Human Cytochrome P450 1A2 Involvement in the Formation of Reactive Metabolites from a Species-Specific Hepatotoxic Pyrazolopyrimidine Derivative, 5-n-Butyl-7-(3,4,5-trimethoxybenzoylamino)pyrazolo[1,5-a]pyrimidine

• Published in: Chemical research in toxicology 2009-02, Vol.22 (2), p.323-331
• Main Authors: Kuribayashi, Shunji, Goto, Kiyoto, Naito, Shinsaku, Kamataki, Tetsuya, Yamazaki, Hiroshi
• Format: Article
• Language: English
• Subjects: Animals; Chemical and Drug Induced Liver Injury - etiology; Chemical and Drug Induced Liver Injury - metabolism; Cysteine - chemistry; Cytochrome P-450 CYP1A2 - metabolism; Cytochrome P-450 CYP1A2 Inhibitors; Humans; Hydrolysis; Magnetic Resonance Spectroscopy; Microsomes, Liver - enzymology; Pyrazoles - chemistry; Pyrazoles - metabolism; Pyrazoles - pharmacology; Pyrimidines - chemistry; Pyrimidines - metabolism; Pyrimidines - pharmacology; Rats; Species Specificity
• ISSN: 0893-228X; 1520-5010
• DOI: 10.1021/tx8003592

5-n-Butyl-7-(3,4,5-trimethoxybenzoylamino)pyrazolo[1,5-a]pyrimidine) (OT-7100) is a pyrazolopyrimidine derivative with potential analgesic effects. Exclusively limited elevations in serum levels of aspirate- and alanine-aminotransferase were abnormally observed in a clinical study, in contrast to no toxicological potential to experimental animals. The aim of this study was to clarify the mechanism responsible for species-specific hepatotoxicity of this model compound. OT-7100 was primarily metabolized to a carboxylic acid derivative and an amino derivative (5-n-butyl-pyrazolo[1,5-a]pyrimidine, M-5) by hydrolysis in humans and rats. In human liver, pyrazolo[1,5-a]pyrimidine derivative M-5 was further metabolized to mainly M-23OH (a C-3-position hydroxyl derivative, 3-hydroxy-5-n-butyl-pyrazolo[1,5-a]pyrimidine). Studies with recombinant cytochrome P450s (P450s), correlation analysis using a panel of human liver microsomes as well as immunoinhibition with anti-P450 antibodies collectively suggested that human liver microsomal P450 1A2 preferentially metabolized M-5 to predominantly M-23OH. Human liver microsomes were capable of activating M-5 to a covalently bound metabolite faster than rat liver microsomes: reduced glutathione prevented the bindings. A cysteine adduct derivative of M-23OH at the C-6-position was structurally confirmed. On the contrary, rat liver microsomal P450 1A2 could metabolize M-5 to equally M-23OH, M-22OH (a C-6-position hydroxyl derivative, 6-hydroxy-5-n-butyl-pyrazolo[1,5-a]pyrimidine), or an unknown metabolite. These results suggest that differences in the regiospecific metabolic function of human and rat P450 1A2 would be responsible for the human-specific metabolic activation of the primary metabolite of OT-7100 to a proximate form. It is presumed that hepatotoxicity associated with OT-7100 could be likely related to the formation of a human-specific reactive metabolite from M-23OH. OT-7100 activation by inducible P450 1A2 may therefore exhibit marked individual differences.