The hepatocellular uptake of the glutathione conjugate of bromosulfophthalein (BSPGSH) was examined in Eisai hyperbilirubinemic rats (EHBR; originating from Sprague-Dawley rats), which lacked the ATP-dependent canalicular transport for non-bile acid organic anions, a trend common to other mutant rat strains (TR- and GY, originating from Wistar rats). Single-pass perfused rat liver experiments were conducted with BSPGSH (26-257 microM) using the multiple indicator dilution technique. The steady-state extraction ratio of BSPGSH was close to zero due to lack of biliary excretion. After the introduction of a bolus dose containing vascular (51Cr-labeled red blood cells), interstitial (125I-labeled albumin and [14C]sucrose) and cellular space (D2O) indicators and [3H]BSPGSH into the portal vein, the outflow dilution profile of [3H]BSPGSH was found to display a protracted declining profile (tailing) at low input BSPGSH concentrations; the tail disappeared at higher BSPGSH concentrations. When data were fitted with the barrier-limited model of Goresky as used previously for BSPGSH for the Sprague-Dawley rat (SDR), model fitting was found to evoke an additional "deep pool" within the hepatocyte to account for the "tail" component. The deep pool became evident for the EHBR because biliary excretion of BSPGSH was absent and the rate of return from the deep pool was slow. The concentration of BSPGSH within the deep pool was estimated to be 12 +/- 8 times that in the cytosol. The binding of BSPGSH to EHBR S9 (effective binding concentration of 53 microM and a binding association constant KA of 2.4 x 10(4) M-1), however, was found to be lower than that of SDR S9 and could not account for the late-in-time data. The influx permeability-surface area product was concentration dependent and decreased from 0.27 to 0.01 ml.sec-1.g-1 with increasing BSPGSH concentration; the throughput component, or the portion of the dose that goes through the liver without entering the hepatocyte, increased with increasing concentration. The trends were characteristic of carrier-mediated transport and were similar to those found for the uptake of BSPGSH in SDR.
"The glutathione conjugate of bromosulfophthalein ( BSPGSH ) is a substrate of rat Oatp1a1 ( Pang et al 1998a ) and is excreted by Mrp2 ( Geng et al 1998 ) . When added to the perfused rat liver , the extraction ratio was found to vary from 0 . "
[Show abstract][Hide abstract] ABSTRACT: The administration of metabolites arising from new drug entities is often employed in drug discovery to investigate their associated toxicity. It is expected that administration of metabolites can predict the exposure of metabolites originating from the administration of precursor drug. Whether exact and meaningful information can be obtained from this has been a topic of debate. This communication summarizes observations and theoretical relationships based on physiological modelling for the liver, kidney and intestine, three major eliminating organs/tissues. Theoretical solutions based on physiological modelling of organs were solved, and the results suggest that deviations are expected. Here, examples of metabolite kinetics observed mostly in perfused organs that did not match predictions are provided. For the liver, discrepancies in fate between formed and preformed metabolites may be explained by the heterogeneity of enzymes, the presence of membrane barriers and whether transporters are involved. For the kidney, differences have been attributed to glomerular filtration of the preformed but not the formed metabolite. For the intestine, the complexity of segregated flows to the enterocyte and serosal layers and differences in metabolism due to the route of administration are addressed. Administration of the metabolite may or may not directly reflect the toxicity associated with drug use. However, kinetic data on the preformed metabolite will be extremely useful to develop a sound model for modelling and simulations; in-vitro evidence on metabolite handling at the target organ is also paramount. Subsequent modelling and simulation of metabolite data arising from a combined model based on both drug and preformed metabolite data are needed to improve predictions on the behaviours of formed metabolites.
Journal of Pharmacy and Pharmacology 11/2008; 60(10):1247-75. DOI:10.1211/jpp.60.10.0001 · 2.26 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Oatp1, the organic anion transport polypeptide, is an integral membrane protein cloned from rat liver that mediates the uptake of various organic anions such as bromosulfophthalein (BSP) and taurocholate (TCA). Recent studies by others revealed that the thrombin inhibitor, CRC 220, a modified dipeptide, was transported by oatp1. The present study was designed to examine whether another modified peptide, enalapril, an angiotensin-converting enzyme inhibitor, was also a substrate. Transport was studied with enalapril (1 to 800 micromol/L, with [3H]enalapril) in a HeLa cell line stably transfected with oatp1-cDNA under the regulation of a Zn2+-inducible promoter. Noninduced transfected cells (without zinc) that did not express oatp1 failed to take up enalapril. In contrast, cells expressing oatp1 transported enalapril, estrone sulfate (E1S), taurolithocholic acid sulfate (TLCAS), and the glutathione conjugate of BSP (BSPGSH). Uptake of enalapril by oatp1 at 37 degreesC was substantially higher than that at 4 degreesC. The rate at 37 degreesC (uptake rates for induced - noninduced, transfected cells) was linear over 5 minutes and was concentration-dependent, characterized by a Km of 214 +/- 67 micromol/L and a Vmax of 0.51 +/- 0.15 nmol/min/mg protein. Enalapril uptake was inhibited competitively by BSP (at 1, 5, 10, and 50 micromol/L) and TCA (at 5, 25, and 100 micromol/L) with inhibition constants (Ki) of 2 and 32 micromol/L, respectively. The metabolite enalaprilat was, however, not transported by oatp1. That oatp1 is not a general transporter of anionic compounds was further shown by the lack of transport of harmol sulfate, benzoate, and hippurate. These observations attest to the role of oatp1 as a specific transporter for at least two classes of pharmacologically important peptides.
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