1521-0103/365/3/519–525$35.00 https://doi.org/10.1124/jpet.117.245993 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS J Pharmacol Exp Ther 365:519–525, June 2018 Copyright ª 2018 by The American Society for Pharmacology and Experimental Therapeutics In Vitro Assessment of Pharmacokinetic Drug-Drug Interactions of Direct Oral Anticoagulants: Type 5-Phosphodiesterase Inhibitors Are Inhibitors of Rivaroxaban and Apixaban Efflux by P-Glycoprotein Victor Margelidon-Cozzolino, Sophie Hodin, Elodie Jacqueroux, Olivier Delézay, Laurent Bertoletti, and Xavier Delavenne INSERM UMR 1059, Equipe Dysfonctions Vasculaires et de l’ Hémostase, Faculté de Médecine de St-Etienne, Université Jean Monnet, Saint-Etienne, France (V.M.-C., S.H., E.J., O.D., X.D.); and Service de Médecine Vasculaire et Thérapeutique, CHU de St-Etienne, Saint-Etienne, France (L.B.) Received October 30, 2017; accepted March 21, 2018 ABSTRACT Because of their lower bleeding risk and simplicity of use, direct oral anticoagulants (DOACs) could represent an interesting alternative to conventional anticoagulant treatment with vitamin K antagonists for patients with pulmonary arterial hypertension (PAH). P-glycoprotein (P-gp) plays a key role in DOAC pharmacokinetics. Type 5-phosphodiesterase inhibitors (PDE5is), a drug class commonly used in the treatment of PAH, have been shown to strongly inhibit P-gp. This work aimed to assess potential P-gp–mediated drug-drug interactions be- tween PDE5is and DOACs using in vitro methods. A cellular model of drug transport assay, using P-gp–overexpressing Madin-Darby canine kidney cells (transfected with the human P-gp gene), was used to determine the bidirectional permeabil- ities of two DOACs (rivaroxaban and apixaban) in the absence and presence of increasing concentrations (0.5–100 mM) of three PDE5is (sildenafil, tadalafil, and vardenafil). Permeabilities and efflux ratios were calculated from DOAC concentrations, were measured with liquid chromatography coupled with mass spectrometry, and were subsequently used to determine the PDE5i percentage of inhibition and half maximal inhibitory concentration (IC 50 ). Rivaroxaban efflux was inhibited by 99%, 66%, and 100% with 100 mM sildenafil, tadalafil, and vardenafil, respectively. Similarly, apixaban efflux was inhibited by 97%, 74%, and 100%, respectively. The IC 50 values of the three PDE5is were 8, 28, and 5 mM for rivaroxaban and 23, 15, and 3 mM for apixaban, respectively. This study showed strong in vitro inhibition of DOAC efflux by PDE5is. In vivo studies are required to determine the clinical relevance of these interactions. Introduction Anticoagulant therapy remains a challenging issue in the field of pulmonary hypertension treatment. Pulmonary arte- rial hypertension (PAH) is a vascular pulmonary disease characterized by elevated mean pulmonary arterial pressure that leads to right heart failure and death (Galiè et al., 2016). Oral anticoagulants, particularly vitamin K antagonists (VKAs), are widely used in PAH as a supportive therapy (e.g., in 60% of participants in a recent French cohort; Gabriel et al., 2016). The risk/benefit ratio of VKAs seems to be associated with a significant bleeding risk in these patients (Henkens et al., 2013). In addition to anticoagulants, patients with PAH receive specific drugs called “targeted therapies,” which include prostanoid analogs, endothelin receptor antagonists, and type 5-phosphodiesterase inhibitors (PDE5is) (Galiè et al., 2016). These targeted therapies are noticeably involved in drug-drug interactions (DDIs), especially with VKAs (Fernández and Romá, 2003; Ciracì et al., 2014; Said, 2014). Indeed, some of these drugs are P-glycoprotein (P-gp) and/or CYP3A4 sub- strates and can act as P-gp and CYP3A4 competitive inhibi- tors toward other drugs prescribed simultaneously. Interestingly, PDE5is were shown to be in vitro P-gp inhibi- tors when tested with chemotherapy in P-gp–overexpressing neoplastic cells (Ding et al., 2011; Shi et al., 2011). However, these studies only monitored intracellular accumulation of P-gp substrates and did not specifically study drug transport. Direct oral anticoagulants (DOACs) are direct specific activated factor X (anti-Xa) or thrombin (Anti-IIa) inhibitors, which are increasingly prescribed for approved indications This work was supported by La Fondation du Souffle and Le Fonds de Recherche en Santé Respiratoire. https://doi.org/10.1124/jpet.117.245993. ABBREVIATIONS: AF, atrial fibrillation; DDI, drug-drug interaction; DMSO, dimethylsulfoxide; DOAC, direct oral anticoagulation; ER, efflux ratio; FA, formic acid; FDA, U.S. Food and Drug Administration; MDCK, Madin-Darby canine kidney; m/z, mass-to-charge ratio; P-gp, P-glycoprotein; PAH, pulmonary arterial hypertension; P app A→B, apical-to-basolateral permeability; P app B→A, basolateral-to-apical permeability; PDE5i, type 5-phosphodiesterase inhibitor; RER, relative efflux ratio; TEER, transepithelial electric resistance; VKA, vitamin K antagonist; VTE, venous thromboembolism. 519 at ASPET Journals on May 31, 2020 jpet.aspetjournals.org Downloaded from