Derivation of a System-Independent K i for P-glycoprotein Mediated Digoxin Transport from System-Dependent IC 50 Data

• Published in: Drug metabolism and disposition 2018-03, Vol.46 (3), p.279
• Main Authors: Chaudhry, Aqsaa, Chung, Git, Lynn, Adam, Yalvigi, Akshata, Brown, Colin, Ellens, Harma, O'Connor, Michael, Lee, Caroline, Bentz, Joe
• Format: Article
• Language: English
• Subjects: Animals; ATP Binding Cassette Transporter, Subfamily B, Member 1 - metabolism; Biological Transport - physiology; Caco-2 Cells; Cell Line; Cell Line, Tumor; Digoxin - metabolism; Humans; Inhibitory Concentration 50; Kinetics; LLC-PK1 Cells; Swine
• ISSN: 1521-009X

It has been previously demonstrated that IC values for inhibition of digoxin transport across confluent polarized cell monolayers are system-dependent. Digoxin IC data from five laboratories participating in the P-glycoprotein (P-gp) IC Initiative, using Caco-2, MDCKII-hMDR1 or LLC-PK1-hMDR1 cells, were fitted by the structural mass action kinetic model for P-gp-mediated transport across confluent cell monolayers. We determined their efflux-active P-gp concentration [T(0)], inhibitor elementary dissociation rate constant from P-gp ( ), digoxin basolateral uptake clearance ( ), and inhibitor binding affinity to the digoxin basolateral uptake transporter ( ). We also fitted the IC data for inhibition of digoxin transport through monolayers of primary human proximal tubule cells (HPTCs). All cell systems kinetically required a basolateral uptake transporter for digoxin, which also bound to all inhibitors. The inhibitor was cell system-independent, thereby allowing calculation of a system-independent i. The variability in efflux-active P-gp concentrations and basolateral uptake clearances in the five laboratories was about an order of magnitude. These laboratory-to-laboratory variabilities can explain more than 60% of the IC variability found in the principal component analysis plot in a previous study, supporting the hypothesis that the observed IC variability is primarily due to differences in expression levels of efflux-active P-gp and the basolateral digoxin uptake transporter. HPTCs had 10- to 100-fold lower efflux-active P-gp concentrations than the overexpressing cell lines, whereas their digoxin basolateral uptake clearances were similar. HPTC basolateral uptake of digoxin was inhibited 50% by 10 M ouabain, suggesting involvement of OATP4C1.