Pharmacokinetics, Pharmacodynamics and Drug MetabolismSodium fluorescein is a probe substrate for hepatic drug transport mediated by OATP1B1 and OATP1B3
Section snippets
INTRODUCTION
In addition to hepatic metabolism and biliary excretion, hepatic drug uptake is an important aspect of hepatobiliary drug disposition of endogenous and exogenous compounds. In the last decennia, several transporters influencing drug clearance have been identified.1 In the basolateral hepatocyte membrane, members of the organic anion transporting polypeptide family [OATP (human), Oatp (rat); SLCO/Slco gene family] support the sodium‐independent uptake of bile salts [e.g., taurocholate (TC)] in
Materials
Amprenavir (APV) was kindly provided by GlaxoSmithKline (Middlesex, UK). Ritonavir (RTV), indinavir (IDV) sulfate, saquinavir (SQV) mesylate, and nelfinavir mesylate were donated by Hetero Drugs Limited (Hyderabad, India). Atazanavir (ATV) was obtained from Bristol–Myers Squibb (New Brunswick, New Jersey) and darunavir (DRV) was obtained through the National Institutes of Health AIDS Reagent Program. CGamF was kindly provided by Professor Alan Hofmann (University of California, San Diego).
Determination of NaFluo Uptake Kinetics in Suspended Rat and Human Hepatocytes
Preliminary experiments on time dependency of NaFluo uptake in suspended rat hepatocytes demonstrated linearity during the first 90 s of the uptake process (data not shown). Initial uptake rates were concentration dependent and could be described by a model incorporating Michaelis–Menten type kinetics including the Hill factor n (Fig. 1a). Mean (± SD, n = 3 batches of hepatocytes) estimates for Km and Vmax were 22.5 ± 4.4 µM and 98.3 ± 8.7 pmol/(million cells • min) and Hill factor 1.7 ± 0.6.
DISCUSSION
The aim of this study was to characterize the uptake mechanism and kinetics of NaFluo in suspended rat and human hepatocytes and in CHO cells transfected with hNTCP/rNtcp or the human hepatic OATP isoforms (OATP1B1, OATP1B3, and OATP2B1).
Our results support saturable NaFluo uptake in both rat and human hepatocytes, indicative of the involvement of one or more uptake transporters. Kinetic analysis of the uptake data revealed that the Km value for NaFluo uptake is comparable between rat and human
Acknowledgements
Tom De Bruyn received a PhD scholarship from the Agency for Innovation by Science and Technology, Flanders. This study was supported by grants from the following independent funders: “Fonds voor Wetenschappelijk Onderzoek” Flanders and “Onderzoeksfonds” of the Katholieke Universiteit Leuven, Belgium.
REFERENCES (46)
- et al.
Drug and bile acid transporters in rosuvastatin hepatic uptake: Function, expression, and pharmacogenetics
Gastroenterology
(2006) - et al.
Fluorometric screening for metabolism‐based drug–drug interactions
J Pharmacol Toxicol Methods
(2000) - et al.
Species‐specific interaction of HIV protease inhibitors with accumulation of cholyl‐glycylamido‐fluorescein (CGamF) in sandwich‐cultured hepatocytes
J Pharm Sci
(2010) - et al.
Development of a fluorescence‐based assay for screening of modulators of human organic anion transporter 1B3 (OATP1B3)
Eur J Pharm Biopharm
(2006) Fluorescence‐based assays for the assessment of drug interaction with the human transporters OATP1B1 and OATP1B3
Anal Biochem
(2010)- et al.
Transport of fluorescent chenodeoxycholic acid via the human organic anion transporters OATP1B1 and OATP1B3
J Lipid Res
(2006) - et al.
Measurement of bile salt export pump transport activities using a fluorescent bile acid derivative
Drug Metab Pharmacokinet
(2010) - et al.
Different pathways of canalicular secretion of sulfated and non‐sulfated fluorescent bile acids: A study in isolated hepatocyte couplets and TR‐ rats
J Hepatol
(1999) - et al.
Screening of OATP1B1/3 and OCT1 inhibitors in cryopreserved hepatocytes in suspension
Eur J Pharm Sci
(2010) - et al.
Function of uptake transporters for taurocholate and estradiol 17beta‐d‐glucuronide in cryopreserved human hepatocytes
Drug Metab Pharmacokinet
(2003)
Characterization of transport in isolated human hepatocytes: A study with the bile acid taurocholic acid, the uncharged ouabain and the organic cations vecuronium and rocuronium
Biochem Pharmacol
Sensitivity of bile acid transport by organic anion‐transporting polypeptides to intracellular pH
Biochim Biophys Acta.
Identification of multispecific organic anion transporter 2 expressed predominantly in the liver
FEBS Lett
Effect of pregnane X receptor ligands on transport mediated by human OATP1B1 and OATP1B3
Eur J Pharmacol
Coordinate induction of PPAR alpha and SREBP2 in multifunctional protein 2 deficient mice
Biochim. Biophys. Acta
Membrane transporters in drug development
Nat Rev Drug Discov
Xenobiotic transporters of the human organic anion transporting polypeptides (OATP) family
Xenobiotica
The role of the sodium‐taurocholate cotransporting polypeptide (NTCP) and of the bile salt export pump (BSEP) in physiology and pathophysiology of bile formation
Handb Exp Pharmacol
in vitro investigation of the hepatobiliary disposition mechanisms of the antifungal agent micafungin in humans and rats
Drug Metab Dispos
Drug transport in the liver
Rate‐limiting steps in hepatic drug clearance: Comparison of hepatocellular uptake and metabolism with microsomal metabolism of saquinavir, nelfinavir, and ritonavir
Drug Metab Dispos
Uptake is the rate‐limiting step in the overall hepatic elimination of pravastatin at steady‐state in rats
Pharm Res
in vitro evidence for the role of OATP and OCT uptake transporters in drug–drug interactions
Expert Opin Drug Metab Toxicol
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2021, Stem Cell ReportsCitation Excerpt :Because MRP4 showed a partial role in the basolateral export in BSEPR1090X i-Heps, we explored the possible contribution of other transporters that are capable of carrying bile acids. Among the SLC family, OST⍺/SLC51A, OSTβ/SLC51B, OATP3A1/SLCO3A1, and OATP1B3/SLCO1B3 are known to export conjugated bile acid from the basolateral domain (Alrefai and Gill, 2007; Ballatori et al., 2009; Briz et al., 2006; Bruyn et al., 2011; Pan et al., 2018). We found gene upregulation of OSTβ/SLC51B and OATP3A1/SLCO3A1 in BSEPR1090X i-Heps and downregulation of OST⍺/SLC51A (Figure 4E).