title: Extrapolation of Elementary Rate Constants of P-glycoprotein-Mediated Transport from MDCKII-hMDR1-NKI to Caco-2 Cells.
abstract: The best parameters for incorporation into mechanistic physiologically based pharmacokinetic models for transporters are system-independent kinetic parameters and active (not total) transporter levels. Previously, we determined the elementary rate constants for P-glycoprotein (P-gp)-mediated transport (on- and off-rate constants from membrane to P-gp binding pocket and efflux rate constant into the apical chamber) using the structural mass action kinetic model in confluent MDCKII-hMDR1-NKI cell monolayers. In the present work, we extended the kinetic analysis to Caco-2 cells for the first time and showed that the elementary rate constants are very similar compared with MDCKII-hMDR1-NKI cells, suggesting they primarily depend on the interaction of the compound with P-gp and are therefore mostly independent of the in vitro system used. The level of efflux active (not total) P-gp is also fitted by our model. The estimated level of efflux active P-gp was 5.0 ± 1.4-fold lower in Caco-2 cells than in MDCKII-hMDR1-NKI cells. We also kinetically identified the involvement of a basolateral uptake transporter for both digoxin and loperamide in Caco-2 cells, as found previously in MDCKII-hMDR1-NKI cells, due to their low passive permeability. This demonstrates the value of our P-gp structural model as a diagnostic tool in detecting the importance of other transporters, which cannot be unambiguously done by the Michaelis-Menten approach. The system-independent elementary rate constants for P-gp obtained in vitro are more fundamental parameters than those obtained using Michaelis-Menten steady-state equations. This suggests they will be more robust mechanistic parameters for incorporation into physiologically based pharmacokinetic models for transporters.
authors.affiliation: Drexel University, Department of Biology, Philadelphia, Pennsylvania (Z.M., J.B.); and GlaxoSmithKline Pharmaceuticals, Drug Metabolism and Pharmacokinetics, King of Prussia, Pennsylvania (Z.M., H.E.).
authors.forename: Zhou
authors.initials: Z
authors.lastname: Meng
authors.affiliation: Drexel University, Department of Biology, Philadelphia, Pennsylvania (Z.M., J.B.); and GlaxoSmithKline Pharmaceuticals, Drug Metabolism and Pharmacokinetics, King of Prussia, Pennsylvania (Z.M., H.E.).
authors.forename: Harma
authors.initials: H
authors.lastname: Ellens
authors.affiliation: Drexel University, Department of Biology, Philadelphia, Pennsylvania (Z.M., J.B.); and GlaxoSmithKline Pharmaceuticals, Drug Metabolism and Pharmacokinetics, King of Prussia, Pennsylvania (Z.M., H.E.) bentzj@drexel.edu.
authors.forename: Joe
authors.initials: J
authors.lastname: Bentz
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D018938:Caco-2 Cells; D002219:Carbamates / Q000493:pharmacokinetics; D018929:Cell Culture Techniques; D002462:Cell Membrane / Q000378:metabolism*; D002463:Cell Membrane Permeability; D004077:Digoxin / Q000493:pharmacokinetics; D004285:Dogs; D005663:Furans; D006801:Humans; D007700:Kinetics; D008139:Loperamide / Q000493:pharmacokinetics; D061985:Madin Darby Canine Kidney Cells; D008954:Models, Biological; D011802:Quinidine / Q000493:pharmacokinetics; D013449:Sulfonamides / Q000493:pharmacokinetics
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D002219:Carbamates; D005663:Furans; D013449:Sulfonamides; C095108:amprenavir; D008139:Loperamide; D004077:Digoxin; D011802:Quinidine