Characterization of unusual pyrazole ring-rearranged metabolites of DS-1971a specific to monkeys
Daigo Asano1, Kazuyoshi Michiba1, Kayoko Masuda2
1Drug Metabolism and Pharmacokinetics Research Laboratories, Daiichi Sankyo Co., Ltd., Tokyo, Japan.
Abstract:
We previously reported that various metabolites were detected in mice, monkeys, and humans after oral administration of radiolabeled DS-1971a. Among these metabolites, mono- (M6) and di-oxidized (M12 and M13) forms were recognized as being specific to monkeys. Here, we report the definitive structures and formation mechanisms of these metabolites to elucidate the monkey-specific metabolism of DS-1971a. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis indicated that mass fragments of standard compounds for M6, M12 and M13, which were biosynthesized by a microorganism, were identical to those of in vivo samples from monkeys, but their full structures could not be identified. Therefore, M6, M12, and M13 were additionally subjected to UV, nuclear magnetic resonance (NMR) and X-ray crystal structure analyses, revealing that these metabolites possess 4-hydroxy (M6), and 4-hydroxy and 5-oxo pyrazole ring moieties (M12 and M13). Notably, M12 and M13 were rearranged stereoisomers of each other. Incubation study with recombinant monkey cytochrome P450s (P450s) suggested that monkey CYP2C8 is responsible for producing not only M6 from DS-1971a but also M12 and M13, both of which are further oxidized metabolites of M6. These results demonstrate novel and unusual metabolism of the pyrazole ring moiety of DS-1971a via monkey CYP2C8-mediated oxidation and rearrangement.
Insights
Monkey-specific metabolism of DS-1971a involves novel oxidation and rearrangement of its pyrazole ring. Researchers identified the structures and formation mechanisms of key metabolites (M6, M12, M13) mediated by monkey CYP2C8.
Area of Science:
- Pharmacokinetics and Drug Metabolism
- Medicinal Chemistry
- Biochemistry
Background:
- Oral administration of DS-1971a results in various metabolites across species.
- Specific metabolites (M6, M12, M13) were previously identified in monkeys, suggesting unique metabolic pathways.
Purpose of the Study:
- To definitively elucidate the structures and formation mechanisms of monkey-specific DS-1971a metabolites.
- To understand the role of cytochrome P450 enzymes in the metabolism of DS-1971a in monkeys.
Main Methods:
- Liquid chromatography-tandem mass spectrometry (LC-MS/MS) for fragment analysis.
- UV, nuclear magnetic resonance (NMR), and X-ray crystal structure analyses for definitive structure elucidation.
- Incubation studies with recombinant monkey cytochrome P450s (CYP2C8).
Main Results:
- The structures of metabolites M6, M12, and M13 were determined, revealing 4-hydroxy (M6) and 4-hydroxy/5-oxo pyrazole ring moieties (M12, M13).
- M12 and M13 were identified as rearranged stereoisomers.
- Monkey CYP2C8 was identified as the enzyme responsible for the oxidation of DS-1971a to M6, and further oxidation to M12 and M13.
Conclusions:
- DS-1971a undergoes a novel and unusual metabolic pathway in monkeys involving pyrazole ring oxidation and rearrangement.
- Monkey CYP2C8 plays a crucial role in the formation of these unique metabolites, M6, M12, and M13.
