IN-VITRO METABOLISM OF CHLORO-2-METHYLPHENYL)-N'-(2-METHYLPROPYL)THIOUREA - SPECIES COMPARISON AND IDENTIFICATION OF A NOVEL THIOCARBAMIDE-GLUTATHIONE ADDUCT

The in. vitro metabolism of SDZ HDL 376, a thiocarbamide developed for the treatment of atherosclerosis, was investigated in rat, dog, monke y, and human liver microsomes, as well as in rat and human liver slice s. [C-14]SDZ HDL 376 was extensively metabolized in all the species ex cept human. In rat liver microsomes an S-oxide was the major metabolit e. In human and monkey microsomes, carbon hydroxylation was favored; T he NADPH-dependent oxidation of SDZ HDL 376 resulted in covalent bindi ng to microsomal protein. Addition of GSH to the incubations decreased protein binding in a concentration-dependent manner and resulted in a novel SDZ HDL 376-GSH adduct. Adduct formation required NADPH and was mediated predominately by cytochrome P450. Inhibition of cytochrome P 450 by 1-aminobenzotriazole resulted in a 95% decrease in adduct forma tion, while heat inactivation of flavin-containing monooxygenases resu lted in a 10% decrease. Unlike other thiocarbamides which form disulfi de adducts with GSH, the SDZ HDL 376 adduct contained a thioether link age as characterized by LC/MS/MS and reference to a synthetic standard . Reactions performed with [S-35]GSH resulted in a [S-35]SDZ HDL 376-G SH adduct, demonstrating the sulfur was derived from GSH. Adduct forma tion was faster in rat microsomal reactions compared to human microsom es. Other structurally unrelated thiocarbamides (phenylthiourea, methi mazole, 2-mercaptobenzimidazole, 2-mercaptoquinazoline, and 2-propyl-6 -thiouracil) did not form similar adducts in rat liver microsomes supp lemented with GSH. Therefore, the GSH adduct of SDZ HDL 376 is unique fdr this type of thiocarbamide. These results suggest that the bioacti vation and detoxification bf SDZ HDL 316 differ significantly from oth er thiocarbamides. Furthermore, the in vitro formation of S-oxides and GSH adducts in rat hepatic tissue, and ring hydroxylation and glucuro nidation in human hepatic tissue, suggests rats may be more susceptibl e to the toxicity of SDZ HDL 376 compared to humans.