ABSTRACT
The multidrug resistance protein (MRP) 2 is predominantly expressed in liver, intestine, and kidney, where it plays an important role in the excretion of a range of drugs and their metabolites or endogenous compounds into bile, feces, and urine. Mrp knockout [Mrp2(-/-)] mice have been used recently to study the role of MRP2 in drug disposition. Here, we describe the first generation and initial characterization of a mouse line humanized for MRP2 (huMRP2), which is nulled for the mouse Mrp2 gene and expresses the human transporter in the organs and cell types where MRP2 is normally expressed. Analysis of the mRNA expression for selected cytochrome P450 and transporter genes revealed no major changes in huMRP2 mice compared with wild-type controls. We show that human MRP2 is able to compensate functionally for the loss of the mouse transporter as demonstrated by comparable bilirubin levels in the humanized mice and wild-type controls, in contrast to the hyperbilirubinemia phenotype that is observed in MRP2(-/-) mice. The huMRP2 mouse provides a model to study the role of the human transporter in drug disposition and in assessing the in vivo consequences of inhibiting this transporter by compounds interacting with human MRP2.
Subject(s)
Membrane Transport Proteins/genetics , Membrane Transport Proteins/metabolism , Multidrug Resistance-Associated Proteins/genetics , Multidrug Resistance-Associated Proteins/metabolism , Animals , Bilirubin/blood , Bilirubin/genetics , Bilirubin/metabolism , Bilirubin/urine , Biological Transport , Cell Line , Cytochrome P-450 Enzyme System/genetics , Cytochrome P-450 Enzyme System/metabolism , Embryonic Stem Cells/metabolism , Gene Knock-In Techniques , Humans , Intestinal Mucosa/metabolism , Kidney/metabolism , Liver/metabolism , Male , Mice , Mice, Inbred C57BL , Multidrug Resistance-Associated Protein 2 , RNA, Messenger/geneticsABSTRACT
Compared with rodents and many other animal species, the human cytochrome P450 (P450) Cyp2c gene cluster varies significantly in the multiplicity of functional genes and in the substrate specificity of its enzymes. As a consequence, the use of wild-type animal models to predict the role of human CYP2C enzymes in drug metabolism and drug-drug interactions is limited. Within the human CYP2C cluster CYP2C9 is of particular importance, because it is one of the most abundant P450 enzymes in human liver, and it is involved in the metabolism of a wide variety of important drugs and environmental chemicals. To investigate the in vivo functions of cytochrome P450 Cyp2c genes and to establish a model for studying the functions of CYP2C9 in vivo, we have generated a mouse model with a deletion of the murine Cyp2c gene cluster and a corresponding humanized model expressing CYP2C9 specifically in the liver. Despite the high number of functional genes in the mouse Cyp2c cluster and the reported roles of some of these proteins in different biological processes, mice deleted for Cyp2c genes were viable and fertile but showed certain phenotypic alterations in the liver. The expression of CYP2C9 in the liver also resulted in viable animals active in the metabolism and disposition of a number of CYP2C9 substrates. These mouse lines provide a powerful tool for studying the role of Cyp2c genes and of CYP2C9 in particular in drug disposition and as a factor in drug-drug interaction.
Subject(s)
Aryl Hydrocarbon Hydroxylases/genetics , Cytochrome P-450 Enzyme System/genetics , Multigene Family/genetics , Animals , Aryl Hydrocarbon Hydroxylases/metabolism , Cytochrome P-450 CYP2C9 , Cytochrome P-450 Enzyme Inhibitors , Cytochrome P-450 Enzyme System/metabolism , Humans , Hydroxylation , Inactivation, Metabolic , Liver/drug effects , Liver/enzymology , Mice , Mice, Inbred C57BL , Mice, Knockout , Multigene Family/drug effects , Tolbutamide/metabolism , Tolbutamide/pharmacokineticsABSTRACT
BACKGROUND: Hypoxia-inducible factor-1alpha (HIF-1alpha) expression was reported to be associated with tumor growth, progression and resistance to radio-/chemotherapy. Whether HIF-1alpha mRNA or protein expression is associated with histomorphological response or prognosis following neoadjuvant chemoradiation and surgery in resectable, locally-advanced esophageal cancer was analyzed. PATIENTS AND METHODS: Fifty-three patients received neoadjuvant chemoradiation (cisplatin, 5-fluorouracil, 36 Gy) followed by transthoracic en bloc esophagectomy. HIF-1alpha mRNA and protein expressions were analyzed by quantitative RT-PCR and immunostaining. RESULTS: In squamous cell carcinoma, HIF-1alpha mRNA expression was significantly higher than in paired normal epithelium (p < 0.001). Normal squamous epithelium showed significant elevated expression in adenocarcinomas, suggesting a field effect (p < 0.04). HIF-1alpha protein expression showed a significant regulation following chemoradiation. Neither HIF-1alpha mRNA nor protein expression was associated with histomorphological regression or prognosis. CONCLUSION: HIF-1alpha mRNA expression is differentially upregulated in esophageal squamous cell carcinoma compared to adenocarcinomas, but does not predict tumor regression or prognosis.
