Expression of Human Cytochromes P450 in Chimeric Mice With Humanized Liver

Katoh, Miki; Matsui, Tomohito; Nakajima, Miki; Tateno, Chise; Kataoka, Mikiko; Soeno, Yoshinori; Horie, Tetsuhiro; Iwasaki, Katsunori; Yoshizato, Katsutoshi; Yokoi, Tsuyoshi

doi:10.1124/dmd.104.001347

Cited by 107 publications

(87 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Many recent publications have unambiguously demonstrated the value of humanized mouse livers for the study of drug metabolism and hepatitis research [7][8][9]21,[23][24][25][26][27][28][29] . However, the albumin-uPA transgenic mouse used for these studies has several major practical limitations.…”

Section: Discussionmentioning

confidence: 99%

Robust expansion of human hepatocytes in Fah−/−/Rag2−/−/Il2rg−/− mice

et al. 2007

View full text Add to dashboard Cite

Mice that could be highly repopulated with human hepatocytes would have many potential uses in drug development and research applications. The best available model of liver humanization, the uroplasminogen-activator transgenic model, has major practical limitations. To provide a broadly useful hepatic xenorepopulation system, we generated severely immunodeficient, fumarylacetoacetate hydrolase (Fah)-deficient mice. After pretreatment with a urokinaseexpressing adenovirus, these animals could be highly engrafted (up to 90%) with human hepatocytes from multiple sources, including liver biopsies. Furthermore, human cells could be serially transplanted from primary donors and repopulate the liver for at least four sequential rounds. The expanded cells displayed typical human drug metabolism. This system provides a robust platform to produce high-quality human hepatocytes for tissue culture. It may also be useful for testing the toxicity of drug metabolites and for evaluating pathogens dependent on human liver cells for replication.The liver is the principal site for the metabolism of xenobiotic compounds, including medical drugs. Because many hepatic enzymes are species specific, it is necessary to evaluate the metabolism of candidate pharmaceuticals using cultured primary human hepatocytes 1,2 . Today, hepatocytes are isolated primarily from cadaveric organs and then shipped to the location where testing will be performed. The condition (viability and state of differentiation) of hepatocytes from cadaveric sources is highly variable, and many cell preparations are of marginal quality. The availability of high-quality human hepatocytes is further hampered by the fact that they cannot be substantially expanded in tissue culture 3,4 . Hepatocytes from readily available mammalian species, such as the mouse, are not suitable for drug testing because they have a different complement of metabolic enzymes and

show abstract

Section: Discussionmentioning

confidence: 99%

Robust expansion of human hepatocytes in Fah−/−/Rag2−/−/Il2rg−/− mice

et al. 2007

View full text Add to dashboard Cite

show abstract

“…Hydroxylation at the 49-position is one of the major metabolic routes of dmd.aspetjournals.org diclofenac, and metabolite II has been used as a marker of human CYP2C9 activity in humanized animal models. The in vitro assessment using liver microsome preparations from humanized uPA/severe combined immunodeficient chimeric mice with varying replacement ratios showed that the formation of II correlated well with hepatic replacement ratio, relative expression levels of CYP2C9 mRNA, and CYP2C9 protein content (Katoh et al, 2004;Tateno et al, 2004). Recently, Hu et al (2013) produced humanized TK-NOG chimeric mice using hepatocytes from donors with different genetic backgrounds.…”

Section: Discussionmentioning

confidence: 99%

Formation of the Accumulative Human Metabolite and Human-Specific Glutathione Conjugate of Diclofenac in TK-NOG Chimeric Mice with Humanized Livers

et al. 2014

View full text Add to dashboard Cite

39-Hydroxy-49-methoxydiclofenac (VI) is a human-specific metabolite known to accumulate in the plasma of patients after repeated administration of diclofenac sodium. Diclofenac also produces glutathioneconjugated metabolites, some of which are human-specific. In the present study, we investigated whether these metabolites could be generated in humanized chimeric mice produced from TK-NOG mice. After a single oral administration of diclofenac to humanized mice, the unchanged drug in plasma peaked at 0.25 hour and then declined with a half-life (t 1/2 ) of 2.4 hours. 49-Hydroxydiclofenac (II) and 39-hydroxydiclofenac also peaked at 0.25 hour and were undetectable within 24 hours. However, VI peaked at 8 hours and declined with a t 1/2 of 13 hours. When diclofenac was given once per day, peak and trough levels of VI reached plateau within 3 days. Studies with administration of II suggested VI was generated via II as an intermediate. Among six reported glutathione-conjugated metabolites of diclofenac, M1 (5-hydroxy-4-(glutathion-S-yl)diclofenac) to M6 (29-(glutathion-S-yl)monoclofenac), we found three dichlorinated conjugates [M1, M2 (49-hydroxy-39-(glutathion-S-yl)diclofenac), and M3 (5-hydroxy-6-(glutathion-S-yl)diclofenac)], and a single monochlorinated conjugate [M4 (29-hydroxy-39-(glutathion-S-yl)monoclofenac) or M5 (49-hydroxy-29-(glutathion-S-yl)monoclofenac)], in the bile of humanized chimeric mice. M4 and M5 are positional isomers and have been previously reported as human-specific in vitro metabolites likely generated via arene oxide and quinone imine-type intermediates, respectively. The biliary monochlorinated metabolite exhibited the same mass spectrum as those of M4 and M5, and we discuss whether this conjugate corresponded to M4 or M5. Overall, humanized TK-NOG chimeric mice were considered to be a functional tool for the study of drug metabolism of diclofenac in humans.

show abstract

“…The diclofenac 4Ј-hydroxylase activity was determined by the method used by Katoh et al (2004) with a 30-min incubation time. The product formation was determined using HPLC with a TSK-GEL ODS-80T M column (4.6 ϫ 250 mm; 5 m; TOSOH, Tokyo, Japan) and monitored at 280 nm.…”

Section: Methodsmentioning

confidence: 99%

Progesterone Receptor Membrane Component 1 Modulates Human Cytochrome P450 Activities in an Isoform-Dependent Manner

Oda

Nakajima²,

Toyoda³

et al. 2011

Drug Metab Dispos

Self Cite

View full text Add to dashboard Cite

ABSTRACT:Cytochromes P450 (P450s) catalyze the metabolism of a wide spectrum of compounds. Recently, progesterone receptor membrane component 1 (PGRMC1), which shares a key structural motif with cytochrome b 5 , has been reported to bind to sterol-or steroidsynthesizing P450s, enhancing their activities. In this study, we investigated whether PGRMC1 affects human drug-metabolizing P450 activities. Using coexpression systems for PGRMC1 and P450s (CYP3A4, CYP2C9, or CYP2E1) in HepG2 cells, we found that PGRMC1 decreased the V max values and increased the K m values of the CYP3A4 activities, and it decreased the V max values but did not affect the K m values of the CYP2C9 activities. In contrast, PGRMC1 hardly affected the CYP2E1 activities. These results suggest that PGRMC1 negatively modulates the drug-metabolizing activities of P450, although it was isoform but not substrate dependent. It is worth noting that coimmunoprecipitation analysis using coexpression systems for FLAG-PGRMC1 and Myc-P450s in human embryonic kidney 293 cells revealed that PGRMC1 interacts with all three P450s, although the affinity seemed to vary. In 29 human liver microsomes (HLMs), there was a 5-fold variability in the PGRMC1 protein levels. By the correlation analyses using the P450 activities and the PGRMC1 levels, we could neither observe the contribution of PGRMC1 to the P450 activities in HLMs nor that of the NADPH-cytochrome P450 reductase or cytochrome b 5 . In conclusion, in contrast to sterol-or steroid-synthesizing P450s, we found that PGRMC1 negatively modulates the human drug-metabolizing activities of P450 through direct interaction. Further studies are needed to determine the clinical significance of PGRMC1 in the pharmacokinetics of drugs.

show abstract

Expression of Human Cytochromes P450 in Chimeric Mice With Humanized Liver

Cited by 107 publications

References 32 publications

Robust expansion of human hepatocytes in Fah−/−/Rag2−/−/Il2rg−/− mice

Robust expansion of human hepatocytes in Fah−/−/Rag2−/−/Il2rg−/− mice

Formation of the Accumulative Human Metabolite and Human-Specific Glutathione Conjugate of Diclofenac in TK-NOG Chimeric Mice with Humanized Livers

Progesterone Receptor Membrane Component 1 Modulates Human Cytochrome P450 Activities in an Isoform-Dependent Manner

Contact Info

Product

Resources

About