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Updated: Dec 22, 2025
Generation of a Humanized Mouse Liver Using Human Hepatic Stem Cells
Published on: August 29, 2016
Human Aldehyde Oxidase 1-Mediated Carbazeran Oxidation in Chimeric TK-NOG Mice Transplanted with Human Hepatocytes
Shotaro Uehara1, Nao Yoneda2, Yuichiro Higuchi2
1Laboratory Animal Research Department, Central Institute for Experimental Animals, Kawasaki, Kanagawa, Japan (S.U., N.Y., Y.H., H.S.) and the Laboratory of Drug Metabolism and Pharmacokinetics, Showa Pharmaceutical University, Machida, Tokyo, Japan (H.Y.) s-uehara@ciea.or.jp.
Abstract:
Carbazeran is a potent phosphodiesterase inhibitor with species-dependent metabolic profiles in rats, dogs, and humans. In this study, we investigated the aldehyde oxidase (AOX)-mediated oxidation of carbazeran to 4-oxo derivatives in chimeric NOD/Shi-scid IL2 receptor gamma-null mice expressing a herpes simplex virus type 1 thymidine kinase transgene with humanized livers (humanized-liver mice). Liver cytosolic fractions from humanized-liver mouse effectively catalyzed carbazeran 4-oxidation with high affinity for the substrate, similar to those of the human liver cytosolic fractions and recombinant human AOX1 protein. Furthermore, hepatocytes prepared from humanized-liver mice and humans also exhibited substantial metabolism via carbazeran 4-oxidation. After a single oral administration of carbazeran (10 mg/kg), plasma levels of 4-oxo-carbazeran, N-desethyl-4-oxo-carbazeran, and 6,7-dimethoxy-1-[4-(hydroxy)-piperidino]-4-phthalazinone (three human metabolites formed via 4-oxidation) were higher in humanized-liver mice than in the control mice. In contrast, plasma levels of O-desmethylcarbazeran (a major metabolite in dogs) in control mice were higher than those in the humanized-liver mice. Relative excreted amounts of the three 4-oxidation-derived human-specific metabolites in the urine and feces were greater for humanized-liver mice than control mice, whereas the relative excreted amounts of O-desmethylcarbazeran were predominant in the urine and feces of control mice. Thus, the production of carbazeran 4-oxo derivatives was elevated in humanized-liver mice compared with control mice, in agreement with our in vitro enzyme-mediated oxidation data. These results suggest that hepatic human AOX1 functions in humanized-liver mice at the in vivo level and that humanized-liver mice may be useful for predicting drug metabolism in humans, at least with regard to human AOX1-dependent metabolism. SIGNIFICANCE STATEMENT: We found that the production of carbazeran 4-oxo derivatives was higher in humanized-liver mice than in control mice. These results were supported by the fact that carbazeran was rapidly metabolized to 4-oxo-carbazeran in humanized-liver mouse hepatocytes expressing human aldehyde oxidase 1. These results suggest that human aldehyde oxidase 1 is functional in humanized-liver mice in vivo and that chimeric NOD/Shi-scid IL2 receptor gamma-null mice expressing a herpes simplex virus type 1 thymidine kinase transgene transplanted with human hepatocytes may be a suitable model animal for predicting aldehyde oxidase-dependent biotransformation of drugs in humans.
Insights
Humanized-liver mice effectively metabolize carbazeran via aldehyde oxidase 1 (AOX1), producing human-specific metabolites. This suggests these mice are a valuable model for predicting human drug metabolism, particularly AOX1-dependent pathways.
Area of Science:
- Pharmacology
- Drug Metabolism
- Toxicology
Background:
- Carbazeran is a phosphodiesterase inhibitor with varied metabolic profiles across species.
- Aldehyde oxidase (AOX) is a key enzyme in drug metabolism, but its activity can differ significantly between species.
- Predicting human drug metabolism often relies on animal models, necessitating accurate species-specific representations.
Purpose of the Study:
- To investigate the role of aldehyde oxidase 1 (AOX1) in the metabolism of carbazeran in humanized-liver mice.
- To evaluate the utility of humanized-liver mice as a model for predicting human AOX1-dependent drug metabolism in vivo.
- To compare carbazeran metabolism in humanized-liver mice with control mice and human liver fractions.
Main Methods:
- Utilized chimeric NOD/Shi-scid IL2 receptor gamma-null mice with humanized livers (expressing human AOX1).
- Assessed carbazeran 4-oxidation in liver cytosolic fractions and hepatocytes from humanized-liver mice and humans.
- Administered carbazeran orally to humanized-liver and control mice, analyzing plasma and excreta for metabolites.
Main Results:
- Humanized-liver mouse liver fractions and hepatocytes efficiently metabolized carbazeran to 4-oxo derivatives, similar to human samples.
- Plasma levels of human-specific 4-oxo carbazeran metabolites were significantly higher in humanized-liver mice post-administration.
- Excretion patterns in humanized-liver mice favored human-specific metabolites, while control mice showed higher levels of dog-specific metabolites.
Conclusions:
- Hepatic human AOX1 is functional in vivo in humanized-liver mice, catalyzing carbazeran 4-oxidation.
- Humanized-liver mice accurately reflect human AOX1-dependent metabolism of carbazeran.
- These mice represent a valuable preclinical model for predicting aldehyde oxidase-mediated drug biotransformation in humans.
