The in vitro metabolism of SDZ HDL 376, a thiocarbamide developed for the treatment of atherosclerosis, was investigated in rat, dog, monkey, and human liver microsomes, as well as in rat and human liver slices. [14C]SDZ HDL 376 was extensively metabolized in all the species except human. In rat liver microsomes an S-oxide was the major metabolite. In human and monkey microsomes, carbon hydroxylation was favored. The NADPH-dependent oxidation of SDZ HDL 376 resulted in covalent binding 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 predominantly by cytochrome P450. Inhibition of cytochrome P450 by 1-aminobenzotriazole resulted in a 95% decrease in adduct formation, while heat inactivation of flavin-containing monooxygenases resulted in a 10% decrease. Unlike other thiocarbamides which form disulfide adducts with GSH, the SDZ HDL 376 adduct contained a thioether linkage as characterized by LC/MS/MS and reference to a synthetic standard. Reactions performed with [35S]GSH resulted in a [35S]SDZ HDL 376-GSH adduct, demonstrating the sulfur was derived from GSH. Adduct formation was faster in rat microsomal reactions compared to human microsomes. Other structurally unrelated thiocarbamides (phenylthiourea, methimazole, 2-mercaptobenzimidazole, 2-mercaptoquinazoline, and 2-propyl-6-thiouracil) did not form similar adducts in rat liver microsomes supplemented with GSH. Therefore, the GSH adduct of SDZ HDL 376 is unique for this type of thiocarbamide. These results suggest that the bioactivation and detoxification of SDZ HDL 376 differ significantly from other thiocarbamides. Furthermore, the in vitro formation of S-oxides and GSH adducts in rat hepatic tissue, and ring hydroxylation and glucuronidation in human hepatic tissue, suggests rats may be more susceptible to the toxicity of SDZ HDL 376 compared to humans.

The pulsed field gradient diffusion edited experiment, bipolar LED, has been combined with the Carr-Purcell-Meiboom-Gill (CPMG) spin-echo sequence for the analysis of solid phase resin samples. Spin-echo enhanced diffusion filtered spectroscopy (SPEEDY), when optimized, filters both the compounds that demonstrate fast diffusion rates as well as the compounds that demonstrate fast T(2) relaxation rates. Using this technique, compounds that are not covalently attached to the resin are not observed and contributions from the resin matrix are greatly attenuated. The interpretation of the resulting spectrum is more readily accessible. This technique lessens the importance of completely removing reaction residues or the wash solvents simply for analytical evaluation. The utility of the combined filtering scheme was demonstrated by the implementation into a NOESY sequence.

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The endocannabinoid system (ECS) plays a diverse role in human physiology ranging from the regulation of mood and appetite to immune modulation and the response to pain. Drug development that targets the cannabinoid receptors (CB and CB) has been explored; however, success in the clinic has been limited by the psychoactive side effects associated with modulation of the neuronally expressed CB that are enriched in the CNS. CB, however, are expressed in peripheral tissues, primarily in immune cells, and thus development of CB-selective drugs holds the potential to modulate pain among other indications without eliciting anxiety and other undesirable side effects associated with CB activation. As part of a collaborative effort among industry and academic laboratories, we performed a high-throughput screen designed to discover selective agonists or positive allosteric modulators (PAMs) of CB. Although no CB PAMs were identified, 167 CB agonists were discovered here, and further characterization of four select compounds revealed two with high selectivity for CB versus CB. These results broaden drug discovery efforts aimed at the ECS and may lead to the development of novel therapies for immune modulation and pain management with improved side effect profiles.