It has been reported that both nicardipine and lovastatin are substrates of both the cytochrome P450 (CYP) 3A subfamily and P-glycoprotein (P-gp), and P-gp transport is unlikely to be a significant factor. Thus, the effects of oral lovastatin on the pharmacokinetics of intravenous and oral nicardipine were investigated in rats. Nicardipine was administered intravenously (4 mg/kg) and orally (12 mg/kg) with 0 (control), 0.3 and 1 mg/kg of oral lovastatin to rats. Lovastatin was administered 30 min before nicardipine administration. After intravenous administration of nicardipine with 0, 0.3 and 1 mg/kg of lovastatin, the total areas under the plasma concentration-time curve from time zero to infinity (AUCs) of nicardipine were not changed by lovastatin. However, after oral administration of nicardipine with 1 mg/kg of oral lovastatin, the AUC of nicardipine was significantly greater (by 67.4%), and the extent of absolute oral bioavailability (F) of nicardipine was increased (by 38.5%). The above data suggest that lovastatin did not considerably inhibit the metabolism of nicardipine via the hepatic CYP3A subfamily, but inhibited intestinal P-gp and/or the CYP3A subfamily. Copyright (c) 2010 John Wiley & Sons, Ltd.

The clinical assessment of new formulations of human insulin is problematic due to the inability to distinguish between endogenous insulin and exogenously administered insulin. The usual methods to surmount the problem of distinguishing between endogenous and exogenous human insulin include evaluation in subjects with no or little endogenous insulin, hyper-insulinemic clamp studies or the administration of somatostatin to suppress endogenous insulin secretion. All of these methods have significant drawbacks. This paper describes a method for C-Peptide correction based upon a mixed effects linear regression of multiple time point sampling of C-Peptide and insulin. This model was able to describe each individual's insulin to C-Peptide relationship using the data from four different phase I clinical trials involving both subjects with and without type 2 diabetes in which insulin and C-Peptide were measured. These studies used hyper-insulinemic euglycemic clamps or meal challenges and subjects received insulin or Glucagon-like peptide 1 (GLP-1). It was possible to determine the exogenously administered insulin concentration from the measured total insulin concentration. A simple statistical technique can be used to determine each individual's insulin to C-Peptide relationship to estimate exogenous and endogenous insulin following the administration of regular human insulin. This technique will simplify the assessment of new formulations of human insulin. Copyright (c) 2010 John Wiley & Sons, Ltd.

A diabetic patient may suffer simultaneously from cardiovascular disease; thus, lipid-lowering or anti-hypertensive agents could be given together with nateglinide. The pharmacokinetics of nateglinide were investigated in the presence and absence of HMG-CoA reductase inhibitors (fluvastatin, lovastatin) and calcium channel blockers (verapamil, nifedipine) in rabbits. A pharmacokinetic modeling approach was used to quantify the effects of the drugs that significantly influenced the pharmacokinetics of nateglinide. Fluvastatin and nifedipine shifted the time course of serum nateglinide concentrations upwards; there was no significant change with verapamil or lovastatin. The C(max) and AUC(inf) increased 1.5- (p<0.05) and 1.3-fold in the presence of fluvastatin and 1.8- (p<0.01) and 2.4-fold (p<0.01) in the presence of nifedipine, respectively. In a simultaneous nonlinear regression, fluvastatin and nifedipine decreased the elimination rate constant, by 76% and 32%, respectively. Fluvastatin and nifedipine increased the systemic exposure of nateglinide in rabbits, probably due to their inhibitory action on the metabolism of nateglinide by CYP2C5 (human CYP2C9). The concomitant use of fluvastatin and/or nifedipine with nateglinide is quite likely; therefore, the clinical consequences of long-term treatments must be considered. Copyright (c) 2010 John Wiley & Sons, Ltd.

The pharmacokinetic (PK) and pharmacodynamic (PD) parameters of ONO-4641 in humans were estimated using preclinical data in order to provide essential information to better design future clinical studies. The characterization of PK/PD was measured in terms of decreased lymphocyte counts in blood after administration of ONO-4641, a sphingosine 1-phosphate receptor modulator. Using a two-compartment model, human PK parameters were estimated from preclinical PK data of cynomolgus monkey and in vitro human metabolism data. To estimate human PD parameters, the relationship between lymphocyte counts and plasma concentrations of ONO-4641 in cynomolgus monkeys was determined. The relationship between lymphocyte counts and plasma concentrations of ONO-4641 was described by an indirect-response model. The indirect-response model had an I(max) value of 0.828 and an IC(50) value of 1.29 ng/ml based on the cynomolgus monkey data. These parameters were used to represent human PD parameters for the simulation of lymphocyte counts. Other human PD parameters such as input and output rate constants for lymphocytes were obtained from the literature. Based on these estimated human PK and PD parameters, human lymphocyte counts after administration of ONO-4641 were simulated. In conclusion, the simulation of human lymphocyte counts based on preclinical data led to the acquisition of useful information for designing future clinical studies. Copyright (c) 2010 John Wiley & Sons, Ltd.

Purpose. Compared with traditional macroemulsion propofol formulations currently in clinical use, microemulsion formulations of this common intravenous anesthetic may offer advantages. The pharmacokinetics and coagulation effects as assessed by thromboelastography of these formulations were characterized in swine. Methods. Yorkshire swine (20-30 kg, either sex, n=15) were sedated, anesthetized with isoflurane, and instrumented to obtain a tracheostomy, internal jugular access and carotid artery catheterization. Propofol (2 mg/kg, 30 s) was administered as a macroemulsion (10 mg/ml; Diprivan((R)); n=7) or a custom (2 mg/kg, 30 s) microemulsion (10 mg/ml; n=8). Arterial blood specimens acquired pre- and post-injection (1 and 45 min) were used for thromboelastography. Arterial blood specimens (n=12 samples/subject, 60 min) were serially collected, centrifuged and analysed with solid-phase extraction with UPLC to determine propofol plasma concentrations. Non-compartmental pharmacokinetic analysis was applied to plasma concentrations. Results. No changes were noted in the thromboelastographic R time (p=0.74), K time (p=0.41), alpha angle (p=0.97), or maximal amplitude (p=0.71) for either propofol preparation. Pharmacokinetic parameters k (p=0.45), t(1/2) (p=0.26), C(o) (p=0.89), AUC(0-infinity) (p=0.23), CL (p=0.14), MRT (p=0.47), V(ss) (p=0.11) of the two formulations were not significantly different. Conclusion. The microemulsion and macroemulsion propofol formulations had similar pharmacokinetics and did not modify thromboelastographic parameters in swine. Copyright (c) 2010 John Wiley & Sons, Ltd.

The present study explored the feasibility of a differential setup for the in situ perfusion technique with mesenteric cannulation in rats to assess drug interactions at the level of intestinal absorption. In contrast to the classic, parallel in situ perfusion setup, the differential approach aims to identify intestinal drug interactions in individual animals by exposing the perfused segment to a sequence of multiple conditions. First, the setup was validated by assessing the interaction between the P-glycoprotein (P-gp) inhibitor verapamil and the transport probes atenolol (paracellular transport), propranolol (transcellular) and talinolol (P-gp mediated efflux). While transport of atenolol and propranolol remained constant for the total perfusion time (2 h), a verapamil-induced increase in talinolol transport was observed within individual rats (between 3.2- and 5.2-fold). In comparison with the parallel setup, the differential in situ perfusion approach enhances the power to detect drug interactions with compounds that exhibit strong subject-dependent permeability. This was demonstrated by identifying an interaction between amprenavir and ketoconazole (P-gp and CYP3A inhibitor) in five out of seven rats (permeability increase between 1.9- and 4.2-fold), despite high inter-individual differences in intrinsic permeability for amprenavir. In combination with an increased throughput (up to 300%) and a reduced animal use (up to 50%), the enhanced power of the differential approach improves the utility of the biorelevant in situ perfusion technique with mesenteric blood sampling to elucidate the intestinal interaction profile of drugs and drug candidates.

The pharmacokinetics (PK) and pharmacodynamics (PD) of recombinant human erythropoietin (rHuEPO) upon its repeated administrations were investigated. Two groups (A and B) of normal Wistar rats received rHuEPO intravenously at 450 or 1350 IU/kg thrice weekly for 2 and 6 weeks. PK studies were conducted following days 0 and 4 for group (A) and days 0, 17 and 28 for group (B), then, washout PK were assessed on days 11 and 36 for both groups. Reticulocytes (RET), red blood cells (RBC) and hemoglobin (Hb) were evaluated daily until day 14, then every 2 days until day 30 for group (A) and 59 for group (B). The total clearance CL(Total) increased with the dose but decreased over time. Its decay reached 20% and 55% between the first and last full PK in both treatment arms. RET peaked on day 5 and were 77.6% and 87.3% higher than baselines for the two dosing regimen. Their nadirs occurred on days 22 and 55 and were 37.9% and 47.3% below normal values. Hb peaked on days 10 and 34 and was 28.9% and 38.6% above the baseline level, its nadirs occurred on days 25 and 57 and were 13.1% and 16% below baselines. Control animals showed stable baselines over the study but with moderate variability. In conclusion, rHuEPO exhibits a nonlinear PK with a time-dependent decrease of its CL(Total). During exposure, RET, RBC and Hb showed a tolerance effect. After exposure, the rebound was characterized for RET, RBC, but not Hb. Copyright (c) 2010 John Wiley & Sons, Ltd.

The pharmacokinetics of SP-8203, a potential protective agent for the treatment of cerebral infarction, were evaluated after its intravenous (10, 20 and 30 mg/kg) and oral (10, 20, 30 and 100 mg/kg) administration in rats. After the intravenous administration of SP-8203, the AUCs of SP-8203 were dose-dependent; the dose-normalized AUCs were significantly greater with increasing doses. After the oral administration of SP-8203, plasma concentrations of SP-8203 were much lower than those after intravenous administration. This could be due to considerable hepatic and intestinal metabolism and the high percent of the dose recovered from the gastrointestinal tract (including its contents and feces) at 24 h as unchanged drug. Copyright (c) 2010 John Wiley & Sons, Ltd.

The objective was to develop a population pharmacokinetic-pharmacodynamic model of caffeine's psychomotor effects in healthy, non-habitual users of caffeine. Twenty Chinese males each received a single dose of 250 mg of caffeine orally. Plasma concentrations of caffeine were determined at various times within 24 h after dosing. The subjects' psychomotor performance was evaluated before and at various times after dosing by a test battery consisting of oculomotor assessment (saccadic velocity) as well as the computerised Swedish Performance Evaluation System. Nonlinear mixed-effects modelling to analyse the pharmacokinetic-pharmacodynamic relationships was performed using NONMEM. Model robustness was assessed by a nonparametric bootstrap. The results showed that caffeine caused significant improvements in psychomotor functioning. The time course of these effects was best described by pharmacokinetic/pharmacodynamic models involving an effect compartment. The transfer half-lives between plasma and effect site for different domains of psychomotor functioning were in the range 24.8-49.5 min. Evaluation of the final models showed close agreement between pairs of bootstrapped and final model parameter estimates (all differences<10%). These results provided the first suggestive evidence that caffeine effects on psychomotor performance occur after some time delay relative to changes in plasma caffeine concentration. The models for the neurobehavioural tests provided similar transfer half-lives between plasma and effect site. Copyright (c) 2010 John Wiley & Sons, Ltd.

Introduction. Sotrastaurin is an immunosuppressant that inhibits protein kinase C and blocks T-lymphocyte activation. The authors determined the effect of combining sotrastaurin with the calcineurin inhibitor cyclosporine on the pharmacokinetics and biomarker responses to both drugs. Methods. This was a randomized, 4-period, crossover study in 20 healthy subjects who received single oral doses of (1) sotrastaurin 100 mg, (2) cyclosporine 400 mg, (3) 100 mg sotrastaurin with 100 mg cyclosporine and (4) 100 mg sotrastaurin with 400 mg cyclosporine. Blood samples were collected to measure drug levels and biomarkers of T-lymphocyte activation (interleukin-2 and tumor necrosis factor producing T-cells and interleukin-2 messenger RNA levels) and of T-lymphocyte proliferation (thymidine uptake). Results. Sotrastaurin did not alter cyclosporine AUC; however, low-dose and high-dose cyclosporine increased sotrastaurin AUC by 1.2-fold [90% confidence interval, 1.1-1.4] and 1.8-fold [1.6-2.1], respectively. Adding high-dose cyclosporine to a low-therapeutic dose of sotrastaurin significantly enhanced the inhibition of cytokine production by 31% [95% confidence interval, 25-36%], of interleukin-2 messenger RNA levels by 13% [7-19%], and of thymidine uptake by 37% [32-42%] compared with sotrastaurin alone. Addition of low-dose cyclosporine elicited slightly lower enhancements in inhibition by 21% [14-28%], 6% [-4-16%], and 26% [21-30%], respectively, compared with sotrastaurin alone. Conclusions. Sotrastaurin did not alter the pharmacokinetics of cyclosporine, but cyclosporine increased sotrastaurin AUC up to 1.8-fold. The combined drugs elicited a significantly greater inhibition of T-cell activation and proliferation than sotrastaurin alone. Copyright (c) 2010 John Wiley & Sons, Ltd.

Racemic licarbazepine (Lic) is the active metabolite of oxcarbazepine (OXC) and eslicarbazepine acetate (ESL), appearing in human plasma as S-licarbazepine (S-Lic) and R-licarbazepine (R-Lic). However, human metabolism of OXC and ESL to Lic differs in the S-Lic/R-Lic enantiomeric ratio observed in plasma. S-Lic appears in higher proportion after ESL administration than after OXC (95% versus 80%). Enantioselective pharmacokinetics of Lic enantiomers have been found in mice after their separate administration and in humans following OXC treatment. Since protein binding of drugs may be enantioselective and a determining factor of pharmacokinetics, the binding of S-Lic and R-Lic to mouse and human total plasma proteins and, specifically, to human serum albumin (HSA) and alpha(1)-acid glycoprotein (AGP) were herein investigated for the first time. Free and bound fractions of S-Lic and R-Lic were separated by ultrafiltration after previous in vitro incubation of spiked plasma samples and protein solutions with each enantiomer at 10, 25 and 50 microg/ml. The results revealed that the extent of binding of Lic enantiomers to total plasma proteins was 30% and independent of the drug concentration and species considered. The data also suggest that the binding of Lic enantiomers to HSA is greater than that to AGP. Moreover, absence of enantioselectivity in the binding of Lic enantiomers to mouse and human plasma proteins and to HSA and AGP is evident. In conclusion, these findings suggest that the enantioselectivity observed in vivo in the biodisposition of S-Lic and R-Lic is not dependent on their affinity to plasma proteins. Copyright (c) 2010 John Wiley & Sons, Ltd.

The objective of this study was to predict R(b) (blood/plasma ratio) in humans using a simple method. Human and rat R(b) and free fraction in plasma (f(p)) values were obtained from the literature. The ratio of total red blood cell concentration to the free concentration in plasma (K(b)) was calculated using f(p) and R(b). Four methods were used for the prediction of R(b): (A) use of rat R(b); (B) use of R(b) calculated from rat K(b) and human f(p); (C) correlation of human log ((1-f(p))/f(p)) and human log K(b); and (D) correlation of log D with human log K(b). The R(b) of 96 compounds in humans ranged from 0.52 to 2.00, with an average of 0.89. A significant correlation was observed among human log K(b), human log ((1-f(p))/f(p)), and log D; however, no obvious correlation was observed among human R(b), human log ((1-f(p))/f(p)), and log D. The errors within 1.25-fold for methods A-D were 68.3%, 77.6%, 61.5% and 64.8%, respectively. All predictive methods considered here were superior to the use of the average value of human R(b) or R(b)=1. Rat R(b) corrected by human f(p) improved the accuracy of the prediction. Method B was the most accurate of the four methods. Copyright (c) 2010 John Wiley & Sons, Ltd.

Asenapine is a psychopharmacologic agent approved in the United States for the acute treatment of schizophrenia in adults and the acute treatment of manic or mixed episodes associated with bipolar I disorder with or without psychotic features in adults. It is pending approval for the treatment of schizophrenia and manic episodes associated with bipolar I disorder in Europe. Asenapine is administered as a sublingual formulation. To determine whether the pharmacokinetics of asenapine are impacted by placing the tablet buccally ('cheeking') or allowing the tablet to dissolve on the top of the tongue, pharmacokinetics were compared following buccal and supralingual administration versus sublingual administration. In this open-label, randomized, 3-way crossover trial, healthy men (n=36) received single 5 mg doses of asenapine via sublingual, supralingual and buccal routes, at least 1 week apart. With buccal administration, the area under the concentration-over-time curve (AUC(0-infinity)) and peak concentration (C(max)) were, respectively, 24%, and 19% higher than with sublingual administration; these routes were not bioequivalent. With supralingual administration, AUC(0-infinity) and C(max) were 6% and 13% lower than with sublingual administration; bioequivalence was established based on AUC(0-infinity) only; bioequivalence based on C(max) could not be assessed due to 40% within-subject variability. The most common adverse events were oral paresthesia (sublingual, 75.8%; supralingual, 55.9%; buccal, 45.7%) and somnolence (81.8%; 76.5%; 68.6%). Compared with the recommended sublingual route of asenapine administration, exposure was 24% higher with buccal administration and comparable to supralingual administration. However, differences in exposure associated with variable placement in the oral cavity did not compromise safety in healthy subjects. Copyright (c) 2010 John Wiley & Sons, Ltd.

The aim of our study was to investigate the effect of biochanin A on the accumulation and transport of mitoxantrone in breast cancer resistance protein (BCRP)-expressing normal cells and its impact on the pharmacokinetics (PK) and tissue distribution of mitoxantrone. In accumulation studies, the intracellular level of mitoxantrone was significantly increased in the presence of 2.5 or 25 microM of biochanin A in both murine and human BCRP-expressing Madin-Darby canine kidney (MDCK) cells, with no effect in corresponding MDCK/Mock cells. In bi-directional transport studies, the P(app,B-A) value of mitoxantrone with biochanin A co-treatment was much lower (6.66+/-0.84x10(-7) cm/s) than that in the absence of biochanin A (21.4+/-4.14x10(-7) cm/s), indicating inhibition of Bcrp1-mediated efflux. To evaluate whether our in vitro results might translate into an in vivo interaction, mitoxantrone PK and tissue distribution, with and without co-administration of biochanin A, was investigated. In contrast to our in vitro results, biochanin A (10 mg/kg, i.v.) had no impact on the concentration of mitoxantrone in plasma and most tissues collected (brain, heart, liver and lung). Surprisingly, the concentrations of mitoxantrone in spleen and kidney were even decreased when biochanin A was co-administered. Interestingly, it was found that the intracellular fluorescence of mitoxantrone was decreased 31.9% when co-incubated with 10 microM biochanin A in P-glycoprotein (P-gp) expressing MCF-7/ADR cells, indicating potential P-gp stimulation. The species difference of the inhibitory effect of biochanin A on BCRP, the extensive metabolism of biochanin A, as well as the stimulation effect of biochanin A on P-gp, may contribute to this in vitro-in vivo disconnect. Copyright (c) 2010 John Wiley & Sons, Ltd.

Paclitaxel is a substrate of the efflux transporters such as P-glycoprotein, and is mainly metabolized by the liver. Schisandrol B (Sch B), one of the active components in Schisandra, has been reported to be able to inhibit the activity of P-gp and CYP3A. It might be possible that Sch B would alter the pharmacokinetic behavior of paclitaxel. Therefore, the purpose of this study was to investigate the effect of Sch B on the pharmacokinetics of paclitaxel administered orally and intravenously in rats. Paclitaxel were administered to rats orally (30 mg/kg) or intravenously (0.5 mg/kg) with or without the concomitant administration of Sch B (10 or 25 mg/kg). Oral pharmacokinetic parameters of paclitaxel were significantly altered when pretreated with Sch B. There were significant increases in AUC(0-24h) (from 297.7+/-110.3 to 838.9+/-302.1 h*ng/ml; p<0.05) and C(max) (from 51.7+/-20.1 to 136.4+/-35.5 ng/ml; p<0.05) in the presence of Sch B (25 mg/kg). The pharmacokinetic parameters for i.v. paclitaxel were not significantly affected by Sch B in contrast to that of oral administration. Since the presence of Sch B enhanced the systemic exposure of paclitaxel, their pharmacokinetic interaction should be taken into consideration. As the oral bioavailability of paclitaxel was increased about 3-fold in the presence of Sch B, the concomitant use of Sch B may provide a benefit in the oral delivery of paclitaxel.

In this report, the utility of a commonly used interspecies scaling method to predict the systemic clearance (CL) of therapeutic proteins in humans was evaluated. Based on analysis of a pharmacokinetic data set of 34 therapeutic proteins, including 12 monoclonal antibodies (mAbs) and Fc fusion proteins, human CL can generally be predicted reasonably well with simple allometric scaling and a fixed exponent of 0.8: approximately 95% of the cases predicted values within 2-fold of the observed values when using CL data from multiple species, or approximately 90% simply using CL from monkeys. Specific to mAbs/Fc fusion proteins, scaling from monkey CL using a fixed exponent of 0.8 gave an excellent prediction; all predicted CL values were within 2-fold of the corresponding observed values. Compared with the simple allometric scaling method that uses a fitted exponent from CL data of > or =3 preclinical species, the fixed exponent approach with 1-2 preclinical species is simple, resource-saving and minimizes systematic bias. Together with its overall satisfactory prediction accuracy, especially in the absence of non-linear pharmacokinetics and species-specific clearance mechanisms, this fixed exponent method affords a viable alternative to other published allometric methods, including the Rule of Exponents (ROE).

Naloxone, a potent and specific opioid antagonist, has been shown in previous studies to have an influx clearance across the rat blood-brain barrier (BBB) two times greater than the efflux clearance. The purpose of the present study was to characterize the influx transport of naloxone across the rat BBB using the brain uptake index (BUI) method. The initial uptake rate of [(3)H]naloxone exhibited saturability in a concentration-dependent manner (concentration range 0.5 microM to 15 mM) in the presence of unlabeled naloxone. These results indicate that both passive diffusion and a carrier-mediated transport mechanism are operating. The in vivo kinetic parameters were estimated as follows: the Michaelis constant, K(t), was 2.99+/-0.71 mM; the maximum uptake rate, J(max), was 0.477+/-0.083 micromol/min/g brain; and the nonsaturable first-order rate constant, K(d), was 0.160+/-0.044 ml/min/g brain. The uptake of [(3)H]naloxone by the rat brain increased as the pH of the injected solution was increased from 5.5 to 8.5 and was strongly inhibited by cationic H(1)-antagonists such as pyrilamine and diphenhydramine and cationic drugs such as lidocaine and propranolol. In contrast, the BBB transport of [(3)H]naloxone was not affected by any typical substrates for organic cation transport systems such as tetraethylammonium, ergothioneine or L-carnitine or substrates for organic anion transport systems such as p-aminohippuric acid, benzylpenicillin or pravastatin. The present results suggest that a pH-dependent and saturable influx transport system that is a selective transporter for cationic H(1)-antagonists is involved in the BBB transport of naloxone in the rat.

Glyburide (GLB) is a widely used oral sulfonylurea for the treatment of gestational diabetes. The therapeutic use of GLB is often complicated by a substantial inter-individual variability in the pharmacokinetics and pharmacodynamics of the drug in human populations, which might be caused by inter-individual variations in factors such as GLB metabolism. Therefore, there has been a continued interest in identifying human cytochrome P450 (CYP) isoforms that play a major role in the metabolism of GLB. However, contrasting data are available in the present literature in this regard. The present study systematically investigated the contributions of various human CYP isoforms (CYP3A4, CYP3A5, CYP2C8, CYP2C9 and CYP2C19) to in vitro metabolism of GLB. GLB depletion and metabolite formation in human liver microsomes were most significantly inhibited by the CYP3A inhibitor ketoconazole compared with the inhibitors of other CYP isoforms. Furthermore, multiple correlation analysis between GLB depletion and individual CYP activities was performed, demonstrating a significant correlation between GLB depletion and the CYP3A probe activity in 16 individual human liver microsomal preparations, but not between GLB depletion and the CYP2C19, CYP2C8 or CYP2C9 probe activity. By using recombinant supersomes overexpressing individual human CYP isoforms, it was found that GLB could be depleted by all the enzymes tested; however, the intrinsic clearance (V(max)/K(m)) of CYP3A4 for GLB depletion was 4-17 times greater than that of other CYP isoforms. These results confirm that human CYP3A4 is the major enzyme involved in the in vitro metabolism of GLB.

Interactions are expected between transporters and enzymes that compete for the substrate within the cell, and controversy exists for data interpretation on the interplay between transporters and enzymes. In the Caco-2 cell monolayer, the increase in mean residence time (MRT) of drug accompanying increased secretion has been construed as the reason for increased metabolism, whereas others hold an opposite view that increased secretion would evoke decreased metabolism in this closed system. A catenary Caco-2 cell model was used to simulate the effects of altered secretion on metabolism, estimated as the fraction of dose metabolized (f(met)) or the extraction ratio (ER). The simulations showed that both f(met) and ER varied inversely with the transporter-mediated intrinsic clearance for apical secretion (CL(int,sec)) under linear conditions. Under non-linear conditions of saturable metabolism, apical absorption, basolateral influx or efflux, the simulated f(met) consistently bore a reciprocal relationship with CL(int,sec). For saturable apical absorption or basolateral efflux, a reciprocal relationship between the ER and CL(int,sec) was also found. However, under conditions of saturable metabolism or basolateral influx, the pattern of change in the ER became dependent on the administration sites, showing increasing patterns for apical dosing but decreasing patterns for basolateral dosing with increasing values of CL(int,sec). In general, f(met) consistently demonstrated an inverse relationship with CL(int,sec), whereas ER, failing to include the drug in the donor side, would not show the same pattern of change in metabolism especially for apical dosing, since a substantial amount of drug was back-secreted into the apical compartment.

Hepatotoxicity has been reported as a side-effect in some patients on HIV protease inhibitors (PI). Since transporter interaction has been implicated as a mechanism underlying drug-mediated hepatotoxicity and drug-drug interactions, the interaction of PI with the hepatic canalicular efflux transporter ABCC2 (MRP2; multidrug resistance associated protein-2) was studied. Interaction with ABCC2/Abcc2 was evaluated in human and rat sandwich-cultured hepatocytes using 5(6)-carboxy-2',7'-dichlorofluorescein (CDF) as substrate. In rat hepatocytes, interaction with estradiol-17-beta-D-glucuronide (E17G) efflux was also studied. In human hepatocytes, saquinavir, ritonavir and atazanavir were the most efficient inhibitors of ABCC2-mediated biliary excretion of CDF, whereas in rat hepatocytes indinavir, lopinavir and nelfinavir were the most efficient. No species-similarity was found for ABCC2/Abcc2 inhibition. In rat hepatocytes, the effects on Abcc2 were substrate-dependent as inhibition of biliary excretion of E17G was most pronounced for saquinavir (completely blocked), amprenavir (82% inhibition) and indinavir (68% inhibition). In conclusion, several HIV PI showed substantial ABCC2 inhibition, which, combined with the effects of PI on other hepatobiliary disposition mechanisms, will determine the clinical relevance of these in vitro interaction data.