Research ArticlesKinetic Analyses for Species Differences in P-glycoprotein-Mediated Drug Transport
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INTRODUCTION
P-glycoprotein (P-gp) is a product of multidrug resistant 1 (MDR1) gene and is one of the most important transporters affecting the pharmacokinetics of drugs in normal cells as well as in cancer cells. A rational approach is needed to determine whether a compound is a P-gp substrate or not, especially in drug development. The Caco-2 cells and human MDR1 transfected cell lines such as LLC-GA5-COL150 and L-MDR1 established by Ueda et al.1 and Schinkel et al.,2 respectively, are frequently used
Materials
[mebmt-β-3H]Cyclosporin A (333.0 GBq/mmol) and [1,2,4-3H]dexamethasone (1480.0 GBq/mmol) were obtained from Amersham Biosciences (Buckinghamshire, UK). cis-(+)-[N-methyl-3H]Diltiazem (3145.0 GBq/mmol) was purchased from PerkinElmer Life Sciences (Boston, MA). Colchicine, cyclosporin A, dexamethasone, and diltiazem hydrochloride were obtained from Wako Pure Chemical Industries (Osaka, Japan). The mouse monoclonal antibodies (C219) and rabbit polyclonal antibodies (H241) against MDR1-encoded P-gp
Immunoblot Analysis of MDR1 Transfected Cells
The results of the immunoblot analysis using anti-P-gp antibodies, C219 and H241, are shown in Figure 1. In the case of the C219 antibodies, the densities of the bands from pMDR1, cMDR1, rMDR1b, and mmdr1a were higher than that from hMDR1 (Fig. 1A). The relative densities of hMDR1, pMDR1, cMDR1, rMDR1a, rMDR1b, mmdr1a, and mmdr1b were 1.0, 1.6, 2.6, 0.9, 1.6, 1.6, and 1.1, respectively. On the other hand, in the case of the H241 antibodies, the densities of the bands from pMDR1 and cMDR1 were
DISCUSSION
Absorption, distribution, metabolism, and excretion are important factors in the pharmacokinetics of a drug and are influenced by the effects of ATP-binding cassette proteins such as P-gp. In vitro studies using MDR1 transfected cells and in vivo studies using mdr1 knockout mice are frequently performed for drug discovery and the assessment of drug interactions.13,14 To estimate and extrapolate the pharmacokinetics and bioavailability of a drug in human, information about species differences is
ACKNOWLEDGMENTS
We acknowledge Brent Bell for reviewing the manuscript.
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