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Scheer, Nico ; Henderson, Colin James ; Kapelyukh, Yury ; Rode, Anja ; Mclaren, Aileen W. ; MacLeod, Alastair Kenneth ; Lin, De ; Wright, Jayne ; Stanley, Lesley ; Wolf, C. Roland
Generation and Characterization of a Novel Multidrug Resistance Protein 2 Humanized Mouse Line
(2012)
Scheer, Nico ; Balimane, Praveen ; Hayward, Michael D. ; Buechel, Sandra ; Kauselmann, Gunther ; Wolf, C. Roland
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.
Scheele, Sandra ; Oertel, Dan ; Bongaerts, Johannes ; Evers, Stefan ; Hellmuth, Hendrik ; Maurer, Karl-Heinz ; Bott, Michael ; Freudl, Roland
Genetically humanized mice for proteins involved in drug metabolism and toxicity and mice engrafted with human hepatocytes are emerging as promising in vivo models for improved prediction of the pharmacokinetic, drug–drug interaction, and safety characteristics of compounds in humans. This is an overview on the genetically humanized and chimeric liver-humanized mouse models, which are illustrated with examples of their utility in drug metabolism and toxicity studies. The models are compared to give guidance for selection of the most appropriate model by highlighting advantages and disadvantages to be carefully considered when used for studies in drug discovery and development.
Salpati, Laurent ; Chu, Xiaoyan ; Chen, Liangfu ; Prasad, Bhagwat ; Dallas, Shannon ; Evers, Raymond ; Mamaril-Fishman, Donna ; Geier, Ethan G. ; Kehler, Jonathan ; Kunta, Jeevan ; Mezler, Mario ; Laplanche, Loic ; Pang, Jodie ; Soars, Matthew G. ; Unadkat, Jashvant D. ; van Waterschoot, Robert A.B. ; Yabut, Jocelyn ; Schinkel, Alfred H. ; Scheer, Nico ; Rode, Anja
Organic anion transporting polypeptide (Oatp) 1a/1b knockout and OATP1B1 and -1B3 humanized mouse models are promising tools for studying the roles of these transporters in drug disposition. Detailed characterization of these models will help to better understand their utility for predicting clinical outcomes. To advance this approach, we carried out a comprehensive analysis of these mouse lines by evaluating the compensatory changes in mRNA expression, quantifying the amounts of OATP1B1 and -1B3 protein by liquid chromatography–tandem mass spectrometry, and studying the active uptake in isolated hepatocytes and the pharmacokinetics of some prototypical substrates including statins. Major outcomes from these studies were 1) mostly moderate compensatory changes in only a few genes involved in drug metabolism and disposition, 2) a robust hepatic expression of OATP1B1 and -1B3 proteins in the respective humanized mouse models, and 3) functional activities of the human transporters in hepatocytes isolated from the humanized models with several substrates tested in vitro and with pravastatin in vivo. However, the expression of OATP1B1 and -1B3 in the humanized models did not significantly alter liver or plasma concentrations of rosuvastatin and pitavastatin compared with Oatp1a/1b knockout controls under the conditions used in our studies. Hence, although the humanized OATP1B1 and -1B3 mice showed in vitro and/or in vivo functional activity with some statins, further characterization of these models is required to define their potential use and limitations in the prediction of drug disposition and drug-drug interactions in humans.
Rösch, C. ; Kratz, F. ; Hering, T. ; Trautmann, S. ; Umanskaya, N. ; Tippkötter, Nils ; Müller-Renno, C.M. ; Ulber, Roland ; Hannig, M. ; Ziegler, C.
The interplay of albumin (BSA) and lysozyme (LYZ) adsorbed simultaneously on titanium was analyzed by gel electrophoresis and BCA assay. It was found that BSA and lysozyme adsorb cooperatively. Additionally, the isoelectric point of the respective protein influences the adsorption. Also, the enzymatic activity of lysozyme and amylase (AMY) in mixtures with BSA was considered with respect to a possible influence of protein-protein interaction on enzyme activity. Indeed, an increase of lysozyme activity in the presence of BSA could be observed. In contrast, BSA does not influence the activity of amylase.