The application of cassette dosing for pharmacokinetic screening in small-molecule cancer drug discovery

Pharmacokinetic evaluation is an essential component of drug discovery and should be conducted early in the process so that those compounds with the best chance of success are prioritized and progressed. However, pharmacokinetic analysis has become a serious bottleneck during the 'hit-to-lead' and lead optimization phases due to the availability of new targets and the large numbers of compounds resulting from advances in synthesis and screening technologies. Cassette dosing, which involves the simultaneous administration of several compounds to a single animal followed by rapid sample analysis by liquid chromatography/tandem mass spectrometry, was developed to increase the throughput of in vivo pharmacokinetic screening. Although cassette dosing is advantageous in terms of resources and throughput, there are possible complications associated with this approach, such as the potential for compound interactions. Following an overview of the cassette dosing literature, this article focuses on the application of the technique in anticancer drug discovery. Specific examples are discussed, including the evaluation of cassette dosing to assess pharmacokinetic properties in the development of cyclin-dependent kinase and heat shock protein 90 inhibitors. Subject to critical analysis and validation in each case, the use of cassette dosing is recommended in appropriate chemical series to enhance the efficiency of drug discovery and reduce animal usage.     

Preclinical pharmacokinetics and metabolism of a novel diaryl pyrazole resorcinol series of heat shock protein 90 inhibitors

CCT018159 was recently identified as a novel inhibitor of heat shock protein (Hsp) 90, a promising target for cancer therapy. Pharmacokinetic and metabolic properties are likely to be important for efficacy and need to be optimized during drug development. Here, we define the preclinical metabolism and pharmacokinetics of CCT018159 and some early derivatives. In addition, we assess in vitro metabolic stability screening and in vivo cassette dosing (simultaneous administration of several compounds to a single animal) as approaches to investigate these compounds. The plasma clearance following individual i.v. administration to mice was rapid (0.128-0.816 L/h), exceeding hepatic blood flow. For CCT066950 and CCT066952, this could be attributed in part to extensive (>80%) blood cell binding. Oral bioavailability ranged from 1.8% to 29.6%. Tissue distribution of CCT066952 was rapid and moderate, and renal excretion of the compounds was minimal (<1% of dose excreted). Compounds underwent rapid glucuronidation both in vivo and following incubation with mouse liver microsomes. However, whereas CCT066965 was metabolized to the greatest extent in vitro, this compound displayed the slowest plasma clearance. The rank order of the compounds from the highest to lowest area under the curve was the same following discrete and cassette dosing. Furthermore, pharmacokinetic variables were similar whether the compounds were dosed alone or in combination. We conclude that the pharmacokinetics of CCT018159 are complex. Cassette dosing is currently the best option available to assess the pharmacokinetics of this promising series of compounds in relatively high throughput and is now being applied to identify compounds with optimal pharmacokinetic properties during structural analogue synthesis.     

Effect of Maalox on the bioavailability of oral gemifloxacin in healthy volunteers

This open, randomized, 4-way crossover study investigated the effect of the antacid Maalox on the bioavailability of gemifloxacin, a novel fluoroquinolone antimicrobial. Sixteen healthy male volunteers received gemifloxacin, 320 mg p.o., alone, 3 h after Maalox administration, or 10 min or 2 h before Maalox administration. Blood was sampled for 48 h after dosing to determine pharmacokinetic parameters. Estimates for the differences between regimens and 95% confidence intervals were calculated using the t-test for paired data. The administration of gemifloxacin 10 min before Maalox resulted in an average 85% reduction in the area under the plasma concentration-time curve from time zero extrapolated to infinity (AUC(0-infinity)), whereas administration 3 h after Maalox produced a decrease in AUC(0-infinity), 15% of which was not considered to be clinically significant. The administration of gemifloxacin 2 h before Maalox had no notable effect on the gemifloxacin AUC(0-infinity) (average increase of 3%). Similar results were seen for the maximum gemifloxacin plasma concentration (C(max)). Neither the time to C(max) nor the half-life of gemifloxacin were notably altered by the administration of Maalox at any time relative to gemifloxacin dosing. There were no clinically important adverse experiences or changes in clinical laboratory parameters during this study. The findings of this study support the dosing recommendation that gemifloxacin can be administered either 2 h or more prior to, or 3 h or more after, the administration of Maalox. Copyright Copyright 1999 S. Karger AG, Basel.     

Compartmental pharmacokinetics and tissue distribution of multilamellar liposomal nystatin in rabbits

The plasma pharmacokinetics of multilamellar liposomal nystatin were studied in normal, catheterized rabbits after single and multiple daily intravenous administration of dosages of 2, 4, and 6 mg/kg of body weight, and drug levels in tissues were assessed after multiple dosing. Concentrations of liposomal nystatin were measured as those of nystatin by a validated high-performance liquid chromatography method, and plasma concentration data were fitted into a two-compartment open model. Across the investigated dosage range, liposomal nystatin demonstrated nonlinear kinetics with more than proportional increases in the AUC(0-24) and decreasing clearance, consistent with dose-dependent tissue distribution and/or a dose-dependent elimination process. After single-dose administration, the mean C(max) increased from 13.07 microg/ml at 2 mg/kg to 41.91 microg/ml at 6 mg/kg (P < 0.001); the AUC(0-24) changed from 11.65 to 67.44 microg. h/ml (P < 0.001), the V(d) changed from 0.205 to 0. 184 liters/kg (not significant), the CL(t) from 0.173 to 0.101 liters/kg. h (P < 0.05), and terminal half-life from 0.96 to 1.51 h (P < 0.05). There were no significant changes in pharmacokinetic parameters after multiple dosing over 14 days. Assessment of tissue concentrations of nystatin near peak plasma levels after multiple dosing over 15 days revealed preferential distribution to the lungs, liver, and spleen at that time point. Substantial levels were also found in the urine, raising the possibility that renal excretion may play a significant role in drug elimination. Liposomal nystatin administered to rabbits was well tolerated and displayed nonlinear pharmacokinetics, potentially therapeutic peak plasma concentrations, and substantial penetration into tissues. Pharmacokinetic parameters were very similar to those observed in patients, thus validating results derived from infection models in the rabbit and allowing inferences to be made about the treatment of invasive fungal infections in humans.     

Pharmacokinetics and pharmacodynamics of TMC207 and its N-desmethyl metabolite in a murine model of tuberculosis

TMC207 is a first-in-class diarylquinoline with a new mode of action against mycobacteria targeting the ATP synthase. It is metabolized to an active derivative, N-desmethyl TMC207, and both compounds are eliminated with long terminal half-lives (50 to 60 h in mice) reflecting slow release from tissues such as lung and spleen. In vitro, TMC207 is 5-fold more potent against Mycobacterium tuberculosis than N-desmethyl TMC207, and the effects of the two compounds are additive. The pharmacokinetic and pharmacodynamic (PK-PD) response was investigated in the murine model of tuberculosis (TB) infection following oral administration of different doses of TMC207 or N-desmethyl TMC207 at 5 days per week for 4 weeks starting the day after intravenous infection with M. tuberculosis and following administration of different doses of TMC207 at various dosing frequencies for 6 weeks starting 2 weeks after infection. Upon administration of N-desmethyl TMC207, maximum plasma concentration (C(max)), area under the plasma concentration-time curve from time zero to 168 h postdose (AUC(168h)), and minimum plasma concentration (C(min)) were approximately dose proportional between 8 and 64 mg/kg, and the lung CFU counts were strongly correlated with these pharmacokinetic parameters using an inhibitory sigmoid maximum effect (E(max)) model. Administration of the highest dose (64 mg/kg) produced a 4.0-log(10) reduction of the bacillary load at an average exposure (average concentration [C(avg)] or AUC(168h) divided by 168) of 2.7 μg/ml. Upon administration of the highest dose of TMC207 (50 mg/kg) 5 days per week for 4 weeks, the total reduction of the bacillary load was 4.7 log(10). TMC207 was estimated to contribute to a 1.8-log(10) reduction and its corresponding exposure (C(avg)) was 0.5 μg/ml. Optimal bactericidal activity with N-desmethyl TMC207 was reached at a high exposure compared to that achieved in humans, suggesting a minor contribution of the metabolite to the overall bactericidal activity in TB-infected patients treated with TMC207. Following administration of TMC207 at a total weekly dose of 15, 30, or 60 mg/kg fractionated for either 5 days per week, twice weekly, or once weekly, the bactericidal activity was correlated to the total weekly dose and was not influenced by the frequency of administration. Exposures (AUC(168h)) to TMC207 and N-desmethyl TMC207 mirrored this dose response, indicating that the bactericidal activity of TMC207 is concentration dependent and that AUC is the main PK-PD driver on which dose optimization should be based for dosing frequencies up to once weekly. The PK-PD profile supports intermittent administration of TMC207, in agreement with its slow release from tissues.     

Bioavailability of two manufacturers' sustained-release quinidine gluconate tablets at steady state

Steady-state bioavailability of sustained-release quinidine gluconate tablets manufactured by two companies was compared in a crossover study. The tablets were Quinaglute Dura-Tabs, manufactured by Berlex Laboratories, Inc., and generic quinidine gluconate tablets, manufactured by Bolar Pharmaceutical Company. Sixteen healthy male volunteers were given multiple doses of the two products in randomized sequence. Blood samples were obtained immediately before administration of the seventh dose (hour 72) and at 1, 2, 3, 4, 5, 6, 8, 10, and 12 hours after administration. Plasma samples were assayed for quinidine content by high-performance liquid chromatography. The tablets manufactured by Berlex provided statistically significantly higher plasma levels during the second half of the dosing interval (six to 12 hours postdose). A 29% difference in plasma levels was observed between the products at the end of the dosing interval. The Bolar quinidine gluconate tablets had a statistically significant lower area under the curve (AUC). The greatest difference in AUC occurred during the last six hours of the dosing period. The six- to 12-hour AUC for the Bolar tablets was 24% less than that for Berlex tablets. The generic tablets also achieved peak plasma level 31% sooner than did Quinaglute Dura-Tabs. The pharmacokinetic characteristics of the two products at steady state indicate that the Bolar quinidine gluconate tablet exhibited a more rapid onset of peak plasma levels and a more rapid decline to minimum plasma levels. In summary, the data from this multiple-dose study, performed using commercially available material, indicate that differences exist in pharmacokinetic performance of the products. However, the exact correlation between pharmacokinetic data and clinical effectiveness has not been established.     

The importance of software for the planning and adjusting of dosage. Application with aminoglycosides

We are presenting a Mac Intosh--Apple computer program (Excel*) for the aminoglycosides (AG) drug monitoring useful in the following situations: --initiation of an AG's dosing regimen depending on the desired peak and through concentration (AG's pharmacokinetic parameters have been calculated according to the AG's literature values). --an adequate dosing regimen after revision of the patient's parameters (calculated with the observed concentrations). The technique has been validated comparing desired (des. C) and measured concentrations (mes. C). --either at the beginning of the treatment, --or after revision of the individual parameters. In the first case, the correlation coefficient obtained between 14 des C and 14 mes. C is equal to 0.91 and is improved when individual parameters are calculated (0.94).     

Phase I study of multiple-dose cefprozil and comparison with cefaclor.

The objectives of this study were to assess the safety and tolerance of cefprozil, to characterize the pharmacokinetics of cefprozil after administration of multiple doses of the drug, and to compare these pharmacokinetic parameters with those obtained with cefaclor. The volunteers received 28 doses of 250, 500, or 1,000 mg of cefprozil or 500 mg of cefaclor every 8 h for 10 days. Serial blood samples and the total volume of urine voided by each individual were collected for pharmacokinetic evaluation on days 1, 5, and 10. Both cephalosporins were well tolerated after multiple oral dosing. The peak levels in plasma (Cmax) of cefprozil ranged from 5.7 to 18.3 micrograms/ml after oral administration of 250- to 1,000-mg doses. The regression analysis of Cmax on cefprozil dose showed a dose-linear response. The mean Cmax of cefaclor ranged from 15.2 to 16.7 micrograms/ml and did not change significantly on multiple dosing. The overall mean terminal half-life of cefprozil was 1.2 h and was invariant with respect to dose or duration of dosing. The area under the plasma-concentration-versus-time curve from 0 h to infinity (AUC0-infinity) of cefprozil increased in a dose-proportional manner with an increase in dose. The overall urinary recovery (61% of dose) and renal clearance values of cefprozil were generally invariant with respect to dose and duration of dosing. While cefprozil was apparently absorbed less rapidly and achieved lower Cmax values than cefaclor, the AUC0-infinity of cefprozil was nearly twofold greater than that of cefaclor. The half-life of cefprozil was also twofold longer than that observed for cefaclor. Although the urinary recovery of cefaclor (75% of dose) was significantly higher than that of cefprozil (61% of dose), the concentrations of cefprozil in urine remained significantly higher than those of cefaclor from 2 to 8 h postdosing. If the therapeutic concept is maintained that levels of beta-lactam antibiotics in plasma should exceed the MIC for the offending organisms over a period that approximates the dosing interval, then cefprozil would appear to be suitable for twice-daily administration, whereas cefaclor should probably be administered three or even four times a day.     

Modest but variable effect of rifampin on steady-state plasma pharmacokinetics of efavirenz in healthy African-American and Caucasian volunteers

Efavirenz-based antiretroviral regimen is preferred during rifampin-containing tuberculosis therapy. However, current pharmacokinetic data are insufficient to guide optimized concurrent dosing. This study aimed to better characterize the effects of rifampin on efavirenz pharmacokinetics. Subjects were randomized to receive 600 mg efavirenz/day or 600 mg efavirenz with 600 mg rifampin/day for 8 days, with plasma samples collected for pharmacokinetic analysis over 24 h on day 8. Treatments were then crossed over after at least a 2-week washout period, and procedures were repeated. Efavirenz concentrations were determined by high-performance liquid chromatography (HPLC), and pharmacokinetic parameters were estimated by noncompartmental analysis. Efavirenz pharmacokinetic differences between treatment periods were evaluated by paired t test. The coefficients of variation in efavirenz plasma AUC(0-24) (area under the concentration-time curve from 0 to 24 h) were 50% and 56% in the absence and presence of rifampin, respectively. Of the 11 evaluable subjects (6 white, 5 black; 6 women, 5 men), the geometric mean AUC(0-24) ratio on/off rifampin (90% confidence interval) was 0.82 (0.72, 0.92), with individual AUC(0-24) ratios varying from 0.55 to 1.18. Five subjects had a 24-hour efavirenz concentration (C(24)) of <1,000 ng/ml on rifampin. They were more likely to have received a lower dose in milligrams/kilogram of body weight and to have lower efavirenz AUC(0-24) values in the basal state. Although rifampin resulted in a modest reduction in efavirenz plasma exposure in subjects as a whole, there was high variability in responses between subjects, suggesting that efavirenz dose adjustment with rifampin may need to be individualized. Body weight and genetic factors will be important covariates in dosing algorithms.     

Morphine and metabolite behavior after different routes of morphine administration: demonstration of the importance of the active metabolite morphine-6-glucuronide

The pharmacokinetic parameters of morphine, morphine-6-glucuronide, and morphine-3-glucuronide were studied after single-dose morphine administration by five different routes. The quantitative significance of the active metabolite morphine-6-glucuronide was assessed, and the effects of novel dosing forms on morphine metabolism and distribution were examined. After administration of intravenous morphine the morphine-6-glucuronide plasma AUC exceeded that of morphine. After administration of oral morphine very low morphine levels were observed--the morphine-6-glucuronide plasma AUC exceeded that of morphine by a factor of 9:1. Sublingual, buccal, and sustained-release buccal morphine tablet administration resulted in delayed absorption, with attenuation and delay of peak morphine and metabolite levels. Morphine bioavailability and morphine glucuronide production were not altered.     

Mycophenolate pharmacokinetics in early period following lung or heart transplantation

BACKGROUND: The available pharmacokinetic and pharmacodynamic data on mycophenolic acid (MPA), the pharmacologically active metabolite of mycophenolate mofetil (MMF), are derived largely from renal transplant patients, not thoracic transplant recipients.OBJECTIVE: To evaluate, in a pilot study, the pharmacokinetics of MPA at 3 different times in the early period (up to the first 9 mo) following lung or heart transplantation.METHODS: Nine patients were entered into this open-label study. Upon administration of a steady-state morning MMF dose, blood samples were collected at 0, 20, 40, 60, and 90 minutes and at 2, 4, 6, 8, 10, and 12 hours after the dose at 3 times (denoted as sampling periods 1, 2, and 3) in the early posttransplant period. Total MPA concentrations were measured by a validated HPLC method with ultraviolet detection and followed by ultrafiltration of pooled samples for unbound MPA concentrations. Pharmacokinetic parameters (maximal concentration [C(max)], dose-normalized C(max), time to C(max), minimum concentration, predose concentration, AUC, dose-normalized AUC, free fraction, free AUC) were calculated by traditional noncompartmental methods.RESULTS: Patient characteristics included 7 men and 2 women, 5 lung and 4 heart transplant recipients, mean +/- SD age 53 +/- 11 years, and weight 77 +/- 14 kg. All patients were receiving prednisone and cyclosporine (with the exception of 2 pts. on tacrolimus during sampling periods 2 and 3). Sampling periods 1, 2, and 3 occurred on posttransplant days 15 +/- 13, 56 +/- 33, and 125 +/- 73, respectively. No significant differences were found between sampling periods in any pharmacokinetic parameter. Drug exposure as evaluated by AUC was 39.95 +/- 44.86, 25.24 +/- 25.68, and 43.96 +/- 38.67 micro g*h/mL during sampling periods 1, 2, and 3, respectively, (p > 0.05).CONCLUSIONS: As of September 26, 2003, this is the first study to systematically evaluate MPA pharmacokinetics in thoracic transplant recipients at 3 different time points during the early posttransplant period. Wide interpatient variability in MPA pharmacokinetics was observed, thus emphasizing the need to individualize dosing of MMF and to further evaluate important pharmacokinetic/pharmacodynamic parameters and endpoints that impact on clinical outcomes. Further studies involving more patients and pharmacodynamic outcomes are underway to help identify optimal MMF strategies.     

Pharmacokinetics of oral propafenone in patients with supraventricular arrhythmia

The efficacy of propafenone (P), a class IC antiarrhythmic drug with weak beta-blocking properties was studied over a four day period in 10 patients with supraventricular arrhythmias (atrial fibrillation 7, flutter 1 and tachycardia 2). Group 1 included five patients (3M, 2F) who received 300 mg of P on days 1 and 4. Group 2 included five patients (4M, 1F) who received 600 mg on days 1 and 4. All the patients received 1200 mg/day on days 2 and 3. Pharmacokinetics parameters were calculated for the first and the final dosing. Half of the patients were converted to sinus rythm after a delay ranging from 12 to 55 h after the first dosing. The duration of arrhythmia was shorter and the left atrial diameter was significantly lower in the responder group than in the non-responders. No relationship was observed between clinical efficacy and dose or plasma concentration of P. After the first administration of P, major interindividual variability in pharmacokinetic parameters was observed. Seven patients correspond to the extensive metabolizer phenotype with t1/2 el less than 10 h (mean: 5.4 +/- 2.2 SD). In this group t1/2 el increased from day 1 to day 4 and the AUC final/AUC initial ratio ranged between 4 to 17.5. Three patients showed the non-extensive metabolizer phenotype with t1/2 el ranging from 12.4 to 13.7 h and a moderate increase in AUC over chronic dosing. Adverse effects (cardiac conduction abnormalities, visual and digestive disturbances) were observed in the 3 oldest patients (70-73 yrs).(ABSTRACT TRUNCATED AT 250 WORDS)     

Metabolism of nomifensine after oral and intravenous administration

The metabolism of nomifensine was studied after single oral and intravenous administration and after 2 weeks of oral dosing. The three principal metabolites reached maximum plasma concentrations rapidly (in 1 to 1.5 hours) after nomifensine administration. Less than 10% was detected as a free, unconjugated form. All three metabolites were eliminated rapidly (elimination t1/2 values between 6.8 and 9.0 hours). Only very low concentrations of free metabolites were found in plasma after 24 hours of nomifensine administration. AUC values for free metabolites were between 0.27 to 0.46 hr X mumol/L after all nomifensine schedules. Two weeks of dosing had no significant influence on the elimination t1/2 or AUC values of the metabolites, indicating no change in the hydroxylation and methylation reactions. In addition, there were no changes in the conjugation reactions during prolonged nomifensine dosing. Nomifensine has a very short t1/2 and no tendency for accumulation after repeated doses. We conclude that nomifensine's clinical pharmacokinetic profile is not significantly changed by the kinetic behavior of its three main metabolites after the usual maintenance doses.     

Simultaneous determination and pharmacokinetic study of roxithromycin and ambroxol hydrochloride in human plasma by LC-MS/MS

BACKGROUND: Although roxithromycin and ambroxol HCl were often administered concomitantly for the treatment of respiratory infections, the pharmacokinetic interactions between them have not been reported. We investigated the interactions between these drugs in health male Chinese volunteers by LC-MS/MS in human plasma. METHODS: The pharmacokinetics were studied in 12 healthy male Chinese volunteers after an overnight fast by a single oral dose, 4-way crossover design with a period of 7-day washout. Each subjects was randomized to receive a single oral dose of 1 compound roxithromycin (150 mg) and ambroxol HCl (30 mg) dispersible tablet (test formulation, treatment A), one 150 mg roxithromycin dispersible tablet together with one 30 mg ambroxol HCl tablet (combined reference formulations, treatment B), one 150 mg roxithromycin dispersible tablet (reference formulation I, treatment C), or one 30 mg ambroxol HCl tablet (reference formulation II, treatment D) with 250 ml of water. Venous blood was collected at pre-dose (0 h) and 0.25, 0.5, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 12, 24, 48, 72 h after dosing. The plasma concentrations of roxithromycin and ambroxol HCl were simultaneously determined by using a validated internal standard LC-MS/MS method. RESULTS: No significant differences were observed for the major pharmacokinetic parameters such as C(max), T(max), t(1/2) and AUC of both roxithromycin and ambroxol HCl between different treatments. CONCLUSION: The pharmacokinetics of both roxithromycin and ambroxol HCl are not affected by their concomitant oral administration. Therefore, there are no obvious pharmacokinetic interactions between roxithromycin and ambroxol HCl after oral administration. Roxithromycin and ambroxol HCl dispersible tablets were bioequivalent with reference to the roxithromycin dispersible tablets and ambroxol HCl tablets in combination usage.     

Restoration of oral all-trans retinoic acid bioavailability after a brief drug holiday.

We evaluated the utility of a 7-day drug holiday in the restoration of chronic dosing all-trans-retinoic acid (tRA) blood levels in 11 non-small cell lung carcinoma patients. Baseline kinetic studies (day 1) were compared to postchronic dosing (day 7) and drug holiday kinetics (day 14). High levels of baseline pharmacokinetic variability and variability in response to prolonged tRA therapy were evident. Median area under the curve (AUC) decreased from a baseline level of 1.2 to 0.69 microg/ml/h (p = 0.03). t (1/2) showed no significant alterations. A near-significant increase in T (lag) (p = 0.08) was noted, which suggests modulation of absorbance parameters. Trends in AUC were strongly correlated with C (max) as was T (max) with T (lag). After a 7-day drug holiday the median AUC significantly increased from the day 7 value to 1.8 microg/ml/h p = 0.01). Since post-drug holiday values for parameters were not statistically different from baseline pharmacokinetic values, this suggests a complete restoration of tRA bioavailability.     

Pharmacokinetic and pharmacodynamic modeling of anidulafungin (LY303366): reappraisal of its efficacy in neutropenic animal models of opportunistic mycoses using optimal plasma sampling

The compartmental pharmacokinetics of anidulafungin (VER-002; formerly LY303366) in plasma were characterized with normal rabbits, and the relationships between drug concentrations and antifungal efficacy were assessed in clinically applicable infection models in persistently neutropenic animals. At intravenous dosages ranging from 0.1 to 20 mg/kg of body weight, anidulafungin demonstrated linear plasma pharmacokinetics that fitted best to a three-compartment open pharmacokinetic model. Following administration over 7 days, the mean (+/- standard error of the mean) peak plasma concentration (C(max)) increased from 0.46 +/- 0.02 microg/ml at 0.1 mg/kg to 63.02 +/- 2.93 microg/ml at 20 mg/kg, and the mean area under the concentration-time curve from 0 h to infinity (AUC(0-infinity)) rose from 0.71 +/- 0.04 to 208.80 +/- 24.21 microg. h/ml. The mean apparent volume of distribution at steady state (V(ss)) ranged from 0.953 +/- 0.05 to 1.636 +/- 0.22 liter/kg (nonsignificant [NS]), and clearance ranged from 0.107 +/- 0.01 to 0.149 +/- 0.00 liter/kg/h (NS). Except for a significant prolongation of the terminal half-life and a trend toward an increased V(ss) at the higher end of the dosage range after multiple doses, no significant differences in pharmacokinetic parameters were noted in comparison to single-dose administration. Concentrations in tissue at trough after multiple dosing (0.1 to 10 mg/kg/day) were highest in lung and liver (0.85 +/- 0.16 to 32.64 +/- 2.03 and 0.32 +/- 0.05 to 43.76 +/- 1.62 microg/g, respectively), followed by spleen and kidney (0.24 +/- 0.65 to 21.74 +/- 1.86 and <0.20 to 16.92 +/- 0.56, respectively). Measurable concentrations in brain tissue were found at dosages of > or =0.5 mg/kg (0.24 +/- 0.02 to 3.90 +/- 0.25). Implementation of optimal plasma sampling in persistently neutropenic rabbit infection models of disseminated candidiasis and pulmonary aspergillosis based on the Bayesian approach and model parameters from normal animals as priors revealed a significantly slower clearance (P < 0.05 for all dosage groups) with a trend toward higher AUC(0-24) values, higher plasma concentrations at the end of the dosing interval, and a smaller volume of distribution (P < 0.05 to 0.193 for the various comparisons among dosage groups). Pharmacodynamic modeling using the residual fungal tissue burden in the main target sites as the primary endpoint and C(max), AUC(0-24), time during the dosing interval of 24 h with plasma drug concentration equaling or exceeding the MIC or the minimum fungicidal concentration for the isolate, and tissue concentrations as pharmacodynamic parameters showed predictable pharmacokinetic-pharmacodynamic relationships in experimental disseminated candidiasis that fitted well with an inhibitory sigmoid maximum effect pharmacodynamic model (r(2), 0.492 to 0.819). However, no concentration-effect relationships were observed in experimental pulmonary aspergillosis using the residual fungal burden in lung tissue and survival as parameters of antifungal efficacy. Implementation of optimal plasma sampling in discriminative animal models of invasive fungal infections and pharmacodynamic modeling is a novel approach that holds promise of improving and accelerating our understanding of the action of antifungal compounds in vivo.     

Compartmental pharmacokinetics and tissue distribution of the antifungal triazole ravuconazole following intravenous administration of its di-lysine phosphoester prodrug (BMS-379224) in rabbits

OBJECTIVES: Ravuconazole is a broad-spectrum antifungal triazole in clinical development. We investigated the compartmental plasma pharmacokinetics and tissue distribution of ravuconazole following administration of its novel intravenous (i.v.) di-lysine phosphoester prodrug, BMS-379224. METHODS: Normal catheterized rabbits received the prodrug at 1.25, 2.5, 5, 10, 20 and 40 mg/kg once daily as 5 min i.v. bolus for 8 days. Serial plasma levels were collected at days 1 and 7, and tissues were obtained 30 min after the eighth dose. Concentrations of ravuconazole were determined by a validated HPLC method. Plasma concentration data were fitted to a three-compartment pharmacokinetic model. Pharmacokinetic parameters were estimated by weighted non-linear least squares regression analysis using the WinNonlin computer program. RESULTS: Following single dosing, ravuconazole demonstrated linear plasma pharmacokinetics across the investigated dosage range. Cmax, AUC(0-infinity), V(ss), CL and terminal half-life (means +/- SEM) ranged from 2.03 to 58.82 mg/L, 5.80 to 234.21 mg x h/L, 5.16 to 6.43 L/kg, 0.25 to 0.18 L/h/kg and 20.55 to 26.34 h, respectively. Plasma data after multiple dosing revealed non-linear disposition at the 20 and 40 mg/kg dosage levels as evidenced by a dose-dependent decrease in CL (from 0.104-0.147 to 0.030 and 0.022 L/h/kg; P = 0.1053) and an increase in the dose-normalized AUC(0-infinity) (from 2.40-3.01 up to 11.90 and 14.56 mg x h/L; P = 0.0382). Tissue concentrations 30 min after the last dose were highest in the liver (12.91-562.68 microg/g), adipose tissue (10.57-938.55 microg/g), lung (5.46-219.12 microg/g), kidney (3.95-252.44 microg/g) and brain tissue (2.37-144.85 microg/g). CONCLUSIONS: The pharmacokinetics of ravuconazole fitted best to a three-compartment pharmacokinetic model. The compound revealed non-linear pharmacokinetics at higher dosages, indicating saturable clearance and/or protein binding. Ravuconazole displayed a long elimination half-life and achieved substantial plasma and tissue concentrations including in the brain.