species. The research has health-related implications because of its potential application to protection in cancer radiotherapy or radioprotection from nuclear disaster. Furthermore, the thiol studies are relevant to the current interest in oxygen radicals as damaging agents in chemotherapy, carcinogenesis, aging and arthritis. Radioprotection and toxicity will be studied using both cell survival and DNA damage assays. Cell survival will be measured using classical clongenic assays in rodent and human tumor cell lines. DNA damage measurements will be made using elution techniques. Cells for both survival and DNA studies will be irradiated using suspension and thin-layer techniques. The repair studies include measurements of DNA repair, potentially lethal damage repair, sublethal damage repair, and use of repair-deficient mammalian cells. The thiols to be used are dithiothreitol, glutathione, and cysteamine and its derivatives, including WR-1065. All studies will be designed to control for oxygen level and metal- catalyzed thiol oxidation, and resulting peroxide formation.