Population pharmacokinetics of sirolimus in kidney transplant patients.

OBJECTIVE: To characterize the dose-related pharmacokinetics of the immunosuppressant agent sirolimus (formerly rapamycin) in kidney transplant patients by use of two-stage and nonlinear mixed-effect model population methods. METHODS: Patients (n = 36) from three centers (Germany, the United Kingdom, and Sweden) who received steady-state oral doses of cyclosporine (ciclosporin) were assessed after single oral administration of sirolimus at doses of 3, 5, 10, and 15 mg/m2. Plasma and whole blood sirolimus samples were analyzed by a high-performance liquid chromatographic/mass spectrophotometric method. Simultaneous fitting used biexponential functions with intercept/slope or clearance/volume terms, as well as first-order absorption (ka) and a lag-time. RESULTS: The nonlinear mixed-effect model method (P-Pharm) provided a better characterization of sirolimus kinetics, especially for the absorption and distribution phases where fewer data were available per patient. Sirolimus distribution between whole blood and plasma was concentration-independent, with a mean blood/plasma ratio (coefficient of variation) of 30.9 (48.5%). Elimination was not influenced by dose, as shown by estimates of the terminal half-life of 63 hours (27.5%) and apparent oral blood clearance of 8.9 L/hr (38.2%). Sirolimus distribution parameters were influenced by body weight and surface area. Sirolimus was rapidly absorbed, as shown by the absorption lag-time of 0.27 hour (35.1%), and ka of 2.77 hr-1 (48.4%). The concomitant administration of sirolimus and cyclosporine did not reveal any pharmacokinetic interactions. CONCLUSION: This report provides an initial population pharmacokinetics of sirolimus in kidney transplant recipients receiving cyclosporine concurrently. Sirolimus blood and plasma pharmacokinetics were biexponential and linear for doses from 3 to 15 mg/m2. No pharmacokinetic interaction was found between sirolimus and cyclosporine.

Population distribution and effects on drug metabolism of a genetic variant in the 5' promoter region of CYP3A4.

BACKGROUND: There are large interindividual differences in CYP3A4 expression and in the metabolism of drug substrates for this enzyme. We and others have identified a polymorphism in the 5' promotor region of the CYP3A4 gene; however, its functional significance is not currently known. This study was conducted to determine whether this polymorphism plays a clinically important role in determining CYP3A4 phenotype. METHODS: An adenine (A) to guanine (G) transition was identified in the 5' promotor region of the CYP3A4 gene at position -292 (from the start codon), in a sequence motif known as the nifedipine-specific element. The frequency of this polymorphism was assessed in 802 healthy volunteers from five broadly defined racial groups. The population distribution of the G allele in these groups was as follows: white Americans (3.6%; n = 273), black Americans (54.6%, n = 186), Hispanic Americans (9.3%; n = 188), Japanese Americans (0.0%; n = 77), and Chinese Americans (0.0%; n = 78). In a subsequent study, 90 additional black Americans were genotyped, and a subset of the homozygous subjects (AA, n = 8; GG, n = 23) were given the CYP3A4 probe substrates erythromycin and nifedipine to allow genotype-phenotype comparisons to be made. RESULTS: There was no difference in the rate of CYP3A4-dependent demethylation of erythromycin (erythromycin breath test) or the pharmacokinetics of nifedipine or its CYP3A4-dependent metabolite dehydronifedipine between the two genotype groups (AA or GG). CONCLUSIONS: This promotor region polymorphism does not appear to play a major role in determining constitutive CYP3A4 expression.

Gender-related assessment of cyclosporine/prednisolone/sirolimus interactions in three human lymphocyte proliferation assays.

BACKGROUND: Cyclosporine (CsA), prednisolone (Pred), and sirolimus (Sir) are immunosuppressive compounds inhibiting lymphocyte proliferation at the cytokine gene transcription (CsA and Pred) or signal transduction (Sir) levels. METHODS: Double- and triple-drug interactions were simultaneously studied using lectin-induced proliferation of isolated cell lymphocytes (ICLP) and whole blood lymphocytes from men and women as well as two-way mixed lymphocyte reaction assays. Drug interactions were described with isobolograms and quantitated with the universal response surface approach by estimating the interaction parameter alpha. RESULTS: All compounds inhibited more than 89% of control proliferative responses. In each assay, CsA was less potent than Pred (3- to 14-fold) and Sir (5- to 11-fold). Sir was of similar or higher potency than Pred and 1.5-fold more potent in men than women. Pred was 1.4 times more potent in women but this was found only in the ICLP assay. All combinations were synergistic (alpha>0), with greater synergism found for combinations involving Sir, especially in the ICLP (alpha>13) and two-way mixed lymphocyte reaction (alpha>40) assays. Moreover, the Sir/Pred interaction in the ICLP assay was two to five times more synergistic in women, because their mean alpha was 56 compared with 13 in men. Double-combination alpha values were able to reasonably describe CsA/Pred/Sir triple-interaction effects. CONCLUSIONS: These studies indicate that CsA, Pred, and Sir act and synergistically interact in vitro, with gender and assay as additional factors, and that whole blood lymphocyte proliferation cultures are useful in assessing the nature and intensity of drug interactions.

Pharmacodynamic modeling of pantoprazole's irreversible effect on gastric acid secretion in humans and rats.

The relationship between the pharmacokinetics of pantoprazole, an irreversible proton pump inhibitor, and its effect on gastric acid secretion was evaluated in humans and rats. Pantoprazole pharmacokinetics were studied in 6 rats (5 mg/kg, i.v.) and 22 healthy volunteers (10 to 80 mg, i.v. and oral). Gastric acid secretion under maximum pentagastrin stimulation was measured after i.v. administration of placebo or pantoprazole in 31 rats (0.12 to 1.15 mg/kg) for 4 hours and in 31 subjects (20 to 120 mg) for 24 hours. Pantoprazole has short half-lives of 0.5 hours in rats and 0.8 hours in humans. After administration of the highest dose, acid secretion was fully inhibited within 1 hour and for the whole observation period in both species. An irreversible pharmacodynamic response model was successfully developed and validated. The apparent reaction rate constants of pantoprazole with the proton pumps were 0.691 L/mg/h in rats and 0.751 L/mg/h in humans, and the apparent recovery rates of the pumps were 0.053 h-1 and 0.031 h-1, respectively. The maximum inhibition and the overall effect of pantoprazole are related to exposure, and the onset is related to initial pantoprazole concentrations. It was concluded that this irreversible response model accurately describes the effect of i.v. and oral pantoprazole on gastric secretion and may be used to predict effects under other dosage regimens.

The effect of a high-fat meal on the oral bioavailability of the immunosuppressant sirolimus (rapamycin).

The bioavailability of an oral nonaqueous solution of sirolimus was compared under fasting conditions and after a high-fat meal in a randomized, two-way crossover pharmacokinetic study. Healthy volunteers were administered a 15 mg single dose of sirolimus on two occasions, once while fasting and once after consumption of a high-fat breakfast. Whole blood concentrations of sirolimus were assayed by using a validated method with high-performance liquid chromatography/tandem mass spectrometric detection. Sirolimus was absorbed more slowly when administered after a high-fat meal than when administered after fasting, as shown by statistically significant reductions in peak concentration (Cmax) and the ratio of Cmax to the area under the curve (AUC), and lengthening of the time to peak concentration. The oral availability of sirolimus was increased to a modest extent (35%) and in a uniform manner when administered with a high-fat meal; the geometric mean ratio of the fed/fasting AUC values was 1.35, with a 90% confidence interval of 1.26 to 1.46. Food had no effect on the terminal half-life of sirolimus (mean values of 67 to 68 hours). The 35% increase in AUC obtained after a high-fat meal appears small relative to the intersubject and intrasubject variabilities observed in clinical trials. However, to minimize unnecessary fluctuations in trough whole blood sirolimus concentrations, it is advisable that sirolimus be administered consistently in individual patients, either with or without meals.

Oral absorption characteristics and pharmacokinetics of colchicine in healthy volunteers after single and multiple doses.

Colchicine is an antimitotic drug used to treat gout and familial Mediterranean fever. Absolute bioavailability, pharmacokinetics, and absorption characteristics of colchicine after single 1.0-mg doses in oral solution or tablet form or 0.5-mg intravenous doses were compared in 6 subjects. This study was combined with 14 days of multiple-dose administration of 1.0-mg colchicine tablets in 6 subjects. Serial blood samples were collected for 48 hours after administration of single doses and for 120 hours after the last dose in the multiple-dose regimen. Plasma colchicine profiles as measured by radioimmunoassay were analyzed using deconvolution and compartmental methods. After intravenous bolus injection of colchicine, the area under the concentration-time curve (AUC) was 61.2 +/- 12.7 ng.hr/mL, steady-state volume of distribution (Vss) was 419 +/- 95 L, systemic clearance (Cl) was 8.5 +/- 1.8 L/hr, and the terminal half-life (t1/2) was 57.8 +/- 10.7 hours. After oral administration in solution form, peak plasma concentrations (Cmax) of 6.50 +/- 1.03 ng/mL were reached at time (tmax) 1.07 +/- 0.55 hours, with a rate of 0.109 +/- 0.024 hr-1 (Cmax/AUC); bioavailability was 47 +/- 14%. Oral tablets yielded similar Cmax, tmax, and Cmax/AUC values, but AUC was significantly lower. Most participants exhibited a secondary peak within 6 hours of administration, possibly in relation to a second absorption site or enterohepatic recirculation. This second absorption process was significantly longer than the first one, and accounted for a similar amount of colchicine absorbed. From the multiple-dose study, a model including an alteration of colchicine absorption due to possible drug-induced gastrointestinal modifications allowed better determination of steady-state plasma concentrations of colchicine.

Pharmacokinetics of prednisolone during administration of sirolimus in patients with renal transplants.

The pharmacokinetic interaction of multiple oral doses of sirolimus (rapamycin) and prednisone were evaluated in 40 stable patients with renal transplants receiving concomitant multiple doses of cyclosporine. Nine sirolimus dosage levels from 1 mg/m2/day to 13 mg/m2/day were studied and compared with placebo. Plasma concentrations of prednisone, prednisolone, and cortisol were measured by high-performance liquid chromatography and analyzed by noncompartmental methods. Mean pharmacokinetic values of prednisolone found before sirolimus administration were as follows: peak plasma concentration (Cmax) was 187 ng/mL; time to peak plasma concentration (tmax) was 2.03 hours; rate of reaching peak plasma concentration (Cmax divided by the area under the concentration-time curve [AUC]) was 0.149 hour-1; terminal half-life (t1/2) was 3.60 hours; AUC was 1206 ng.hour/mL; and apparent clearance (Cl/F) was 0.094 L/hour/kg. During the 2 weeks of concomitant administration, prednisolone elimination decreased in relation to sirolimus dosages. These changes were modest, with mean increases of 18% in Cmax and 27% in t1/2 and mean decreases of 27% in Cl/F for the groups receiving 6 mg/m2/day to 13 mg/m2/day. Most patients initially had plasma cortisol concentrations indicative of adrenal suppression. With sirolimus treatment, the Cmax of cortisol did not decrease further, but the AUC (8:00 AM-8:00 PM) values were significantly lower, independent of sirolimus exposure. The AUC for cyclosporine did not correlate with sirolimus and prednisolone exposure. A 2-week course of sirolimus showed a slight pharmacokinetic interaction between sirolimus and prednisolone/prednisone/cortisol in stable patients with renal transplants.

Lack of pharmacokinetic interaction between oral pantoprazole and cisapride in healthy adults.

Pantoprazole, an irreversible proton pump inhibitor, may be administered with cisapride, a prokinetic agent. As increased cisapride concentrations may result in longer electrocardiogram (ECG) QTc intervals, a crossover study was conducted in healthy subjects to evaluate the oral pharmacokinetic interaction between cisapride (20 mg) and pantoprazole (40 mg). After dosing, serial blood samples and 12-lead ECGs were collected, and cisapride plasma concentrations were quantitated. For cisapride alone, mean parameter values were the following: peak concentration (Cmax), 56 ng/mL; time to Cmax (tmax), 1.7 hours; area under the concentration-time curve (AUC), 426 ng x h/mL; and terminal half-life (t1/2), 5.8 hours. Pantoprazole coadministration did not alter cisapride AUC or other pharmacokinetic parameters except for a slight 17% decrease in Cmax' resulting in 90% confidence limits of 79% to 88%, which were marginally outside strict bioequivalence limits. In addition, cisapride did not affect ECG QTc intervals, with or without pantoprazole. Therefore, no dosage adjustment is needed when pantoprazole and cisapride are coadministered.

Pharmacokinetics of pantoprazole in patients with moderate and severe hepatic dysfunction.

BACKGROUND: Patients with impaired hepatic function usually require gastric acid-suppressant therapy but are at increased risk for drug interactions and may require dosage adjustments. The proton pump inhibitor pantoprazole is rapidly absorbed and eliminated, primarily by cytochrome P450 (CYP) 2C19 isozymes. OBJECTIVE: This study sought to determine whether dosage adjustment of pantoprazole is required in patients with moderate or severe hepatic impairment by comparing the pharmacokinetic profile of pantoprazole in such patients with that in healthy slow metabolizers of pantoprazole, in whom no dosage adjustment is required. METHODS: Patients with moderate (Child-Pugh class B) and severe (Child-Pugh class C) hepatic impairment received oral pantoprazole 40 mg once daily on days 1 through 4 and then on alternate days (days 6 and 8). Serial blood samples were collected on days 4 and 8 for analyses of plasma pantoprazole concentrations. Pharmacokinetic data were compared between the 2 groups with hepatic impairment and against historical data from 17 healthy subjects who were genetically slow CYP2C19 metabolizers of pantoprazole. RESULTS: Twenty-two patients participated in the study, 13 in the Child-Pugh class B group and 9 in the Child-Pugh class C group. No clinically significant differences in pantoprazole pharmacokinetics were noted between the patients with hepatic impairment and the healthy slow metabolizers of pantoprazole on days 4 and 8. Pantoprazole was well tolerated. Four Child-Pugh class B patients and 3 Child-Pugh class C patients reported > or = 1 adverse event. Adverse events were generally mild or moderate, and were similar to those reported in healthy subjects. Two patients discontinued the study because of severe events related to their underlying disease. CONCLUSIONS: The pharmacokinetics and tolerability of pantoprazole were similar in patients with moderate hepatic impairment, patients with severe hepatic impairment, and healthy slow metabolizers of pantoprazole, in whom no dosage adjustment is required. Thus, no dosage adjustment of pantoprazole is required in patients with hepatic impairment, regardless of its severity. However, caution should be exercised when giving pantoprazole to patients with severe hepatic impairment.

Species- and gender-related differences in cyclosporine/prednisolone/sirolimus interactions in whole blood lymphocyte proliferation assays.

Cyclosporin A (CyA), prednisolone (Pred) and sirolimus (Sir) inhibit lymphocyte proliferation at the cytokine transcription (CyA and Pred) or signal transduction (Sir) levels. Their double and triple interactions were studied on lectin-induced proliferation of whole blood lymphocytes (WBLP) from male and female humans, rabbits and rats. Isobols along with the Universal Response Surface Approach were used to describe and quantify the nature and intensity of drug interactions by determining alpha values. CyA was always less potent than Pred and Sir while these two compounds were relatively equipotent. Species-related differences were observed with single drugs. Rabbit WBLP were resistant to Pred action (Imax = 67%) and rats were more sensitive to Pred (IC50 = 9.1 nM in females) and Sir (2.8 nM) actions than humans (32 and 55 nM). Gender differences were observed but were not consistent across species. All double-drug combinations were synergistic, and combinations containing Pred were 10 to 100 times more synergistic in rabbits (alpha Pred/Sir = 213 and alpha CyA/Pred = 147 in males) than in rats (12 and 2.1) or humans (3.7 and 5.7) in relation to the lower efficacy of Pred. Double-combination alpha values were able to describe CyA/Pred/Sir triple combination effects. These studies indicate that CyA, Pred and Sir act and synergistically interact in vitro in species- and gender-dependent fashions. Adrenalectomized rats better resemble humans in these responses. WBLP are useful in various species in determining immunosuppressive drug action and interactions.

Species differences in sirolimus stability in humans, rabbits, and rats.

Species differences in the in vitro stability of sirolimus was assessed in plasma and whole blood in relation to red blood cell distribution. Fresh blood and plasma samples obtained from humans, rabbits, and rats were aliquoted and spiked with sirolimus. After incubation from 0 to 144 hr in a shaking water bath maintained at 37 degrees C, sirolimus concentrations were quantified by a specific high-liquid performance chromatographic method. Sirolimus was unstable in both plasma and whole blood. Sirolimus degradation half-life in whole blood was 135 hr (vs. 7.2 hr in plasma) in humans, 62 hr (vs. 3.1 hr) in rabbits, and 15 hr (vs. 2.2 hr) in rats. Sirolimus stability is greater in whole blood compared with plasma in proportion to the whole blood/plasma ratio and hematocrit. In vivo instability may account for up to 36% of sirolimus clearance in humans and 13% in rabbits, and this is an important factor in the pharmacokinetics of this drug.

Pharmacokinetic and pharmacoimmunodynamic interactions between prednisolone and sirolimus in rabbits.

PURPOSE: To assess pharmacokinetic and pharmacoimmunodynamic interactions between prednisolone (Pred, I1 mg/kg) and sirolimus (Sir, 0.25 mg/kg) in rabbits. METHODS: After intravenous administration, plasma concentrations of Pred and corticosterone, and Sir blood concentrations were followed for 24 hours along with blood granulocyte and T-helper cell counts. Ex vivo and in vitro whole blood lymphocyte proliferation marked lymphocyte reactivity. RESULTS: Pred terminal half-life and clearance were 1.1 hr and 0.72 l/hr/kg with no difference after Sir. Sir values were 13 hr and 0.16 l/hr/kg and Pred produced no changes. Corticosterone production (0-12hr) was suppressed by 55% after Pred alone or combined, while Sir did not cause adrenal suppression. Blood T-helper cells and granulocytes displayed circadian rhythms after placebo. Over 12 hr, T-helper cell counts were decreased by Pred (40%) and Sir (19%) while granulocyte numbers increased by 56% and 23%. After coadministration, cell numbers were similar to Pred alone. Pred and Sir decreased lymphocyte reactivity by 41% and 56% over 24 hr and their combination reached 85% inhibition with additive interaction. In vitro studies showed antagonistic or synergistic interactions depending on drug concentration ratios. CONCLUSIONS: At therapeutic concentrations, Sir and Pred do not significantly interact pharmacokinetically and have additive pharmacoimmunodynamics. Thus, the therapeutic application of this combination is promising.

Lipophilic benzamide and anilide derivatives as high-performance liquid chromatography internal standards: application to sirolimus (rapamycin) determination.

Finding a suitable internal standard in reversed-phase high-performance liquid chromatography is often difficult. A general approach for selecting and synthesizing the proper internal standard is presented and applied to a validated method for quantitation of sirolimus in several biological matrices. A series of fifteen N-alkylbenzamides, N-alkyltoluamides and N-alkanoylanilines with a log P range of 3.51 to 6.68 were synthesized as internal standards; N-undecyl-o-toluamide was evaluated most extensively. Sirolimus quantitation involves a simple sample clean-up procedure followed by isocratic chromatography on a heated C18 analytical column with an 70% methanol-water mobile phase and ultraviolet detection at 278 nm. This method was linear from 2.5 to 200 ng with a limit of quantitation of 2.5 ng using a 1-ml blood sample. Sirolimus recovery was above 72.1%. The intra-day and inter-day coefficients of variation were less than 11.7%. Several drugs and sirolimus metabolites do not interfere with the analysis. This method was used to measure sirolimus in blood from rats, rabbits and humans.

Pharmacokinetic and pharmacoimmunodynamic interactions between prednisolone and sirolimus in adrenalectomized rats.

Prednisolone (Pred) and sirolimus (SIR) are immunosuppressive compounds acting through different mechanisms with moderate synergism found in vitro. Both drugs are metabolized partly by CYP3A enzymes. After i.v. administration of placebo, Pred (5 mg/kg), SIR (1 mg/kg), or Pred with SIR (5 and 1 mg/kg doses) to adrenalectomized male rats, Pred plasma and SIR whole blood concentrations were followed for 48 hr along with circulating T-helper and T-cytotoxic cell counts. Ex vivo whole blood lymphocyte proliferation marked host responsiveness. An extended indirect PK/PD model was used to describe responses to these drugs, alone or combined. An interactive two-stage population analysis showed no modification in drug PK. Mean Pred plasma clearance was 0.655 L/hr (interrat++ variability: 11%) and significantly increased with weight. Mean SIR whole blood volume of distribution and clearance were 5.6 L (62%) and 0.28 L/hr (32%), and animal scaling showed weight-power proportionality. In vitro metabolism studies showed no significant inhibition of Pred or prednisone CYP3A metabolism by SIR (50 microM), but this pathway accounted for less than 5% of Pred metabolism. Pred decreased numbers of T-helper lymphocytes with a mean IC50 of 37.8 nM (21%) alone or 12.3 nM (130%) with SIR. Results for T-cytotoxic lymphocytes were similar. SIR increased lymphocyte numbers with a mean IC50 of 52.2 nM (24%) for T-helper and 28.8 nM (51%) for T-cytotoxic cells. Taking into account drug effects on lymphocyte trafficking, Pred directly inhibited ex vivo lymphocyte proliferation with a mean IC50 of 1.08 nM (38%). SIR, after a transduction step, inhibited proliferation with a mean IC50 of 1.00 nM (26%). Responses measured after drug coadministration were reasonably quantitated by addition of single drug effects. Since, at pharmacologic concentrations in rats, Pred and SIR did not interact in their PK but synergistically or additively interact in their dynamics, their joint therapeutic use is promising. The adrenalectomized rat may be a suitable animal model to characterize drug effects on lymphocyte trafficking and reactivity.