The CYP2D6 humanized mouse: Effect of the human CYP2D6 transgene and HNF4 alpha on the disposition of debrisoquine in the mouse

CYP2D6 is a highly polymorphic human gene responsible for a large variabili ty in the disposition of more than 100 drugs to which humans may be exposed . Animal models are inadequate for preclinical pharmacological evaluation o f CYP2D6 substrates because of marked species differences in CYP2D isoforms . To overcome this issue, a transgenic mouse line expressing the human CYP2 D6 gene was generated. The complete wild-type CYP2D6 gene, including its re gulatory sequence, was microinjected into a fertilized FVB/N mouse egg, and the resultant offspring were genotyped by both polymerase chain reaction a nd Southern blotting. CYP2D6-specific protein expression was detected in th e liver, intestine, and kidney from only the CYP2D6 humanized mice. Pharmac okinetic analysis revealed that debrisoquine (DEB) clearance was markedly h igher (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 4 alpha (HNF4 alpha), a hepatic transcription factor known to regul ate in vitro expression of the CYP2D6 gene, could affect the disposition of CYP2D6 drug substrates. To determine whether the HNF4 alpha gene modulates in vivo pharmacokinetics of CYP2D6 substrates, a mouse line carrying both the CYP2D6 gene and the HNF4 alpha conditional mutation was generated and p henotyped using DEB. After deletion of HNF4 alpha, DEB 4-hydroxylase activi ty 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 disp lay significant DEB 4-hydroxylase activity and that HNF4 alpha regulates CY P2D6 activity In vivo. The CYP2D6 humanized mice represent an attractive mo del for future preclinical studies on the pharmacology, toxicology, and phy siology of CYP2D6-mediated metabolism.