American Chemical Society and American Pharmaceutical Association
Journal of Pharmaceutical Sciences vol:100 issue:11 pages:5018-5030
The aim of this study was to characterize the in vitro hepatic uptake kinetics of NaFluo and identify the transporters involved. NaFluo exhibited saturable uptake kinetics in suspended rat and human hepatocytes as reflected by Km values of 22.5 and 14.1 µM, and Vmax values of 98.3 and 5.8 pmol/million cells/min, respectively. Co-incubation with known inhibitors (e.g. rifampicin) of OATP/Oatp (Organic Anion Transporting Polypeptide; SLCO gene family) significantly decreased NaFluo uptake in hepatocytes. In contrast, neither inhibitors/substrates of the OCT/Oct or OAT/Oat 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 CHO cells transfected with OATP1B1, 1B3 and 2B1. Transporter-mediated uptake of NaFluo was observed in OATP1B1- and 1B3-transfected cells (Km = 4.2 and 10.9 µM; Vmax = 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 to the unconjugated compounds.