Non-Adherence in Adult Male Patients with Community-Acquired Pneumonia: Relative Forgiveness of Amoxicillin versus Respiratory Fluoroquinolones.

The consequences of non-adherence to treatment (NAT) on antimicrobial efficacy may depend on drug forgiveness-a property that should account for pharmacokinetics (PK) and pharmacodynamics (PD) as well as interindividual variability. In this simulation study, relative forgiveness (RF) in NAT, defined as the probability of a successful PK/PD target (PTA) attained under perfect adherence compared to imperfect adherence, was evaluated for amoxicillin (AMOX) (oral 1000 mg/8 h) and two respiratory fluoroquinolones-levofloxacin (LFX) (oral 750 mg/24 h) and moxifloxacin (MOX) (oral 400 mg/24 h)-in virtual outpatients with community-acquired pneumonia for S. pneumoniae. Several NAT scenarios (delay in dose intake and a missed dose) were considered. PK characteristics of virtual patients, including variability in creatinine clearance (70-131 mL/min) and S. pneumoniae susceptibility variability associated with geographical location, were simulated in NAT. In this regard, in regions of low MIC delays from 1 h to 7 h or omission of dose ingestion would not have negative consequences on the efficacy of AMOX because of its good RF associated with the AMOX PK and PD properties; RF of LFX 750 mg or MOX 400 mg/24 h regimen vs. AMOX 1000 mg/8 h is one. However, in regions of elevated MIC for S. pneumoniae AMOX loses its RF, LFX and MOX vs. AMOX, showing higher RF (>1) depending on the CLCR of patients. These results illustrate the importance of considering the RF of antimicrobial drugs in NAT and provide a framework for further studying its implications for clinical success rates.

Application of a dual mechanistic approach to support bilastine dose selection for older adults.

The objective of this study was to evaluate bilastine dosing recommendations in older adults and overcome the limitation of insufficient data from phase I studies in this underrepresented population. This was achieved by integrating bilastine physicochemical, in vitro and in vivo data in young adults and the effect of aging in the pharmacology by means of two alternative approaches: a physiologically-based pharmacokinetic (PBPK) model and a semi-mechanistic population pharmacokinetic (Senescence) model. Intestinal apical efflux and basolateral influx transporters were needed in the PBPK model to capture the observations from young adults after single i.v. (10 mg) and p.o. (20 mg) doses, supporting the hypothesis of involvement of gut transporters on secretion. The model was then used to extrapolate the pharmacokinetics (PKs) to elderly subjects considering their specific physiology. Additionally, the Senescence model was develop starting from a published population PK) model, previously applied for pediatrics, and incorporating declining functions on different physiological systems and changes in body composition with aging. Both models were qualified using observed data in a small group of young elderlies (N = 16, mean age = 68.69 years). The PBPK model was further used to evaluate the dose in older subjects (mean age = 80 years) via simulation. The PBPK model supported the hypothesis that basolateral influx and apical efflux transporters are involved in bilastine PK. Both, PBPK and Senescence models indicated that a 20 mg q.d. dose is safe and effective for geriatrics of any age. This approach provides an alternative to generate supplementary data to inform dosing recommendations in under-represented groups in clinical trials.

Model-informed pediatric development applied to bilastine: Analysis of the clinical PK data and confirmation of the dose selected for the target population.

Bilastine is a non-sedating second-generation H1 antihistamine approved for treatment of allergic rhinoconjunctivitis (AR) and urticaria (U) in adults at the oral (p.o.) dose of 20 mg once daily (OD). Optimal attributes can be anticipated for its clinical use in pediatrics due to its favorable safety and tolerability and age-independent PD profile. The aim of this work was to characterize bilastine PK in children through population modeling of data from a limited sampling confirmatory clinical trial in children with AR or U. The objective was also to ascertain whether the proposed dose (10 mg/day) in the target pediatric subset aged 2-<12 years matches the systemic exposure seen in adults at the 20 mg/day dose. A popPK model characterizing bilastine PK behavior in children aged from 4 to <12 years treated with 10 mg oral bilastine daily was successfully developed and qualified. No relationship was found between bilastine PK and age or weight; stopping rules pre-stablished to finalize the trial, i.e., model completeness and no dependence of exposure on decreasing age, were thus fulfilled. On a second step, the popPK model in children was linked to the PD model in adults assuming the same PD as described in adults and used to compare the PD outcome between both populations. Finally, an allometric scaling method and a physiological approximation were used to evaluate the suitability of the selected dose in the youngest children, showing that children from 2 years were deemed to belong to the same population as well. The achievement of comparable PK (i.e., within the range) to that observed in adults after the therapeutic dose of 20 mg, together with the achievement of similar PD and additional integrative analysis, served to confirm the validity of the 10 mg daily dose for the target pediatric subset (2 to <12 years).

Time Scaling for In Vitro-In Vivo Correlation: the Inverse Release Function (IRF) Approach.

In vitro-in vivo correlations (IVIVC) are methods used to create a link between biopharmaceutical properties such as dissolution and physiological response such as plasma concentration. Level A IVIVC defines 1:1 relationship between the percent absorbed in vivo and the percent dissolved in vitro. A successful level A IVIVC provides the capacity to predict in vivo behavior based only on in vitro data with application in formulation development and support of biowaivers recognized by regulatory agencies across the world. Level A regression may be complicated due to differences in time scales as well as the lack of coincident times of similar release in vitro and in vivo leading to approximate time-to-time links and subsequent loss of information. Here, a novel method to establish Levy's plot and to provide time scaling for improved IVIVC predictive capacity is presented. The method is mathematically closed and is an inverse release function (IRF) characterizing the single (or more) phases of dissolution/absorption. It uses the complete set of information available from all time points both in vitro and in vivo. An extended-release formulation development situation is presented with three increasing release rate test products compared in a trial versus a reference product. First, the standard level A regression was made. Prediction errors for internal validation were higher than 10% for Cmax. The IRF method was applied to obtain the in vitro times of percentage dissolved equivalent to percentage absorbed. The prediction errors from the IRF level A correlation were nearly negligible.

Model Informed Pediatric Development Applied to Bilastine: Ontogenic PK Model Development, Dose Selection for First Time in Children and PK Study Design.

PURPOSE: Bilastine is an H1 antagonist whose pharmacokinetics (PK) and pharmacodynamics (PD) have been resolved in adults with a therapeutic oral dose of 20 mg/day. Bilastine has favorable characteristics for use in pediatrics but the PK/PD and the optimal dose in children had yet to be clinically explored. The purpose is to: (1) Develop an ontogenic predictive model of bilastine PK linked to the PD in adults by integrating current knowledge; (2) Use the model to design a PK study in children; (3) Confirm the selected dose and the study design through the evaluation of model predictability in the first recruited children; (4) Consider for inclusion the group of younger children (< 6 years). METHODS: A semi-mechanistic approach was applied to predict bilastine PK in children assuming the same PD as described in adults. The model was used to simulate the time evolution of plasma levels and wheal and flare effects after several doses and design an adaptive PK trial in children that was then confirmed using data from the first recruits by comparing observations with model predictions. RESULTS: PK/PD simulations supported the selection of 10 mg/day in 2 to <12 year olds. Results from the first interim analysis confirmed the model predictions and design hence trial continuation. CONCLUSION: The model successfully predicted bilastine PK in pediatrics and optimally assisted the selection of the dose and sampling scheme for the trial in children. The selected dose was considered suitable for younger children and the forthcoming safety study in children aged 2 to <12 years.

Evaluation of fentanyl disposition and effects in newborn piglets as an experimental model for human neonates.

BACKGROUND: Fentanyl is widely used off-label in NICU. Our aim was to investigate its cerebral, cardiovascular and pulmonary effects as well as pharmacokinetics in an experimental model for neonates. METHODS: Fentanyl (5 µg/kg bolus immediately followed by a 90 minute infusion of 3 µg/kg/h) was administered to six mechanically ventilated newborn piglets. Cardiovascular, ventilation, pulmonary and oxygenation indexes as well as brain activity were monitored from T = 0 up to the end of experiments (T = 225-300 min). Also plasma samples for quantification of fentanyl were drawn. RESULTS: A "reliable degree of sedation" was observed up to T = 210-240 min, consistent with the selected dosing regimen and the observed fentanyl plasma levels. Unlike cardiovascular parameters, which were unmodified except for an increasing trend in heart rate, some of the ventilation and oxygenation indexes as well as brain activity were significantly altered. The pulmonary and brain effects of fentanyl were mostly recovered from T = 210 min to the end of experiment. CONCLUSION: The newborn piglet was shown to be a suitable experimental model for studying fentanyl disposition as well as respiratory and cardiovascular effects in human neonates. Therefore, it could be extremely useful for further investigating the drug behaviour under pathophysiological conditions.

Integration of preclinical and clinical knowledge to predict intravenous PK in human: bilastine case study.

Modern pharmacometrics can integrate and leverage all prior proprietary and public knowledge. Such methods can be used to scale across species or comparators, perform clinical trial simulation across alternative designs, confirm hypothesis and potentially reduce development burden, time and costs. Crucial yet typically lacking in integration is the pre-clinical stage. Prediction of PK in man, using in vitro and in vivo studies in different animal species, is increasingly well theorized but could still find wider application in drug development. The aim of the present work was to explore methods for bridging pharmacokinetic knowledge from animal species (i.v. and p.o.) and man (p.o.) into i.v. in man using the antihistamine drug bilastine as example. A model, predictive of i.v. PK in man, was developed on data from two pre-clinical species (rat and dog) and p.o. in man bilastine trials performed earlier. In the knowledge application stage, two different approaches were used to predict human plasma concentration after i.v. of bilastine: allometry (several scaling methods) and a semi-physiological method. Both approaches led to successful predictions of key i.v. PK parameters of bilastine in man. The predictive i.v. PK model was validated using later data from a clinical study of i.v. bilastine. Introduction of such knowledge in development permits proper leveraging of all emergent knowledge as well as quantification-based exploration of PK scenario, e.g. in special populations (pediatrics, renal insufficiency, comedication). In addition, the methods permit reduction or elimination and certainly optimization of learning trials, particularly those concerning alternative off-label administration routes.

Tacrolimus dose individualization in "de novo" patients after 10 years of experience in liver transplantation: pharmacokinetic considerations and patient pathophysiology.

AIM: To determine how changes in tacrolimus (TAC) immunosuppression clinical practice, in the first 15 days post liver transplantation (LT) and across a decade, impact a clinical covariate - pharmacokinetic (PK) model, developed in data from 1998, thus testing its utility in dose individualization across time. Patient cohorts from 1998 (Reference: R-1998) and 2007 (EVALUATION: E-2007) were compared. METHODS: Analysis of monitoring observations (Cmin and Cmin/dose) and the biochemical variables aspartate aminotransferase (AST), hematocrit (HCT), albumin (ALB) and serum creatinine (SCr) was done for 0 - 3 and 4 - 15 days post transplantation (PT). The population PK model developed for R-1998 [1] was re-evaluated for the two cohorts. RESULTS: Significant differences in R-1998 vs. E-2007 existed in Cmin and Cmin/dose and in covariates AST (as hepatic function marker) and SCr (as toxicity marker). E-2007 had lower levels of Cmin and Cmin/dose (1/CL), lower AST with faster recovery and lower variability in Cmin/dose for similar dose. AST was a covariate on CL/F in the 0 - 3 day PT period. In 4 - 15 days PT for E-2007, low levels of HCT and ALB as CL/F predictors confirmed a subgroup with higher CL/F (23.8 l/h vs. 19.3 l/h). The R-1998 model's original structure was confirmed. CONCLUSIONS: Ten years of use of TAC shows gain in therapeutic targeting efficiency, due to improvement in LT methods, knowledge of the drug and consideration of PK steady state. The remaining uncertainty with TAC monitoring in LT can be resolved with application of PK principles combined with patients' diosyncrasies in the model developed for TAC dose individualization in R-1998. The applicability of the model as nucleus in Bayes individualization remains intact across a decade.

A predictive pharmacokinetic/pharmacodynamic model of fentanyl for analgesia/sedation in neonates based on a semi-physiologic approach.

BACKGROUND AND OBJECTIVES: Fentanyl is a synthetic opioid commonly used as an anesthetic and also increasingly popular as a sedative agent in neonates. Initial dosage regimens in this population are often empirically derived from adults on a body weight basis. However, ontogenic maturation processes related to drug disposition are not necessarily always body weight correlates. We developed a predictive pharmacokinetic/pharmacodynamic model that includes growth and maturation physiologic changes for fentanyl in neonatal care. METHODS: Key pharmacokinetic variables and principles (protein binding, clearance, distribution) as related to fentanyl pharmacokinetic/pharmacodynamic behavior in adults (tricompartmental model) and to neonatal physiologic data (organ weights and blood flows, body composition, renal and hepatic function, etc.) were used to guide the building of a semi-physiologic ontogenic model. The model applies to a normal-term neonate without any other intervention. Then, extension to a pharmacokinetic/pharmacodynamic link model for fentanyl was made. The final model was evaluated by predicting the time course of plasma concentrations and the effect of a standard regimen of 10.5 µg/kg over a 1-h period followed by 1.5 µg/kg/h for 48 h. RESULTS: Hepatic clearance was linked to ontogeny of unbound fraction and of α1-acid glycoprotein. All parameters were reduced in the neonate compared to adults but in a differing proportion due to qualitative changes in physiology that are analyzed and accounted for. Systemic clearance (CLS), volume of the central compartment (V1) and steady-state volume of distribution predicted by the model were 0.028 L/min, 1.26 L, and 22.04 L, respectively. Weight-corrected parameters generally decreased in adults compared with neonates, but differentially, e.g., CLS = 0.0093 versus 0.0088 L/min/kg, while V1 = 0.42 versus 0.18 L/kg (neonates vs. adults). Under such complexity a pharmacokinetic/pharmacodynamic model is the appropriate method for rational efficacy targeting. Fentanyl pharmacodynamics in neonates were considered to be similar to those in adults except for the equilibrium rate constant, which was also scaled on an ontogenic basis. The model adequately predicted the reported mean expected concentration-time profiles for the standard regimen. CONCLUSIONS: Integrated pharmacokinetic/pharmacodynamic modeling showed that the usually prescribed dosage regimens of fentanyl in neonates may not always provide the optimum degree of sedation. The model could be used in optimal design of clinical trials for this vulnerable population. Prospective clinical testing is the reasonable next step.

Tacrolimus pharmacokinetics in the early post-liver transplantation period and clinical applicability via Bayesian prediction.

PURPOSE: To define and validate a pharmacokinetic (PK) model for tacrolimus (TAC) that includes patient pathophysiology and has clinical applicability in the first 2 weeks post-liver transplantation (PLT). METHODS: Routine monitoring records [dose, trough levels (C(min)), demographics, biochemistry] from 75 patients treated with TAC (Prograf®) PLT were used to develop a population PK model (employing NONMEM®) testing for predictors of oral clearance (CL/F) according to bedside evidence and primarily with aspartate aminotransferase (AST), albumin (ALB), and hematocrit (HCT). Patients were catergorized into subgroups with above and below "normal" thresholds for AST (500 U/L), ALB (2.5 g/dL), and HCT (28 %), respectively. The model was validated with ten patients from the same period and 15 more recent patients. An empirical Bayes method was developed and applied to the prediction of individual profiles serving as a dose adjustment tool. RESULTS: The number of days PLT (Days PLT) was a key variable during the first 2 weeks, with a dichotomy in the mono-compartmental parameters for 0-3 Days PLT and 4-15 Days PLT. During 0-3 Days PLT, AST levels, indicative of allograft functionality (and TAC metabolism), were crucial predictors of elimination. Three groups were identified with the following clearances: CL/F0₋3 = 8.93 L/h for AST ≥ 500 U/L and CL/F0₋3 = 11.0 L/h for AST <500 U/L. During 4-15 Day PLT, low values of ALB (<2.5 g/dL) and HCT (<28 %) combined were determinant of a patient subgroup with a tendency to underexposure and complexity in empirical dose adjustment. The CL/F4₋15 = 25.1 L/h for this subgroup compared to CL/F4₋15 = 17.1 L/h for the others in that period. The elimination half-life for individual patients varied over tenfold so that a large number of subjects were not at steady state, making the use of a PK model necessary to achieve rapidly and safely the target concentration for TAC in LT. Validation of the model demonstrated that both bias and precision were within acceptable limits. CONCLUSION: For TAC therapy, covariate models using mixed effects methods are most useful when combined with patient-specific biochemical assays as well as clinical evidence. In such cases, the observed C(min) and Bayes methods can provide the most likely individual PK parameters, hence the optimal next dose to reach individualized target levels for each patient.

Pathophysiological idiosyncrasies and pharmacokinetic realities may interfere with tacrolimus dose titration in liver transplantation.

PURPOSE: To explore the main factors that make it difficult to empirically monitor tacrolimus (TAC) in the early period post-liver transplantation (LTx), with a specific focus on those aspects related to patient idiosyncrasy and clinical status as well as to the pharmacokinetic (PK) assumptions on which drug individualization in clinical practice is based. METHODS: Retrospective monitoring data from 75 de novo liver transplant patients treated with twice daily with TAC and followed for up to 15 days were analyzed. An extensive battery of laboratory measurements were available. Dose adjustment was performed empirically using trough levels (C(min)). The population was separated into two major background groups according to low or high values of aspartate aminotransferase (AST) (Group 1 and 2, respectively) based on AST measurements made during the first 4 days post-LTx. Each of these two major groups was then further subdivided into two subgroups based on elevated (Groups 1A, 2A) or reduced (Groups 1B, 2B) combined albumin (cut-off 2.5 g/dl) and hematocrit (cut-off 28%). RESULTS: The C(min)/Dose ratio [inversely proportional to systemic clearance (CL)] had a variability [coefficient of variation (CV) >80%) that was incongruently higher for the ratio than for C(min) and Dose separately. This was attributed to most patients not being at steady state or physiologically stable in the early post-LTx period. Group 1 patients were more predictable than Group 2 patients, who were responsible for the variability in the ratio. C(min) was lower in the reduced ALB and HCT patient groups when AST conditions were similar (1A vs. 1B and 2A vs. 2B), likely due to increased TAC metabolic clearance (reduced C(min)/Dose). This situation existed for two periods: 0-15 days post-LTx and 4-15 days post-LTx observations. Group 2A patients were the main source of the paradoxical variability in C(min)/Dose (higher ratio of 2.7; CV = 100%), suggesting a lower clearance and difficulty in the recovery of stability. In contrast, Group 2B patients had the lower ratio (1.4; 47%) but required the highest number of dose adjustments as the variability was hard to identify clinically. Group 1A patients were the most predictable empirically. When observations from 15 new patients who entered the clinic in 2007 and 2008 were used for the analysis, the same sub-groups existed in the same proportions in both years. CONCLUSION: The difficulty in empirical dose adjustment of TAC is associated to the inevitable non-fulfillment of PK assumptions early post-LTx and also to the inherent complexity of the clinical condition, leading to increased uncertainty for the clinician regarding dose selection. Identifying these sub-categories provides a rational means of classifying patients akin to a phenotype. The complexity of the kinetics in LTx and TAC treatment does not invalidate C(min) as a biomarker, but a Bayes algorithm including a full PK structure and these covariates would be optimal.

Simulation of sirolimus exposures and population variability immediately post renal transplantation: importance of the patient's CYP3A5 genotype in tailoring treatment.

The rapid achievement of efficacious exposure to sirolimus (SRL) after renal transplantation is crucial. However, there is high unpredictability in the dose to exposure relationship. Part of the variation is related to patients originating from subpopulations of fast or slow metabolizers via the CYP3A5*1/*3 genotype. The probability of achieving therapeutic SRL blood concentrations for each subpopulation under two equal-intensity increasing-frequency protocols after the start of treatment was explored with Monte Carlo simulation. The population pharmacokinetic model and inter-patient variability distributions of Djebli et al. (DRH2006) were sampled. They developed a base and final model with a genotype covariate for CL/F in patients receiving calcineurin inhibitor (CNI)-free therapy with SRL, mycophenolate mofetil and corticosteroids. Fast metabolizers (expressers) had a CL/F of 28.3 l/h whilst slow metabolizers (non-expressers) had a CL/F of 14.1 l/h. Here, in simulation, a standard 10 mg QD SRL was contrasted with a higher frequency of 5 mg BID SRL as related to the proportion of next dosed patients being within the 15-30 ng/ml trough levels on day 7 after transplantation. Near 0% of expressers on either regimen reached or exceeded the 30 ng/ml trough on day 7. Expressers showed protocol dependence for the chance of being within the 15-30 ng/ml range with the 5 mg BID protocol doubling those chances. Non-expressers appeared less protocol dependent for the probability of being above or below the 15-30 ng/ml range. The ability to determine the genotype early on may help to rationalize the initial titration of individual patients receiving CNI-free renal transplantation treatment with SRL.

Population pharmacokinetics of recombinant factor VIII:C (ReFacto) in adult HIV-negative and HIV-positive haemophilia patients.

PURPOSE: The aim of this study was to explore possible differences in the pharmacokinetics (PK) of recombinant factor VIII:C (ReFacto - ReFacto ) in HIV+ vs. HIV- patients and also differences in the chromogenic substrate bioassay (CHS) and one-stage clotting (OSC) methods. METHODS: Twenty-eight haemophilia A adults (20 HIV- and eight HIV+) were assayed with both the CHS and OSC methods. An average of two and six samples were collected per patient for HIV-/+, respectively, after one, and occasionally two more, prophylactic doses (mean 2,003 IU; range 1,000-4,300 IU). The observations were analysed with the mixed-effects (population) compartmental PK modelling package NONMEM (nonlinear mixed-effects modelling) and the FOCE (first-order conditional estimation) method. Base modelling was performed independently for the CHS and OSC bioassays for comparison, and covariate models and simulation tests were done only for the commonly used OSC bioassay. The final covariate model was validated using the bootstrap method. Monte Carlo simulations were used to estimate the expected probability of exceeding 20%, 40% or 60% of normal factor VIII:C in plasma after a single dose, corresponding to required levels for preventing mild, moderate and life-threatening haemorrhages. RESULTS: One-compartment base-model population PK parameters were [mean parameter (interpatient variability %)] for CHS: clearance (CL) = 2.56 dl h(-1) (33.2%); volume of distribution (V) = 34.8 dl (12.8%); and for OSC: CL = 3.83 dl h(-1) (47.8%), V = 53.7 dl (22.4%). The volumes differed significantly between the CHS and OSC methods (p < 0.0001), and variabilities were higher for OSC. Nevertheless, the empirical half-lives (t(1/2) = l n (2) x V/CL) were similar for CHS and OSC, [(mean +/- standard deviation (SD)], 9.5 +/- 3 h and 10.2 +/- 4 h, respectively. In covariate modelling with the OSC-derived model, HIV status (VIR) was a significant categorical predictor (p < 0.005) for V. The final covariate models with OSC were for CL = 3.93 + 0.09 x (WT-75) and for V = 48.6 x (1 + 0.36 x VIR) + 0.55 x (WT-75); therefore, V for the typical HIV+ patient was 36% higher than for the HIV- patient. CONCLUSIONS: Both HIV- and HIV+ patients showed 100% success with the 20% threshold at doses >20 IU/kg. HIV- patients receiving >50 IU/kg had a 100% expected chance of success for all thresholds. HIV+ patients for moderate or life-threatening haemorrhage treatment need 10 IU/kg more than the HIV- patient equivalent to have the same probability of success.

Comparative evaluation of positive tests to Mycobacterium avium subsp. paratuberculosis in clinically healthy sheep and goats in south-west Greece using molecular techniques, serology, and culture.

Mycobacterium avium subsp. paratuberculosis (MAP) is the cause of paratuberculosis, which affects mainly ruminants although there is a growing concern about its possible implication in Crohn's disease in humans especially in connection with environmental spread and risks to the food chain. Retail cheese may represent a significant source of human exposure to MAP and the aim of this study was to assess MAP status in clinically healthy sheep and goats in Greece, comparing techniques routinely used in the positive diagnosis of the disease. From a total of 30 flocks, 632 sheep and goats had faecal, serum, and whole-blood samples examined by culture, complement fixation test (CFT), and polymerase chain reaction (PCR) targeted at IS900, IS1245, and IS6110. PCR produced positive results in 21% of the animals tested, with 5.6%, 3.9%, and 11.5% being identified as MAP, Mycobacterium avium subsp. avium, and Mycobacterium tuberculosis complex, respectively. CFT produced positive and suspicious results in 4.4% and 14.4% of the cases. Faecal cultures were negative in all but a single case that was identified as restriction fragment length polymorphism (RFLP)-type BC1. Agreement between results obtained by PCR and CFT was poor with isolated cases although an assessment of the MAP positive tests produced similar results for both methods. The findings indicate the need for additional measures of control, although the costs may be substantial if public health protection justifies elimination of MAP from livestock.

Population pharmacokinetics of high-dose methotrexate in children with acute lymphoblastic leukaemia.

OBJECTIVE: To develop and a priori validate a methotrexate population pharmacokinetic model in children with acute lymphoblastic leukaemia (ALL), receiving high-dose methotrexate followed by folinic acid rescue, identifying the covariates that could explain part of the pharmacokinetic variability of methotrexate. METHODS: The study was carried out in 49 children (aged 6 months to 17 years) who received high-dose methotrexate (3 g/m(2) per course) in long-term treatment. In an index group (37 individuals; 1236 methotrexate plasma concentrations), a population pharmacokinetic model was developed using a nonlinear mixed-effects model. The remaining patients' data (12 individuals; 278 methotrexate plasma concentrations) were used for model validation. Age, sex, total bodyweight (TBW), height, body surface area, lowest urine pH during infusion, serum creatinine, ALT, AST, folinic acid dose and length of rescue were analysed as possible covariates. The final predictive performance of the pharmacokinetic model was tested using standardised mean prediction errors. RESULTS: The final population pharmacokinetic model (two-compartmental) included only age and total bodyweight as influencing clearance (CL) and volume of distribution of central compartment (V(1)). For children aged < or =10 years: CL (L/h) = 0.287 . TBW(0.876); V(1) (L) = 0.465 . TBW, and for children aged >10 years: CL (L/h) = 0.149 . TBW; V(1) (L) = 0.437 . TBW. From the base to the final model, the inter-individual variabilities for CL and V(1) were significantly reduced in both age groups (30-50%). The coefficients of variation of the pharmacokinetic parameters were <30%, while residual and inter-occasional coefficients maintained values close to 40%. Validation of the proposed model revealed the suitability of the model. CONCLUSION: A methotrexate population pharmacokinetic model has been developed for ALL children. The proposed model could be used in Bayesian algorithms with a limited sampling strategy to estimate the systemic exposure of individual patients to methotrexate and adapt both folinic acid rescue and methotrexate dosing accordingly.

The role of unbound drug in pharmacokinetics/pharmacodynamics and in therapy.

The evolution of research on drug protein binding is discussed with the unbound concentration (Cu) and the unbound fraction (fu) as protagonists. Particular attention is paid to the mechanisms via which alterations in binding affect the pharmacokinetics (PK) and the effect, or independently the pharmacodynamics (PD). Apart from albumin, the important alpha-acid glycoprotein (AGP), as well as specific drug classes and applications in the clinic and development (routine monitoring, cancer and HIV therapy, allometry) are addressed. The flaws with the classical method of indirectly calculating the Cu or the unbound PK/PD parameters, based on the fu in vitro, are related to the intrinsic complexity and variability in the outcomes. Increased focus is urged on directly estimating the unbound PK/PD and also on using population statistical methods.

The infectivity of sarcoid clinical material and its bacterial content inoculated in CBA mice.

BACKGROUND: Sarcoidosis is a multisystemic disorder that is currently viewed as the consequence of chronic immunological response associating genetic susceptibility and specific environmental or transmissible agents. Relevant evidence, although conflicting, justifies a concern about the involvement of specific pathogens to disease causation. In this study we assessed the infectivity of sarcoid clinical material, and of the pathogens found in it, to normal CBA mice used as a model of an immuno-competent host. MATERIALS AND METHODS: One hundred and eleven mice were inoculated into their footpads with fresh, filtered, and autoclaved, sputum and bronchoalveolar lavage homogenates, collected from patients with sarcoidosis, and with the mycobacterial and propionibacterial pathogens isolated from this material. RESULTS: The total number of positive reactors of the animals that received raw clinical material and the pathogens it contained was statistically significant compared to those of the control groups. However, the number of affected mice per group was in most cases less than 50% and inflammation was almost always mild and local. CONCLUSION: Based on the evidence provided by inoculation of normal CBA mice, some of the material under study, although of mild potency, can be infectious to an immuno-competent host.

Mycobacterium avium subsp. paratuberculosis, genetic susceptibility to Crohn's disease, and Sardinians: the way ahead.

The present study was performed to determine what proportion of people in Sardinia with or without Crohn's disease were infected with Mycobacterium avium subspecies paratuberculosis and had a preponderance of allelic variants of Nod 2, an intracellular protein involved in Crohn's disease susceptibility. Genetic analysis of the alleles of the NOD 2/CARD 15 gene (ins C 3020, G 908 R, and R 702 W alleles), linked to susceptibility or genetic predisposition to Crohn's disease in humans, was carried out on specimens from 37 Crohn's disease patients and 34 patients without Crohn's disease. Our results show that more than 70 percent of people in Sardinia with Crohn's disease carry at least one of the susceptibility-associated NOD 2/CARD 15 alleles and were infected with Mycobacterium avium subspecies paratuberculosis.

Pharmacokinetic-pharmacodynamic modeling of the hydroxy lerisetron metabolite L6-OH in rats: an integrated parent-metabolite model.

PURPOSE: The twofold aim of this study was to characterize in vivo in rats the pharmacokinetics (PK) and pharmacodynamics (PD) of L6-OH, a metabolite of lerisetron with in vitro pharmacological activity, and evaluate the extent to which L6-OH contributes to the overall effect. METHODS: The PK of L6-OH was determined directly postmetabolite i.v. dose (PK-1), and also simultaneously for L (lerisetron concentration) and for generated L6-OH after lerisetron dose (200 microg kg(-1), i.v.), using Nonlinear Mixed Effects Modeling with an integrated parent-metabolite PK model (PK-2). Surrogate effect was measured by inhibition of serotonin-induced bradycardia. Protein binding was assayed via ultrafiltration and all quantification was performed via liquid chromatography-electrospray ionization-mass spectrometry. RESULTS: L6-OH showed elevated plasma and renal clearances, and volume of distribution (PK-1). The in vivo potency (PD) of L6-OH was high (EC(50) = 0.098 ng mL(-1) and EC(50unbound) = 0.040 ng mL(-1)). Total clearance for L (PK-2) in the presence of generated L6-OH (CL(L) = CL(-->L6-OH) + CL(n)) was 0.0139 L min(-1). Most of this clearance was L6-OH formation (F(c) = 99.6%), but only an 8.6% fraction of L6-OH was released into the bloodstream. The remainder undergoes biliar and fecal elimination. The parameters estimated from PK-2 were used to predict concentrations of L6-OH (Cp(L6)) generated after a lerisetron therapeutic dose (10 microg kg(-1)) in the rat. These concentrations are needed for the PD model and are below the quantification limit. Cp(L6max) was less than the EC(50) of L6-OH. CONCLUSIONS: We conclude that after lerisetron administration, L6-OH is extensively formed in the rat but it is quickly eliminated; therefore, besides being equipotent with the parent drug, the L6-OH metabolite does not influence the effect of lerisetron.