title: Sodium fluorescein is a probe substrate for hepatic drug transport mediated by OATP1B1 and OATP1B3.
abstract: The aim of this study was to characterize the in vitro hepatic uptake kinetics of sodium fluorescein (NaFluo) and identify the transporters involved. NaFluo exhibited saturable uptake kinetics in suspended rat and human hepatocytes as reflected by K(m) values of 22.5 and 14.1 µM, and V(max) values of 98.3 and 5.8 pmol/(million cells • min), respectively. Coincubation with known inhibitors (e.g. rifampicin) of organic anion transporting polypeptide (OATP/Oatp; SLCO gene family) significantly decreased NaFluo uptake in hepatocytes. In contrast, neither inhibitors/substrates of the organic cation transporter or organic anion transporter family nor depletion of extracellular sodium resulted in significant inhibition of NaFluo uptake. To explore the contribution of individual uptake transporters, NaFluo uptake was determined in Chinese hamster ovary cells transfected with OATP1B1, OATP1B3, and OATP2B1. Transporter-mediated uptake of NaFluo was observed in OATP1B1- and OATP1B3-transfected cells (K(m) = 4.2 and 10.9 µM; V(max) = 30.9 and 135 [pmol/(mg protein • min)], respectively). NaFluo can be used as a probe substrate to study Oatp/OATP1B-mediated drug interactions in fluorescence-based in vitro transport assays of rat and human liver. Labeling of drugs or bile salts with a fluorescein moiety can be expected to result in fluorescent conjugates with substantially altered hepatic uptake characteristics as compared with the unconjugated compounds.
authors.affiliation: Department of Pharmaceutical Sciences, Laboratory for Pharmacotechnology and Biopharmacy, Katholieke Universiteit Leuven, 3000 Leuven, Belgium.
authors.forename: Tom
authors.initials: T
authors.lastname: De Bruyn
authors.forename: Sarinj
authors.initials: S
authors.lastname: Fattah
authors.forename: Bruno
authors.initials: B
authors.lastname: Stieger
authors.forename: Patrick
authors.initials: P
authors.lastname: Augustijns
authors.forename: Pieter
authors.initials: P
authors.lastname: Annaert
mesh_terms: D000818:Animals; D001692:Biological Transport; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D019793:Fluorescein / Q000493:pharmacokinetics*; D017320:HIV Protease Inhibitors / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000378:metabolism*; D027381:Liver-Specific Organic Anion Transporter 1; D008297:Male; D027361:Organic Anion Transporters / Q000502:physiology*; D029362:Organic Anion Transporters, Sodium-Independent / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D060888:Real-Time Polymerase Chain Reaction; D000074542:Solute Carrier Organic Anion Transporter Family Member 1B3
chemical_list: D017320:HIV Protease Inhibitors; D027381:Liver-Specific Organic Anion Transporter 1; D027361:Organic Anion Transporters; D029362:Organic Anion Transporters, Sodium-Independent; C503999:SLCO1B1 protein, human; C421209:SLCO1B3 protein, human; D000074542:Solute Carrier Organic Anion Transporter Family Member 1B3; D019793:Fluorescein