Vectorial Transport of Fexofenadine across Caco-2 Cells: Involvement of Apical Uptake and Basolateral Efflux Transporters

Abstract

Fexofenadine is a nonsedative antihistamine that exhibits good oral bioavailability despite its zwitterionic chemical structure and efflux by P-gp. Evidence exists that multiple uptake and efflux transporters play a role in hepatic disposition of fexofenadine. However, the roles of specific transporters and their interrelationship in intestinal absorption of this drug are unclear. This study was designed to elucidate vectorial absorptive transport of fexofenadine across Caco-2 cells involving specific apical uptake and efflux transporters as well as basolateral efflux transporters. Studies with cellular models expressing single transporters showed that OATP2B1 expression stimulated uptake of fexofenadine at pH 6.0. Apical uptake of fexofenadine into Caco-2 cells was decreased by 45% by pretreatment with estrone 3-sulfate, an OATP inhibitor, at pH 6.0 but not at pH 7.4, indicating that OATP2B1 mediates apical uptake of fexofenadine into these cells. Examination of fexofenadine efflux from preloaded Caco-2 cells in the presence or absence of (i) the MRP inhibitor MK-571 and (ii) the P-gp inhibitor GW918 showed that apical efflux is predominantly mediated by P-gp, with a small contribution by MRP2, whereas basolateral efflux is predominantly mediated by MRP3. These results also showed that while OSTαβ is functionally active in the basolateral membrane of Caco-2 cells, it does not play a role in the export of fexofenadine. MK-571 decreased the absorptive transport of fexofenadine by 17%. However, the decrease in absorptive transport by MK-571 was 42% when P-gp was inhibited by GW918. The results provide a novel insight into a vectorial transport system mainly consisting of apical OATP2B1 and basolateral MRP3 that may play an important role in delivering hydrophilic anionic and zwitterionic drugs such as pravastatin and fexofenadine into systemic circulation upon oral administration.

Zinc Finger Nuclease–Mediated Gene Knockout Results in Loss of Transport Activity for P-Glycoprotein, BCRP, and MRP2 in Caco-2 Cells

“…The potential involvement of a basolateral efflux transporter in the MRP family is supported by the observation that the addition of MK571 increased the efflux ratio in each of the cell lines lacking P-gp (up to 1.52 in the case of MDR1 KO cells). These data support the conclusions drawn by Ming et al (2011) that fexofenadine apical efflux in Caco-2 cells is predominantly mediated by P-gp, whereas basolateral efflux is predominantly mediated by MRP3. Based on data using MK571 and a P-gp/BCRP inhibitor (GW120918), Ming et al (2011) further suggested that MRP2 makes a small contribution to the apical efflux of fexofenadine, although our data using the MRP2 KO cell lines do not support the involvement of MRP2.…”

The Role of a Basolateral Transporter in Rosuvastatin Transport and Its Interplay with Apical Breast Cancer Resistance Protein in Polarized Cell Monolayer Systems

“…Furthermore, in contrast to MDCK cells, Caco-2 failed to sort recombinant NTCP-GFP protein into polarized surface expression on the plasma membrane and showed no polarity of taurocholate uptake (Sun et al, 2001), thus suggesting a lack of meaningful NTCP protein expression and function in Caco-2 systems. Both OATP1A2 and OATP2B1 are expressed at the apical membrane of enterocytes or Caco-2 cells (Sai et al, 2006;Glaeser et al, 2007), where they can contribute to the absorption of drugs such as statins, fexofenadine, levofloxacin, methotrexate, and talinolol (Badagnani et al, 2006;Ho et al, 2006;Maeda et al, 2007;Kitamura et al, 2008;Shirasaka et al, 2010;Ming et al, 2011). For the aforementioned reasons, it is highly unlikely that OATP1B1, OATP1B3, OATP2B1, OATP1A2, or NTCP could be involved in the basolateral uptake of rosuvastatin in Caco-2 cells.…”

“…Kinetic analysis of rosuvastatin basolateral uptake in Caco-2 at 37°C was performed by nonlinear regression analysis using GraphPad Prism 5.0 (GraphPad Software Inc., San Diego, CA). The data were fit to a model containing a saturable and a nonsaturable term (Ming et al, 2011):…”

Intestinal transporters for endogenic and pharmaceutical organic anions: the challenges of deriving in-vitro kinetic parameters for the prediction of clinically relevant drug–drug interactions

“…OATP2B1, the study can be performed at a lower pH (pH 5-6.5) to optimize Pcar. [36,[41][42][43] However, introducing a pH-gradient may be complex when only the anionized form is the exclusive substrate for the transporter since the decreased concentration of this species may reduce Pcar. Consequently, the Km or Ki may appear pH-dependent.…”

“…OATP2B1) may not be accounted for. [36,[41][42][43] Ideally, efflux ratios should be performed both in the presence and the absence of a pH-gradient. In the presence of a pH-gradient, the efflux ratios determined at a low and high concentration (high enough to saturate all transporters) of the ionizable drug substance could be compared to distinguish between transporter effects and pH-partition.…”