Organic Anion Transporting Polypeptide Inhibitor Research Articles

Coproporphyrins I and III as Functional Markers of OATP1B Activity: In Vitro and In Vivo Evaluation in Preclinical Species.

Inhibition of organic anion-transporting polypeptide (OATP)1B function can lead to serious clinical drug-drug interactions, thus a thorough evaluation of the potential for this type of interaction must be completed during drug development. Therefore, sensitive and specific biomarkers for OATP function that could be used in conjunction with clinical studies are currently in demand. In the present study, preclinical evaluations were conducted to characterize the suitability of coproporphyrins (CPs) I and III as markers of hepatic OATP functional activity. Active uptake of CPs I and III was observed in human embryonic kidney (HEK) 293 cells singly expressing human OATP1B1 (hOATP1B1), hOATP1B3, cynomolgus monkey OATP1B1 (cOATP1B1), or cOATP1B3, as well as human and monkey hepatocytes. Cyclosporin A (100 mg/kg, oral) markedly increased the area under the curve (AUC) plasma concentrations of CPs I and III by 2.6- and 5.2-fold, while rifampicin (15 mg/kg, oral) increased the AUCs by 2.7- and 3.6-fold, respectively. As the systemic exposure increased, the excretion of both isomers in urine rose from 1.6- to 4.3-fold in monkeys. In agreement with this finding, the AUC of rosuvastatin (RSV) in cynomolgus monkeys increased when OATP1B inhibitors were coadministered. In Oatp1a/1b gene cluster knockout mice (Oatp1a/1b(-/-)), CPs in plasma and urine were significantly increased compared with wild-type animals (7.1- to 18.4-fold; P < 0.001), which were also in agreement with the changes in plasma RSV exposure (14.6-fold increase). We conclude that CPs I and III in plasma and urine are novel endogenous biomarkers reflecting hepatic OATP function, and the measurements have the potential to be incorporated into the design of early clinical evaluation.

Transport of Corilagin, Gallic Acid, and Ellagic Acid from Fructus Phyllanthi Tannin Fraction in Caco-2 Cell Monolayers.

Objective. To investigate the absorption property of the representative hydrolyzable tannin, namely corilagin, and its hydrolysates gallic acid (GA) and ellagic acid (EA) from the Fructus Phyllanthi tannin fraction (PTF) in vitro. Methods. Caco-2 cells monolayer model was established. Influences of PTF on Caco-2 cells viability were detected with MTT assay. The transport across monolayers was examined for different time points, concentrations, and secretory directions. The inhibitors of P-glycoprotein (P-gp), multidrug resistance proteins (MRPs), organic anion transporting polypeptide (OATP) and sodium/glucose cotransporter 1 (SGLT1), and tight junction modulators were used to study the transport mechanism. LC-MS method was employed to quantify the absorption concentration. Results. The apparent permeability coefficient (P app) values of the three compounds were below 1.0 × 10−6 cm/s. The absorption of corilagin and GA were much lower than their efflux, and the uptake of both compounds was increased in the presence of inhibitors of P-gp and MRPs. The absorption of EA was decreased in the company of OATP and SGLT1 inhibitors. Moreover, the transport of corilagin, GA, and EA was enhanced by tight junction modulators. Conclusion. These observations indicated that the three compounds in PTF were transported via passive diffusion combined with protein mediated transport. P-gp and MRPs might get involved in the transport of corilagin and GA. The absorption of EA could be attributed to OATP and SGLT1 protein.

Near Infrared Imaging of Indocyanine Green Distribution in Pregnant Mice and Effects of Concomitant Medications.

The transfer of indocyanine green (ICG) across the placenta is considered to be very low based on measurements in fetal blood. The goal of this study was to evaluate in mice ICG's distribution within fetuses themselves and effects of concomitant medications on fetal exposure. Mid-gestational (day 12.5) and late-gestational (day 17.5) age mice were imaged after administration of ICG (0.167 mg), in the presence and the absence of the organic anion transporting polypeptide (OATP) inhibitor rifampin (10 mg/kg, n = 11, or 20 mg/kg, n = 1) or the P-glycoprotein inhibitor valspodar (12.5 mg/kg). In vivo ICG emission intensity was followed by ex vivo analysis of blood and tissue emission. Both valspodar and rifampin increased ICG's emission intensity within maternal tissues. In addition, valspodar enhanced the ex vivo signal in mid-pregnancy placentae (2.1-fold; p < 0.01) and fetuses (2.4-fold; p < 0.01), and reduced late-pregnancy placenta:blood and fetus:blood ratios. Rifampin increased placental (1.4-fold, p < 0.05, and 2.3-fold, p < 0.01, in mid- and late-pregnancy, respectively) and fetal (2.2-fold, p < 0.01, and 3.2-fold, p < 0.01, in mid- and late-pregnancy) ICG signal. Similarly to valspodar, late-pregnancy placenta:blood and fetus:blood ratios were reduced by rifampin. Both inhibitors enhanced ICG's emission in fetal leg, liver, and brain. In conclusion, ICG distribution into the mouse fetus can be enhanced when used concomitantly with OATP or P-glycoprotein inhibitors. The greater distribution within individual fetal tissues is likely related to ICG's greater transplacental transfer. Until further data are available on ICG's safety when combined with medications that affect its maternal handling, such combinations should be used with caution.

Evaluation of rosuvastatin as an organic anion transporting polypeptide (OATP) probe substrate: in vitro transport and in vivo disposition in cynomolgus monkeys.

Organic anion transporting polypeptides (OATPs) mediate hepatic drug uptake and serve as the loci of drug-drug interactions (DDIs). Consequently, there is a major need to develop animal models and refine in vitro-in vivo extrapolations. Therefore, the in vivo disposition of a model OATP substrate, [(3)H]rosuvastatin (RSV), was studied in the cynomolgus monkey and reported for the first time. After monkeys had received a 3-mg/kg oral dose, mass balance was achieved after bile duct cannulation (mean total recovery of radioactivity of 103.6%). Forty-two percent of the RSV dose was recovered in urine and bile, and the elimination pathways were similar to those reported for human subjects; 61.7%, 39.0%, and 2.9% of the dose was recovered in the feces, bile, and urine, respectively. The high levels of unchanged RSV recovered in urine and bile (26% of the dose) and the relatively low levels of metabolites observed indicated that RSV was eliminated largely by excretion. Also, for the first time, the in vitro inhibitory potential of cyclosporin A (CsA) toward cynomolgus monkey OATPs and sodium-taurocholate cotransporting polypeptide was studied in vitro (primary hepatocytes and transporter-transfected cells). It is concluded that one can study the CsA-RSV DDI in the cynomolgus monkey. For example, the in vitro IC50 values were within 2-fold (monkey versus human), and the increase (versus vehicle control) in the RSV AUC0-inf (6.3-fold) and Cmax (10.2-fold) with CsA (100 mg/kg) was similar to that reported for humans. The results further support the use of the cynomolgus monkey as a model to assess interactions involving OATP inhibition.

Using Improved Serial Blood Sampling Method of Mice to Study Pharmacokinetics and Drug–Drug Interaction

In pharmacokinetic evaluation of mice, using serial sampling methods rather than a terminal blood sampling method could reduce the number of animals needed and lead to more reliable data by excluding individual differences. In addition, using serial sampling methods can be valuable for evaluation of the drug-drug interaction (DDI) potential of drug candidates. In this study, we established an improved method for serially sampling the blood from one mouse by only one incision of the lateral tail vein, and investigated whether our method could be adapted to pharmacokinetic and DDI studies. After intravenous and oral administration of ibuprofen and fexofenadine (BCS class II and III), the plasma concentration and pharmacokinetic parameters were evaluated by our method and a terminal blood sampling method, with the result that both methods gave comparable results (ibuprofen: 63.8 ± 4.0% and 64.4%, fexofenadine: 6.5 ± 0.7% and 7.9%, respectively, in bioavailability). In addition, our method could be adapted to DDI study for cytochrome P450 and organic anion transporting polypeptide inhibition. These results demonstrate that our method can be useful for pharmacokinetic evaluation from the perspective of reliable data acquisition as well as easy handling and low stress to mice and improve the quality of pharmacokinetic and DDI studies.

Determinants of the stereoselective pharmacokinetics of fexofenadine

Drug transporters play an important role in the clinical pharmacokinetics of many therapeutic agents. Although it is estimated that about half of all therapeutic agents are chiral, there has been little information on the stereoselective pharmacokinetics related to drug transporters. This review focuses on the drug transporters contributing to the stereoselective pharmacokinetics of fexofenadine enantiomers in humans. Fexofenadine is administered clinically as a racemic mixture, and the plasma concentration of (R)-fexofenadine is about 1.5-fold higher than that of the (S)-enantiomer. Because fexofenadine is poorly metabolized by cytochrome P450s, its pharmacokinetics depends on its drug-transporter activities. First, we examined whether drug-transporter polymorphisms influence fexofenadine enantiomer pharmacokinetics. The findings suggested that a combination of multiple transporters involving organic anion transporting polypeptide (OATP) 2B1, P-glycoprotein (P-gp), and multidrug resistance-associated protein 2 (MRP2) react to stereoselective fexofenadine exposure. Subsequently, we evaluated the roles of P-gp and OATPs in fexofenadine enantiomer pharmacokinetics using these inducer/inhibitors. Coadministration of P-gp inducer/inhibitors significantly altered the pharmacokinetics of fexofenadine enantiomers. In addition, the OATP inhibitors rifampicin and apple juice also affected fexofenadine enantiomer pharmacokinetics. Moreover, in in vitro studies, the uptake of both fexofenadine enantiomers into OATP2B1 cRNA-injected oocytes was significantly higher than that into water-injected oocytes, and this effect was greater for (R)-fexofenadine. Taken together, these studies indicated that multiple transporters including P-gp, OATPs, and MRP2 play important roles in fexofenadine enantiomer pharmacokinetics. Furthermore, OATP2B1 is a key determinant of the stereoselective pharmacokinetics of fexofenadine, and drug transporters may have chiral discrimination ability.

Dehydroepiandrosterone sulfate, a useful endogenous probe for evaluation of drug–drug interaction on hepatic organic anion transporting polypeptide (OATP) in cynomolgus monkeys

Since drug–drug interaction (DDI) can affect organic anion-transporting polypeptide (OATP) and cause clinical events, prediction of such DDI is important in early clinical development. Although statins are useful probes for OATP-mediated DDI, endogenous probes would be more practical for predicting such DDI. In this study, we investigate the possible use of dehydroepiandrosterone sulfate (DHEAS), an endogenous OATP substrate, in predicting OATP-mediated DDI in cynomolgus monkeys as a first step toward in human assessment. In in vitro experiments, both human and cynomolgus monkey hepatocytes showed a time- and temperature-dependent DHEAS uptake. Rifampicin (RIF), a typical OATP inhibitor, inhibited this uptake, indicating the involvement of OATP in DHEAS uptake. In in vivo experiments, the area under the plasma concentration–time curve (AUC) and maximum plasma concentration (Cmax) of DHEAS were significantly increased following administration of RIF 10 mg/kg, although the extent of this increase was lower than that observed with the test-statins used in this study. However, based on the results of in vitro hepatic DHEAS uptake, changes in DHEAS concentration are expected to be more prominent in human than in monkey. This shows for the first time that DHEAS may be used as endogenous probe for predicting OATP-mediated DDI.

Fruit juice, organic anion transporting polypeptides, and drug interactions in psychiatry.

Organic anion transporting polypeptides (OATPs) are a group of membrane transport proteins that facilitate the influx of endogenous and exogenous substances across biological membranes. OATPs are found in enterocytes and hepatocytes and in brain, kidney, and other tissues. In enterocytes, OATPs facilitate the gastrointestinal absorption of certain orally administered drugs. Fruit juices such as grapefruit juice, orange juice, and apple juice contain substances that are OATP inhibitors. These fruit juices diminish the gastrointestinal absorption of certain antiallergen, antibiotic, antihypertensive, and β-blocker drugs. While there is no evidence, so far, that OATP inhibition affects the absorption of psychotropic medications, there is no room for complacency because the field is still nascent and because the necessary studies have not been conducted. Patients should therefore err on the side of caution, taking their medications at least 4 hours distant from fruit juice intake. Doing so is especially desirable with grapefruit juice, orange juice, and apple juice; with commercial fruit juices in which OATP-inhibiting substances are likely to be present in higher concentrations; with calcium-fortified fruit juices; and with medications such as atenolol and fexofenadine, the absorption of which is substantially diminished by concurrent fruit juice intake.

Effect of transporter inhibition on the distribution of cefadroxil in rat brain.

BackgroundCefadroxil, a cephalosporin antibiotic, is a substrate for several membrane transporters including peptide transporter 2 (PEPT2), organic anion transporters (OATs), multidrug resistance-associated proteins (MRPs), and organic anion transporting polypeptides (OATPs). These transporters are expressed at the blood–brain barrier (BBB), blood-cerebrospinal fluid barrier (BCSFB), and/or brain cells. The effect of these transporters on cefadroxil distribution in brain is unknown, especially in the extracellular and intracellular fluids within brain.MethodsIntracerebral microdialysis was used to measure unbound concentrations of cefadroxil in rat blood, striatum extracellular fluid (ECF) and lateral ventricle cerebrospinal fluid (CSF). The distribution of cefadroxil in brain was compared in the absence and presence of probenecid, an inhibitor of OATs, MRPs and OATPs, where both drugs were administered intravenously. The effect of PEPT2 inhibition by intracerebroventricular (icv) infusion of Ala-Ala, a substrate of PEPT2, on cefadroxil levels in brain was also evaluated. In addition, using an in vitro brain slice method, the distribution of cefadroxil in brain intracellular fluid (ICF) was studied in the absence and presence of transport inhibitors (probenecid for OATs, MRPs and OATPs; Ala-Ala and glycylsarcosine for PEPT2).ResultsThe ratio of unbound cefadroxil AUC in brain ECF to blood (Kp,uu,ECF) was ~2.5-fold greater during probenecid treatment. In contrast, the ratio of cefadroxil AUC in CSF to blood (Kp,uu,CSF) did not change significantly during probenecid infusion. Icv infusion of Ala-Ala did not change cefadroxil levels in brain ECF, CSF or blood. In the brain slice study, Ala-Ala and glycylsarcosine decreased the unbound volume of distribution of cefadroxil in brain (Vu,brain), indicating a reduction in cefadroxil accumulation in brain cells. In contrast, probenecid increased cefadroxil accumulation in brain cells, as indicated by a greater value for Vu,brain.ConclusionsTransporters (OATs, MRPs, and perhaps OATPs) that can be inhibited by probenecid play an important role in mediating the brain-to-blood efflux of cefadroxil at the BBB. The uptake of cefadroxil in brain cells involves both the influx transporter PEPT2 and efflux transporters (probenecid-inhibitable). These findings demonstrate that drug-drug interactions via relevant transporters may affect the distribution of cephalosporins in both brain ECF and ICF.

Effects of multiple-dose rifampicin 450 mg on the pharmacokinetics of fexofenadine enantiomers in Japanese volunteers.

Rifampicin is a potent inducer of P-glycoprotein (P-gp) and inhibitor of organic anion-transporting polypeptides (OATPs), with fexofenadine acting as a substrate for both mechanisms. Simultaneous administration of single- or multiple-dose rifampicin 600 mg significantly increases the concentrations of fexofenadine enantiomers by inhibiting OATP transporters. However, the effects of rifampicin 450 mg are unknown. Here, we evaluated the effects of multiple doses of rifampicin 450 mg on the pharmacokinetics of fexofenadine enantiomers in healthy Japanese volunteers. In this randomized, two-phase, double-blind crossover study, 10 healthy volunteers received rifampicin 450 mg/day or placebo for 7 days. On day 7, fexofenadine 60 mg was co-administered simultaneously. Rifampicin significantly increased the mean area under the plasma concentration-time curve (AUC) of (R)- and (S)-fexofenadine (3.10-fold and 3.48-fold, respectively) and decreased the renal clearance of (R)- and (S)-fexofenadine (0.40-fold and 0.47-fold, respectively), causing marked differences in the mean amounts of these enantiomers excreted into the urine in the rifampicin phase (P < 0.001). These results indicated that multiple doses of rifampicin 450 mg may be sufficient to inhibit the renal influx transporter and OATP-mediated hepatic uptake of both enantiomers. Moreover, these effects may be greater than the P-gp-inductive effects of rifampicin. Therefore, the interactive mechanism of multidose rifampicin may occur through a combination of OATP and P-gp transporters, thereby altering the pharmacokinetics of fexofenadine enantiomers. In this study of rifampicin 450 mg, the interactive magnitude of the mean AUC values of fexofenadine enantiomers was higher than that observed in the previous study of rifampicin 600 mg, and no dose-dependent inhibitory effects of rifampicin were observed. These effects may be clinically significant in patients receiving fexofenadine and rifampicin.

Clarithromycin substantially increases steady-state bosentan exposure in healthy volunteers

The aim of this study was to assess the effect of the cytochrome P450 (CYP) 3A4 and organic anion-transporting polypeptide (OATP) 1B1 inhibitor clarithromycin on the pharmacokinetics of bosentan. We also aimed to evaluate the impact of CYP2C9 and SLCO1B1 (encoding for OATP1B1) genotypes and their combination. We assessed the effect of the OATP and CYP3A inhibitor clarithromycin on bosentan pharmacokinetics at steady state and concurrently quantified changes of CYP3A activity using midazolam as a probe drug. Sixteen healthy volunteers received therapeutic doses of bosentan (125 mg twice daily) for 14 days and clarithromycin (500 mg twice daily) concomitantly for the last 4 days, and bosentan pharmacokinetics was assessed on days 1, 10 and 14. Clarithromycin significantly increased bosentan area under the plasma concentration-time curve of the dosing interval 3.7-fold and peak concentration 3.8-fold in all participants irrespective of the genotype. Clarithromycin also reduced CYP3A activity (midazolam clearance) in all participants; however, these changes were not correlated to the changes of bosentan clearance. Clarithromycin substantially increases the exposure to bosentan, suggesting that dose reductions may be necessary.

Investigation of the role of transporters on the hepatic elimination of an LAT1 selective inhibitor JPH203

JPH203 has been developed as an anticancer drug that inhibits L-type amino acid transporter 1-mediated essential amino acid uptake into tumor cells. This study sought to elucidate which drug transporters may be involved in JPH203 hepatic elimination, and to estimate human hepatic clearance. In Sprague-Dawley rats, JPH203 total body clearance approached blood flow rate. JPH203 biotransformation via phase II metabolism produces N-acetyl-JPH203 (NAc-JPH203). NAc-JPH203 accumulates in the bile, and NAc-JPH203 canalicular efflux was significantly decreased in Mrp2-deficient mutant rats (Eisai hyperbilirubinemic rats). JPH203 and NAc-JPH203 are organic anion transporters [organic anion transporting polypeptide (OATP)1B1, OATP1B3, OATP2B1, and OAT3] substrates. In human cryopreserved hepatocytes, JPH203 uptake was saturable and inhibited by rifampicin, a prototypical OATP inhibitor. JPH203 metabolic clearance was larger than influx clearance and eventually passive clearance; JPH203 uptake appears to be the rate-determining process in overall hepatic elimination. Furthermore, unlike rats, the human hepatic clearance was predicted to be intrinsic clearance rate limited. These results suggest that the hepatic uptake transporters are determinant factors to determine JPH203 systemic exposure.

Pitavastatin as an In Vivo Probe for Studying Hepatic Organic Anion Transporting Polypeptide-Mediated Drug–Drug Interactions in Cynomolgus Monkeys

Drug-drug interactions (DDIs) caused by the inhibition of hepatic uptake transporters such as organic anion transporting polypeptide (OATP) can affect therapeutic efficacy and cause adverse reactions. We investigated the potential utility of pitavastatin as an in vivo probe substrate for preclinically studying OATP-mediated DDIs using cynomolgus monkeys. Cyclosporine A (CsA) and rifampicin (RIF), typical OATP inhibitors, inhibited active uptake of pitavastatin into monkey hepatocytes with half-maximal inhibitory concentration values comparable with those in human hepatocytes. CsA and RIF increased the area under the plasma concentration-time curve (AUC) of intravenously administered pitavastatin in cynomolgus monkeys by 3.2- and 3.6-fold, respectively. In addition, there was no apparent prolongation of the elimination half-life of pitavastatin due to the decrease in both hepatic clearance and volume of distribution. These findings suggest that DDIs were caused by the inhibition of hepatic uptake of pitavastatin. CsA and RIF increased the AUC of orally administered pitavastatin by 10.6- and 14.8-fold, respectively, which was additionally caused by the effect of the CsA and RIF in the gastrointestinal tract. Hepatic contribution to the overall DDI for oral pitavastatin with CsA was calculated from the changes in hepatic availability and clearance, and it was shown that the magnitude of hepatic DDI was comparable between the present study and the clinical study. In conclusion, pharmacokinetic studies using pitavastatin as a probe in combination with drug candidates in cynomolgus monkeys are useful to support the assessment of potential clinical DDIs involving hepatic uptake transporters.

Substrate-Dependent Inhibition of Organic Anion Transporting Polypeptide 1B1: Comparative Analysis with Prototypical Probe Substrates Estradiol-17β-Glucuronide, Estrone-3-Sulfate, and Sulfobromophthalein

Organic anion transporting polypeptide (OATP) 1B1 plays an important role in the hepatic uptake of many drugs, and the evaluation of OATP1B1-mediated drug-drug interactions (DDIs) is emphasized in the latest DDI (draft) guidance documents from U.S. and E.U. regulatory agencies. It has been suggested that some OATP1B1 inhibitors show a discrepancy in their inhibitory potential, depending on the substrates used in the cell-based assay. In this study, inhibitory effects of 14 compounds on the OATP1B1-mediated uptake of the prototypical substrates [³H]estradiol-17β-glucuronide (E₂G), [³H]estrone-3-sulfate (E₁S), and [³H]sulfobromophthalein (BSP) were studied in OATP1B1-transfected cells. Inhibitory potencies of tested compounds varied depending on the substrates. Ritonavir, gemfibrozil, and erythromycin caused remarkable substrate-dependent inhibition with up to 117-, 14-, and 13-fold difference in their IC₅₀ values, respectively. Also, the clinically relevant OATP inhibitors rifampin and cyclosporin A exhibited up to 12- and 6-fold variation in their IC₅₀ values, respectively. Regardless of the inhibitors tested, the most potent OATP1B1 inhibition was observed when [³H]E₂G was used as a substrate. Mutual inhibition studies of OATP1B1 indicated that E₂G and E₁S competitively inhibited each other, whereas BSP noncompetitively inhibited E₂G uptake. In addition, BSP inhibited E₁S in a competitive manner, but E₁S caused an atypical kinetics on BSP uptake. This study showed substrate-dependent inhibition of OATP1B1 and demonstrated that E₂G was the most sensitive in vitro OATP1B1 probe substrate among three substrates tested. This will give us an insight into the assessment of clinically relevant OATP1B1-mediated DDI in vitro with minimum potential of false-negative prediction.

Effects of Selected OATP and/or ABC Transporter Inhibitors on the Brain and Whole-Body Distribution of Glyburide

Glyburide (glibenclamide, GLB) is a widely prescribed antidiabetic with potential beneficial effects in central nervous system injury and diseases. In vitro studies show that GLB is a substrate of organic anion transporting polypeptide (OATP) and ATP-binding cassette (ABC) transporter families, which may influence GLB distribution and pharmacokinetics in vivo. In the present study, we used [(11)C]GLB positron emission tomography (PET) imaging to non-invasively observe the distribution of GLB at a non-saturating tracer dose in baboons. The role of OATP and P-glycoprotein (P-gp) in [(11)C]GLB whole-body distribution, plasma kinetics, and metabolism was assessed using the OATP inhibitor rifampicin and the dual OATP/P-gp inhibitor cyclosporine. Finally, we used in situ brain perfusion in mice to pinpoint the effect of ABC transporters on GLB transport at the blood-brain barrier (BBB). PET revealed the critical role of OATP on liver [(11)C]GLB uptake and its subsequent impact on [(11)C]GLB metabolism and plasma clearance. OATP-mediated uptake also occurred in the myocardium and kidney parenchyma but not the brain. The inhibition of P-gp in addition to OATP did not further influence [(11)C]GLB tissue and plasma kinetics. At the BBB, the inhibition of both P-gp and breast cancer resistance protein (BCRP) was necessary to demonstrate the role of ABC transporters in limiting GLB brain uptake. This study demonstrates that GLB distribution, metabolism, and elimination are greatly dependent on OATP activity, the first step in GLB hepatic clearance. Conversely, P-gp, BCRP, and probably multidrug resistance protein 4 work in synergy to limit GLB brain uptake.

Pharmacokinetics of a Three-Way Drug Interaction Between Danoprevir, Ritonavir and the Organic Anion Transporting Polypeptide (OATP) Inhibitor Ciclosporin

Danoprevir (RG7227) is a potent macrocyclic inhibitor of the hepatitis C virus NS3/4A protease, which is currently in development in combination with low-dose ritonavir for the treatment of chronic hepatitis C infection. Danoprevir is a substrate of cytochrome P450 3A4, and the organic anion transporting polypeptides (OATP) 1B1 and 1B3. The objective of this study was to evaluate the effect of a potent OATP inhibitor, ciclosporin, on danoprevir pharmacokinetics, when administered as danoprevir/ritonavir. The effect of danoprevir/ritonavir on ciclosporin pharmacokinetics was also investigated. This was a single-dose, randomized, open-label, two-sequence, three-period, crossover study in healthy volunteers. In the first period, subjects were randomized to receive either a single oral dose of danoprevir 100mg in combination with ritonavir 100mg or a single oral dose of ciclosporin 100mg. After a 14-day washout, patients were crossed over to receive the opposite treatment. In period 3, all subjects received the combination of danoprevir/ritonavir and ciclosporin following a 14-day washout from period 2. Blood samples were collected serially with each dose for pharmacokinetic assessment. Pharmacokinetic parameters were estimated using non-compartmental analysis. Geometric mean ratios (GMRs) and 90% confidence intervals (CIs) were used to compare pharmacokinetic parameters [maximum concentration (C max), area under the concentration-time curve from time zero to infinity (AUC∞), and concentration 12h post-dose (C 12h)] of danoprevir/ritonavir and ciclosporin when administered alone or in combination. Measures of safety and tolerability were also evaluated. A total of 18 subjects were enrolled, and 17 completed the study. The C max, AUC∞, and C 12h GMRs (90% CI) when danoprevir/ritonavir and ciclosporin were co-administered versus danoprevir/ritonavir or ciclosporin alone were 7.22 (5.42-9.62), 13.6 (11.2-16.6), and 22.5 (17.4-29.3), respectively, for danoprevir, 1.97 (1.72-2.27), 2.23 (2.07-2.42), and 2.50 (2.22-2.81), respectively, for ritonavir, and 1.42 (1.29-1.57), 3.65 (3.27-4.08), and 6.15 (5.32-7.11), respectively, for ciclosporin. All treatments were well tolerated, with no laboratory abnormalities, and no clinically significant changes in vital signs, electrocardiograms, or physical examinations observed. A significant drug-drug interaction was observed between ciclosporin and danoprevir/ritonavir, leading to substantial increases in exposure to danoprevir and a lesser impact on exposure to ritonavir. Therefore, co-administration of danoprevir/ritonavir with potent OATP inhibitors should be undertaken with appropriate precautions.

Sorafenib Hepatobiliary Disposition: Mechanisms of Hepatic Uptake and Disposition of Generated Metabolites

Sorafenib is an orally active tyrosine kinase inhibitor used in the treatment of renal and hepatocellular carcinoma. This study was designed to establish whether transport proteins are involved in the hepatic uptake of sorafenib and to determine the extent of biliary excretion of sorafenib and its metabolites in human hepatocytes. Initial uptake was assessed in freshly isolated, suspended human hepatocytes in the presence of inhibitors and modulators. [(14)C]Sorafenib (1 µM) uptake at 4°C was reduced by about 61-63% of the uptake at 37°C, suggesting a high degree of passive diffusion. Hepatocyte uptake of [(14)C]sorafenib was not Na(+) dependent or influenced by the organic anion transporter 2 inhibitor ketoprofen. However, initial [(14)C]sorafenib hepatocyte uptake was reduced by 46 and 30% compared with control values in the presence of the organic anion transporting polypeptide inhibitor rifamycin SV and the organic cation transporter (OCT) inhibitor decynium 22, respectively. [(14)C]Sorafenib (0.5-5 µM) uptake was significantly higher in hOCT1-transfected Chinese hamster ovary cells compared with mock cells, and inhibited by the general OCT inhibitor, 1-methyl-4-phenylpryidinium. OCT1-mediated uptake was saturable with a Michaelis-Menten constant of 3.80 ± 2.53 µM and a V(max) of 116 ± 42 pmol/mg/min. The biliary excretion index and in vitro biliary clearance of sorafenib (1 µM) in sandwich-cultured human hepatocytes were low (∼11% and 11 ml/min/kg, respectively). Results suggest that sorafenib uptake in human hepatocytes occurs via passive diffusion, by OCT1, and by organic anion transporting polypeptide(s). Sorafenib undergoes modest biliary excretion, predominantly as a glucuronide conjugate(s).

Comparison of Inhibitory Duration of Grapefruit Juice on Organic Anion-Transporting Polypeptide and Cytochrome P450 3A4

Recently, a new type of interaction has been reported in which fruit juices diminish oral drug bioavailability through inhibition of organic anion-transporting polypeptide (OATP). In this study, we aimed to clarify the duration of OATP inhibition by grapefruit juice (GFJ), and to compare it with the duration of GFJ-induced inhibition of cytochrome P450 (CYP) 3A4 activity. Seven healthy volunteers were enrolled in this open-label, single-sequence study. They were orally administered celiprolol (100 mg) and midazolam (15 µg/kg) with water on the control day. Three days later, they ingested GFJ (200 mL) 3 times a day for 3 d. On day 1, the same drugs were administered with GFJ. On days 3 and 7, the same drugs were administered with water. Pharmacokinetics of both drugs were evaluated on each trial day. The peak plasma concentration (Cmax) and the area under the plasma concentration-time curve from 0 to 8 h (AUC0-8) of celiprolol significantly decreased on day 1, and the mean ratios of these values and the corresponding control-day values were 0.18 and 0.25, respectively. The Cmax and AUC0-8 returned to the control levels on days 3 and 7. In contrast, AUC0-8 of midazolam were higher on days 1 and 3 than on the control day (mean ratio, 2.12 and 1.47, respectively). The AUC0-8 returned to the control level on day 7. In conclusion, results of this study indicated that the OATP inhibition caused by GFJ dissipated faster than GFJ-mediated alterations in CYP3A4 activity, which were sustained for at least 48 h.

Classification of Inhibitors of Hepatic Organic Anion Transporting Polypeptides (OATPs): Influence of Protein Expression on Drug–Drug Interactions

The hepatic organic anion transporting polypeptides (OATPs) influence the pharmacokinetics of several drug classes and are involved in many clinical drug–drug interactions. Predicting potential interactions with OATPs is, therefore, of value. Here, we developed in vitro and in silico models for identification and prediction of specific and general inhibitors of OATP1B1, OATP1B3, and OATP2B1. The maximal transport activity (MTA) of each OATP in human liver was predicted from transport kinetics and protein quantification. We then used MTA to predict the effects of a subset of inhibitors on atorvastatin uptake in vivo. Using a data set of 225 drug-like compounds, 91 OATP inhibitors were identified. In silico models indicated that lipophilicity and polar surface area are key molecular features of OATP inhibition. MTA predictions identified OATP1B1 and OATP1B3 as major determinants of atorvastatin uptake in vivo. The relative contributions to overall hepatic uptake varied with isoform specificities of the inhibitors.

PET Imaging–Based Evaluation of Hepatobiliary Transport in Humans with (15R)-11C-TIC-Me

It is well accepted that drug transporters play a pivotal role in hepatobiliary excretion of anionic drugs, in which drug-drug interactions and genetic polymorphisms are known to cause variations. However, PET probes for in vivo functional characterization of these transporters have not been established yet. We used PET to investigate hepatic uptake and subsequent canalicular efflux of (11)C-labeled (15R)-16-m-tolyl-17,18,19,20-tetranorisocarbacyclin methyl ester [(15R)-(11)C-TIC-Me)] in healthy subjects. Serial PET scans of the abdominal region in healthy male subjects were obtained with or without the organic anion-transporting polypeptide (OATP) inhibitor rifampicin after intravenous injection of (15R)-(11)C-TIC-Me as a radiotracer. Venous blood samples and PET images were obtained at frequent intervals up to 30 min after administration of the PET tracer. Dynamic imaging data were evaluated by integration plots of data collected for 2-10 min and for 10-30 min after tracer administration for the determination of tissue uptake clearance and biliary efflux clearance, respectively. After rapid hydrolysis in blood, the acid form-(11)C-labeled (15R)-16-m-tolyl-17,18,19,20-tetranorisocarbacyclin [(15R)-(11)C-TIC]-accumulated in the liver (37% of the dose by 17 min), and the radioactivity was then excreted into the bile (6.2% by 30 min). Rifampicin (600 mg by mouth), a potent OATP inhibitor, significantly reduced the radioactivity excreted into the bile (by 44%) by inhibiting both uptake (by 45%) and subsequent canalicular efflux (by 62%). (15R)-(11)C-TIC is an in vitro substrate of OATP1B1 and OATP1B3, and clinically relevant concentrations of rifampicin inhibited uptake by OATP1B1 and OATP1B3. These results demonstrated that in humans, (15R)-(11)C-TIC-associated radioactivity is excreted into the bile by organic anion transport systems. We demonstrated that PET image analysis with (15R)-(11)C-TIC-Me is useful for investigating variations in OATP function in the human hepatobiliary transport system.