Dominique Breilh

Steady-state plasma and intrapulmonary concentrations of cefepime administered in continuous infusion in critically ill patients with severe nosocomial pneumonia.

ObjectiveTo determine the steady-state plasma and epithelial lining fluid concentrations of cefepime administered in continuous infusion in critically ill patients with severe bacterial pneumonia. DesignProspective, open-label study. SettingAn intensive care unit and research ward in a university hospital. PatientsTwenty adult patients with severe nosocomial bacterial pneumonia on mechanical ventilation were enrolled. InterventionsAll subjects received a 30-min intravenous infusion of cefepime 2 g followed by a continuous infusion of 4 g over 24 hrs. The concentrations of cefepime in plasma and epithelial lining fluid were determined at steady state after 48 hrs of therapy with high performance liquid chromatography. Measurements and Main ResultsThe mean ± sd steady-state plasma and epithelial lining fluid concentrations of cefepime 4 g in continuous infusion were 13.5 ± 3.3 &mgr;g/mL and 14.1 ± 2.8 &mgr;g/mL, respectively, with a mean percentage penetration of cefepime into epithelial lining fluid of about 100%. ConclusionsThe administration of 4 g of cefepime in continuous infusion in critically ill patients with severe nosocomial pneumonia appears to optimize the pharmacodynamic profile of this &bgr;-lactam by constantly providing concentrations in excess of minimal inhibitory concentration of most of susceptible organisms over the course of therapy in both serum and epithelial lining fluid.

Human Immunodeficiency Virus Type 1 Genotypic and Pharmacokinetic Determinants of the Virological Response to Lopinavir-Ritonavir-Containing Therapy in Protease Inhibitor-Experienced Patients

ABSTRACT The response to regimens including lopinavir-ritonavir (LPV/r) in patients who have received multiple protease (PR) inhibitors (PI) can be analyzed in terms of human immunodeficiency virus type 1 (HIV-1) genotypic and pharmacokinetic (pK) determinants. We studied these factors and the evolution of HIV-1 resistance in response to LPV/r in a prospective study of patients receiving LPV/r under a temporary authorization in Bordeaux, France. HIV-1 PR and reverse transcriptase sequences were determined at baseline LPV/r for all the patients and at month 3 (M3) and M6 in the absence of response to treatment. pK measurements were determined at M1 and M3. Virological failure (VF) was defined as a plasma viral load ≥400 copies/ml at M3. A multivariate analysis of the predictors of VF, including clinical and biological characteristics and the treatment history of the patients, was performed. The PR gene sequence at M0, including individual mutations or a previously defined LPV mutation score (D. J. Kempf, J. D. Isaacson, M. S. King, S. C. Brun, Y. Xu, K. Real, B. M. Bernstein, A. J. Japour, E. Sun, and R. A. Rode, J. Virol. 75:7262-7269, 2001), and the individual exposure to LPV were also included covariates. Sixty-eight patients were enrolled. Thirty-four percent had a virological response at M3. An LPV mutation score of >5 mutations, the presence of the PR I54V mutation at baseline, a high number of previous PIs, prior therapy with ritonavir or indinavir, absence of coprescription of efavirenz, and a lower exposure to LPV or lower LPV trough concentrations were independently associated with VF on LPV/r. Additional PI resistance mutations, including primary mutation I50V, could be selected in patients failing on LPV/r. Genotypic and pK parameters should be used to optimize the virological response to LPV/r in PI-experienced patients and to avoid further viral evolution.

Virological, intracellular and plasma pharmacological parameters predicting response to lopinavir/ritonavir (KALEPHAR study).

Objectives: To assess the impact of HIV-1 protease mutations and intracellular and plasma lopinavir minimum concentrations (Cmin) on virological success or failure on lopinavir/ritonavir-containing highly active antiretroviral therapy (HAART). Design: HIV-1-infected HAART-experienced patients included in an observational study, received lopinavir/ritonavir (400/100 mg twice a day) plus two to three nucleoside reverse transcriptase inhibitors (NRTI) or one NRTI plus one non-NRTI. A viral load less than 50 copies/ml at month 6 defined virological success. Methods: Intracellular and plasma lopinavir concentrations were determined by high-pressure liquid chromatography with mass-spectrometry detection. Reverse transcriptase and protease genes were sequenced at baseline and the time of virological failure. Results: When the 38 patients started the lopinavir/ritonavir-based regimen, baseline median (25–75th percentile) values were: CD4 cell count 218 cells/μl (133–477); plasma HIV-1-RNA load 5.3 log10 copies/ml (3.8–5.1); number of lopinavir mutations four per protease gene (two to six). Univariate analysis associated virological success or failure at month 6 (21/38 patients) with the number of baseline lopinavir mutations, intracellular and plasma lopinavir Cmin, and the genotype inhibitory quotient (GIQ) at months 1 and 6. Multivariate analysis showed that the number of baseline lopinavir mutations and intracellular and plasma lopinavir Cmin were independently associated with virological success or failure. We defined the most discriminating intracellular and plasma lopinavir Cmin efficacy thresholds (8 and 4 μg/ml, respectively) and GIQ thresholds (1 and 3, respectively). Conclusion: The monitoring of lopinavir/rironavir-based HAART efficacy should include the number of baseline lopinavir/ritonavir mutations, intracellular and plasma lopinavir Cmin and GIQ calculation.

Simultaneous determination of five systemic azoles in plasma by high-performance liquid chromatography with ultraviolet detection

A simple, specific and automatable HPLC assay was developed for a simultaneous determination of systemic azoles (fluconazole, posaconazole, voriconazole, itraconazole and its metabolite hydroxyl-itraconazole, and ketoconazole) in plasma. The major advantage of this assay was sample preparation by a fully automatable solid phase extraction with Varian Plexa cartridges. C6-phenyl column was used for chromatographic separation, and UV detection was set at a wavelength of 260 nm. Linezolid was used as an internal standard. The assay was specific and linear over the concentration range of 0.05 to 40 microg/ml excepted for fluconazole which was between 0.05 and 100 microg/ml, and itraconazole between 0.1 and 40 microg/ml. Validation data for accuracy and precision for intra- and inter-day were good and satisfied FDAs guidance: CV between 0.24% and 11.66% and accuracy between 93.8% and 108.7% for all molecules. This assay was applied to therapeutic drug monitoring on patients hospitalized in intensive care and onco-hematologic units.

Alveolar concentrations of piperacillin/tazobactam administered in continuous infusion to patients with ventilator-associated pneumonia.

Objectives:To determine the steady-state serum and alveolar concentrations of piperacillin/tazobactam administered in continuous infusion to critically ill patients with ventilator-associated pneumonia and various degrees of renal failure. Design:Prospective comparative study. Setting:An intensive care unit and research ward in a university hospital. Patients:Forty patients with microbiologically documented ventilator-associated pneumonia. Interventions:Patients were randomized to receive piperacillin/tazobactam daily continuous infusions of 12/1.5 g or 16/2 g after a loading dose of 4/0.5 g. The serum and alveolar piperacillin/tazobactam concentrations were determined at steady-state with high performance liquid chromatography. Measurements and Main Results:The median (interquartile) serum and alveolar piperacillin concentrations were respectively 25.3 mg/L (23.1–32.6) and 12.7 mg/L (6.7–18.0) for 12/1.5 g/day, and 38.9 mg/L (32.9–59.6) and 19.1 mg/L (14.0–21.5), respectively, for 16/2 g/day in patients with no/mild renal failure. In patients with moderate/advance renal failure, the median (interquartile) serum and alveolar piperacillin concentrations were 102.4 mg/L (97.4–112.6) and 44.1 mg/L (33.4–48.3), respectively, for 12/1.5 g/day, and 135.3 mg/L (119.5–146.2) and 54.9 mg/L (45.2–110.3), respectively, for 16/2 g/day. Our results show great variability in piperacillin/tazobactam concentrations, with an alveolar percentage penetration of 40–50% for piperacillin and 65–85% for tazobactam and a negative association between serum or alveolar concentrations and creatinine clearance. Conclusions:A target piperacillin serum concentration of at least 35–40 mg/L is probably required to provide alveolar concentrations exceeding the susceptibility breakpoint for Gram-negative bacteria (16 mg/L) during ventilator-associated pneumonia. In patients with no/mild renal failure, a continuous daily dose of piperacillin/tazobactam 16/2 g allows reaching this target concentration, which might be not observed with 12/1.5 g/day. In patients with moderate/advanced renal failure, both dosages achieve serum concentrations far above the 35–40 mg/L threshold, suggesting that in that case, therapeutic drug monitoring should be performed in order to adjust the daily dose.

Virologic response to nelfinavir-based regimens: pharmacokinetics and drug resistance mutations (viraphar study)

ObjectiveTo assess the impact of HIV-1 protease and reverse transcriptase (RT) mutations, and pharmacokinetic parameters on virological responses to nelfinavir (NFV)-containing highly active antiretroviral therapy. DesignNaive or antiretroviral-experienced HIV-1-infected subjects were included in a non-randomized, observational cohort study and received two nucleoside RT inhibitors + NFV (750 mg three times per day or 1250 mg twice per day). Virologic success was defined as a virus load < 50 copies/ml for > 6 months. MethodsRT and protease genes were sequenced at baseline and at the time of virological failure. Plasma NFV trough concentration (Cmin), maximum concentration (Cmax), and AUC0–τ at steady-state were subjected to population pharmacokinetic analysis. ResultsPatients (n = 154) enrolled between November 1998 and February 2000 started a twice per day (n = 84) or three times per day (n = 70) NFV-based regimen as first- (n = 48) or second-line therapy when protease inhibitor-naive (n = 64) or -experienced (n = 42). Median follow-up duration was 16 months. Virologic failure occurred in 88 patients. No significant differences were observed between twice per day and three times per day regimens. According to multivariate analysis, NFV Cmin and Cmax, CD4 cell count, number of baseline RT + protease gene mutations, D67N, M184V, T215F/Y in RT, and M36I in protease, were independent factors that were significantly predictive of failure. At failure, L10I, D30N, M36I, V77I, N88S/D or L90M protease mutations had emerged since baseline. Pharmacokinetic parameters were similar in patients with or without emergence of these neo-mutations. The more discriminating NFV Cmin efficacy-threshold was estimated to be 1 mg/l. ConclusionsOur data confirm the association among individual pharmacokinetic parameters, genotype pattern and virological response to NFV-containing regimens.

HIV-1 resistance patterns to integrase inhibitors in antiretroviral-experienced patients with virological failure on raltegravir-containing regimens

BACKGROUND Our aim was to study the in vivo viral genetic pathways for resistance to raltegravir, in antiretroviral-experienced patients with virological failure (VF) on raltegravir-containing regimens. METHODS We set up a prospective study including antiretroviral-experienced patients receiving raltegravir-based regimens. Integrase (IN) genotypic resistance analysis was performed at baseline. IN was also sequenced at follow-up points in the case of VF, i.e. plasma HIV-1 RNA>400 copies/mL at month 3 and/or >50 copies/mL at month 6. For phenotyping, the IN region was recombined with an IN-deleted HXB2-based HIV-1 backbone. A titrated amount of IN recombinant viruses was used for antiviral testing against raltegravir and elvitegravir. RESULTS Among 51 patients, 11 (21.6%) had VF. Four different patterns of IN mutations were observed: (i) emergence of Q148H/R with secondary mutations (n=5 patients); (ii) emergence of N155H, then replaced by a pattern including Y143C/H/R (n=3); (iii) selection of S230N (n=1); and (iv) no evidence of selection of IN mutations (n=2). The median raltegravir and elvitegravir fold changes (FCs) were 244 (154-647) and 793 (339-892), respectively, for the Q148H/R pattern, while the median raltegravir and elvitegravir FCs were 21 (6-52) and 3 (2-3), respectively, with Y143C/H/R. The median plasma raltegravir Cmin was lower in patients with selection of the N155H mutation followed by Y143C/H/R compared with patients with Q148H/R and with patients without emerging mutations or without VF. CONCLUSIONS Diverse genetic profiles can be associated with VF on raltegravir-containing regimens, including the dynamics of replacement of mutational profiles. Pharmacokinetic parameters could be involved in this genetic evolution.

Pharmacokinetics and intrapulmonary diffusion of levofloxacin in critically ill patients with severe community-acquired pneumonia.

Objective:To determine the steady-state plasma and epithelial lining fluid concentrations of intravenous levofloxacin, 500 mg, administered once or twice daily in critically ill patients with severe community-acquired pneumonia. Design:Prospective, open-label study. Setting:An intensive care unit and a clinical pharmacokinetic laboratory in two university hospitals. Patients:Twenty-four adult patients with severe community-acquired pneumonia and receiving mechanical ventilation were enrolled. Interventions:All subjects received 1-hr intravenous infusions of 500 mg levofloxacin once or twice daily. The plasma and epithelial lining fluid levofloxacin concentrations were determined at steady-state after 2 days of therapy with high-performance liquid chromatography. Measurements and main results:The median (interquartile range [IQR]) plasma and epithelial lining fluid peak levofloxacin concentrations were 12.6 (IQR, 12.0–14.1) and 11.9 (IQR, 8.7–13.7) mg/L, respectively, in the once-daily group and 19.7 (IQR, 19.0–22.0) and 17.8 (IQR, 16.2–23.5) mg/L in the twice-daily group, showing a pulmonary percentage penetration of >100% in both groups. The median (IQR) total body exposures were 151 (IQR, 137–174) and 416 (IQR, 406–472) mg·hr/L, respectively, in the once-daily and twice-daily groups. Conclusions:Our results suggest that in critically ill patients who are receiving mechanical ventilation and have severe community-acquired pneumonia and creatinine clearance of >40 mL/min, the administration of 500 mg of intravenous levofloxacin once and twice daily allows for the exceeding of pharmacodynamic thresholds predictive of outcome (i.e., peak concentration to minimum inhibitory concentration ratio of >10 or area under concentration-time curve to minimal inhibitory concentration ratio of >125 hrs) both in serum and epithelial lining fluid for pathogens with minimum inhibitory concentration values of ≤1 mg/L and >1 mg/L, respectively.

Prediction of methotrexate elimination after high dose infusion in children with acute lymphoblastic leukaemia using a population pharmacokinetic approach.

Abstract— High‐dose methotrexate (HD‐MTX) with leucovorin rescue is a component of therapy in children with acute lymphoblastic leukaemia. Since MTX toxicity is related to drug exposure, a monitoring of serum MTX concentrations at H24, H48, H72 and until the concentration is less than 0.2 μmol/L is commonly performed. However, a number of patients may reach concentrations of less than 0.2 μmol/L long before the next sampling is scheduled. The aim of our study was to develop a Bayesian method predicting the time at which MTX concentration reaches 0.2 μmol/L in order to decrease the number of samples drawn and to allow for a more rapid patient discharge. Methotrexate population parameters were estimated from a retrospective analysis of 60 infusions in 23 children and MTX concentrations were predicted from an independent set of 20 courses in 14 children with a Bayesian approach using either one (H48) or two (H24 and H48) samples. The following population parameters were obtained using a two‐compartment model: CL = 3.51 L/h (inter‐individual variability: 66%), Vd = 8.67 L (58%), k12 = 0.0044 h−1 (105%), k21 = 0.039 h−1 (25%). Clearance and Vd were found to increase with weight and age respectively. Both sampling schedules tested for the Bayesian estimation enabled accurate prediction of concentrations and provided satisfactory precision despite a small bias. When considering the ability to predict the time at which the threshold was reached, the one‐sample (H48) schedule gave the best results. We conclude that a sampling schedule involving only one sample and Bayesian parameter estimation may be able to predict the delay necessary to reach 0.2 μmol/L in each individual.

Simultaneous determination of ritonavir and saquinavir, two human immunodeficiency virus protease inhibitors, in human serum by high-performance liquid chromatography

The aim of this study was to describe an high-performance liquid chromatographic assay for the simultaneous determination of two HIV protease inhibitors, saquinavir and ritonavir, in human serum. The method involved extraction of ritonavir and saquinavir from serum with the aid of solid-phase extraction C18 cartridges followed by high-performance liquid chromatography with a C8 column and ultraviolet detection set at a wavelength of 240 nm. The assay was linear and has been validated over the concentrations range of 0.5-32 microg/ml for ritonavir and 0.075-4.8 microg/ml for saquinavir, from 600 microl serum extracted. In future, the assay will be used to support human population pharmacokinetic studies, and therapeutic drug monitoring for ritonavir and saquinavir.