The CYP2D6 humanized mouse: effect of the human CYP2D6 transgene and HNF4alpha on the disposition of debrisoquine in the mouse.
CYP2D6 is a highly polymorphic human gene responsible for a large variability in the disposition of more than 100 drugs to which humans may be exposed. Animal models are inadequate for preclinical pharmacological evaluation of CYP2D6 substrates because of marked species differences in CYP2D isoforms. To overcome this issue, a transgenic mouse line expressing the human CYP2D6 gene was generated. The complete wild-type CYP2D6 gene, including its regulatory sequence, was microinjected into a fertilized FVB/N mouse egg, and the resultant offspring were genotyped by both polymerase chain reaction and Southern blotting. CYP2D6-specific protein expression was detected in the liver, intestine, and kidney from only the CYP2D6 humanized mice. Pharmacokinetic analysis revealed that debrisoquine (DEB) clearance was markedly higher (94.1 +/- 22.3 l/h/kg), and its half-life significantly reduced (6.9 +/- 1.6 h), in CYP2D6 humanized mice compared with wild-type animals (15.2 +/- 0.9 l/h/kg and 16.5 +/- 4.5 h, respectively). Mutations in hepatic nuclear factor 4alpha (HNF4alpha), a hepatic transcription factor known to regulate in vitro expression of the CYP2D6 gene, could affect the disposition of CYP2D6 drug substrates. To determine whether the HNF4alpha gene modulates in vivo pharmacokinetics of CYP2D6 substrates, a mouse line carrying both the CYP2D6 gene and the HNF4alpha conditional mutation was generated and phenotyped using DEB. After deletion of HNF4alpha, DEB 4-hydroxylase activity in CYP2D6 humanized mice decreased more than 50%. The data presented in this study show that only CYP2D6 humanized mice but not wild-type mice display significant DEB 4-hydroxylase activity and that HNF4alpha regulates CYP2D6 activity in vivo. The CYP2D6 humanized mice represent an attractive model for future preclinical studies on the pharmacology, toxicology, and physiology of CYP2D6-mediated metabolism.
Corchero
J
J
Laboratory of Metabolism, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Granvil
C P
CP
Akiyama
T E
TE
Hayhurst
G P
GP
Pimprale
S
S
Feigenbaum
L
L
Idle
J R
JR
Gonzalez
F J
FJ
D018663:Adrenergic Agents / Q000493:pharmacokinetics; D000483:Alleles; D000818:Animals; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D019389:Cytochrome P-450 CYP2D6 / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D003647:Debrisoquin / Q000493:pharmacokinetics*; D017353:Gene Deletion; D018628:Gene Dosage; D018014:Gene Transfer Techniques; D051557:Hepatocyte Nuclear Factor 4; D006801:Humans; D008297:Male; D051379:Mice; D018345:Mice, Knockout / Q000378:metabolism; D008822:Mice, Transgenic / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D018663:Adrenergic Agents; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; C496592:HNF4A protein, human; D051557:Hepatocyte Nuclear Factor 4; C496593:Hnf4a protein, mouse; C403128:MLX protein, human; D010750:Phosphoproteins; C501970:Tcfl4 protein, mouse; D014157:Transcription Factors; D019389:Cytochrome P-450 CYP2D6; D003647:Debrisoquin