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Mucormycosis is an emerging invasive fungal infection, primarily affecting immunocompromised patients. The disease is difficult to diagnose and mortality reaches 40% even if treated adequately. Depending on site of infection and risk factors, surgical debridement in combination with systemically active antifungal drugs are the mainstay treatment strategies. Lipid-based amphotericin B is the treatment of choice for first-line therapy while posaconazole may be a promising alternative. We performed a PubMed search on reports of patients with mucormycosis treated with posaconazole. From 2003 to 2011, 96 cases have been published. Diagnosis was based on histology alone in 2 (2.1%) and microbiological evidence in 67 (69.8%), while no data on the diagnostic approach was reported in 27 (28.1%) patients. The most frequent pathogens were Rhizopus spp. (31.2%), followed by Mucor spp. (14.6%). The site of infection was predominantly rhino-orbital (38.5%, of which 43% also had central nervous system [CNS] involvement), followed by disseminated disease (22.1%). A complete response was achieved in 62 (64.6%), partial response in 7 (7.3%) patients, and stable disease in 1 (1%). Overall mortality was 24% (lacking data for three patients). In published case reports on posaconazole treatment for mucormycosis, the drug was frequently and successfully used in combination or as second line therapy.

Caspofungin (CAS) is approved for second-line management of proven or probable invasive aspergillosis at a dose of 50 mg once daily (QD). Preclinical and limited clinical data support the concept of the dose-dependent antifungal efficacy of CAS with preservation of its favorable safety profile. Little is known, however, about the pharmacokinetics (PKs) of higher doses of CAS in patients. In a formal multicenter phase II dose-escalation study, CAS was administered as a 2-h infusion at doses ranging from 70 to 200 mg QD. CAS PK sampling (n = 468 samples) was performed on day 1 and at peak and trough time points on days 4, 7, 14, and 28 (70 mg, n = 9 patients; 100 mg, n = 8 patients; 150 mg, n = 9 patients; 200 mg, n = 20 patients; total, n = 46 patients). Drug concentrations in plasma were measured by liquid chromatography tandem mass spectroscopy. Population pharmacokinetic analysis (PopPK) was performed using NONMEM (version 7) software. Model evaluation was performed using bootstrap analysis, prediction-corrected visual predictive check (pcVPC), as well as standardized visual predictive check (SVPC). The four investigated dose levels showed no difference in log-transformed dose-normalized trough levels of CAS (analysis of variance). CAS concentration data fitted best to a two-compartment model with a proportional-error model, interindividual variability (IIV) fitted best on clearance (CL), central and peripheral volume of distribution (V(1) and V(2), respectively) covariance fitted best on CL and V(1), interoccasion variability (IOV) fitted best on CL, and body weight fitted best as a covariate on CL and V(1) (CL, 0.411 liters/h ± 29% IIV; IOV on CL, 16%; V(1), 5.785 liters ± 29% IIV; intercompartmental clearance, 0.843 liters/h; V2, 6.53 liters ± 67% IIV). None of the other examined covariates (dose level, gender, age, serum bilirubin concentration, creatinine clearance) improved the model further. Bootstrap results showed the robustness of the final PopPK model. pcVPC and SVPC showed the predictability of the model and further confirmed the linear PKs of CAS over the dosage range of 70 to 200 mg QD. On the basis of the final model, geometric mean simulated peak plasma levels at steady state ranged from 13.8 to 39.4 mg/liter (geometric coefficient of variation, 31%), geometric mean trough levels ranged from 4.2 to 12.0 mg/liter (49%), and geometric mean areas under the concentration-time curves ranged from 170 to 487 mg · h/liter (34%) for the dosage range of 70 to 200 mg QD. CAS showed linear PKs across the investigated dosage range of 70 to 200 mg QD. Drug exposure in the present study population was comparable to that in other populations. (This study has been registered with the European Union Drug Regulating Authorities Clinical Trials website under registration no. 2006-001936-30 and at ClinicalTrials.gov under registration no. NCT00404092.).

Pharmacokinetic studies usually focus on serum concentrations, hence little is known about intracellular drug concentrations. In this study, we determined the concentration of anidulafungin within different cellular compartments of the peripheral blood. Blood samples (n=17) from patients receiving anidulafungin were collected and separated by double-discontinuous Ficoll-Hypaque density gradient centrifugation. Intracellular concentrations within the obtained cells, i.e. peripheral blood mononuclear cells (PBMCs), polymorphonuclear leucocytes (PMNs) and red blood cells (RBCs), were determined by liquid chromatography tandem mass spectrometry (LC-MS/MS). Within the PBMCs and PMNs, the intracellular concentration of anidulafungin was significantly increased compared with the plasma concentration (P<0.001). However, the concentration within RBCs did not significantly differ from the plasma concentration. Anidulafungin reaches high concentrations in human PBMCs and PMNs and is present in RBCs. In vitro data showed that intracellular uptake of anidulafungin by PBMCs depends on the albumin concentration of the surrounding medium, but only at low, i.e. non-physiological, protein concentrations., (Copyright © 2013 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for simultaneous quantification of mycophenolic acid (MPA) and its phenolic glucuronide (MPAG) in saliva and plasma. Saliva was collected and processed using a standardized commercially available collection device. Sample preparation comprised of protein precipitation with acetonitrile and subsequent centrifugation, followed by evaporation and reconstitution with mobile phase. A labeled isotope of MPA was used as internal standard, and chromatographic separation was achieved on a C18 column with an isocratic flow. LC-MS/MS detection was performed using a triple-stage quadrupole mass spectrometer working in selected reaction monitoring mode with positive electrospray ionization (ESI). The analytes were quantified in a single run within 2 min. For saliva, linearity was demonstrated over the concentration range of 5.0 to 400.0 ng/ml for both MPA and MPAG, and from 0.08 to 20.00 μg/ml for MPA and 0.4 to 100.0 μg/ml for MPAG in plasma. The lower limits of quantification for MPA and MPAG were 0.07 and 0.80 ng/ml in saliva, and 0.002 and 0.009 μg/ml in plasma, respectively. Inter- and intra-day precisions expressed as relative standard deviation (RSD) and accuracies were less than 15%. The recoveries for MPA and MPAG from the collection device's swab were higher than 90%. Sample stability was confirmed for bench times up to 24 h at room temperature. The method was validated according to International Conference on Harmonization (ICH) guidelines Q2 (R1) Validation of Analytical Procedures. The applicability of the method was tested in a renal pediatric patient. Based on a limited sampling strategy, MPA saliva and plasma concentrations were found in good agreement with each other. We suggest that the described method is suitable to analyze saliva and plasma samples of small volumes for therapeutic drug monitoring (TDM) and pharmacokinetic studies in pediatric patients., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Whilst micafungin serum concentrations are well studied, little is known about its concentrations within cellular compartments of the peripheral blood. Hence, in this study blood samples were collected from patients receiving micafungin (n=26). These samples were separated by double-discontinuous Ficoll-Hypaque density gradient centrifugation. Intracellular concentrations within the obtained cells, i.e. peripheral blood mononuclear cells (PBMCs), polymorphonuclear leukocytes (PMNs) and red blood cells (RBCs), were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Within PBMCs and PMNs, the intracellular micafungin concentration was significantly increased compared with the concentration in plasma (P<0.001). The intracellular concentration within RBCs did not significantly differ from the plasma concentration. Micafungin reaches high concentrations in human PBMCs and PMNs and is present in RBCs. In vitro data showed that intracellular uptake of micafungin by PBMCs depends on the albumin concentration of the surrounding medium, but only at non-physiological protein concentrations., (Copyright © 2011 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

Despite the availability of newer antifungal agents, invasive fungal diseases remain a leading cause of morbidity and mortality in immunocompromised patients. Voriconazole and posaconazole are two extended-spectrum triazoles indicated for treatment and prophylaxis of invasive fungal diseases. Recently, there has been increased interest in the utility of therapeutic drug monitoring to optimize safety and efficacy of antifungals in an attempt to improve patient outcomes. We reviewed the pharmacokinetic and pharmacodynamic characteristics of voriconazole and posaconazole in the context of clinical indications for therapeutic drug monitoring. In addition, the most recent evidence examining the relationship between serum concentrations of voriconazole and posaconazole and their efficacy or toxicities was evaluated. This information was then integrated to formulate recommendations for use of therapeutic drug monitoring in clinical settings.

Therapeutic drug monitoring (TDM) of antifungal plasma concentrations is increasingly recommended. However, data on antifungal concentrations in the other compartments of the peripheral blood are limited. Hence, we collected 23 blood samples from 14 patients receiving posaconazole for prophylaxis of fungal infections. These samples were separated by double-discontinuous Ficoll-Hypaque density gradient centrifugation. The intracellular posaconazole concentrations of the obtained cells, i.e., the peripheral blood mononuclear cells (PBMCs), polymorphonuclear leukocytes (PMNs), and red blood cells (RBCs), were determined by liquid chromatography-tandem mass spectrometry. The intracellular concentrations of the PBMCs and PMNs were significantly higher than those of surrounding media (P < 0.001). The ratios between the intracellular and extracellular concentrations (C/E) were 22.5 +/- 21.2, 7.66 +/- 6.50, and 0.09 +/- 0.05 for the PBMCs, PMNs, and RBCs, respectively. Posaconazole reaches high concentrations within human PBMCs and PMNs and is, to a lesser extent, present in RBCs. The high intracellular concentrations might contribute to posaconazole efficacy and distribution.

A rapid turnaround is a prerequisite of therapeutic drug monitoring (TDM). For antifungals, this need is still unmet, since hardly any method has been established to simultaneously quantitate concentrations of different antifungal classes. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed allowing quantitation of anidulafungin (ANF), caspofungin (CSF), isavuconazole (ISC), micafungin (MCF), posaconazole (PSC), and voriconazole (VRC). Quantitation was successful with diluted plasma samples, peripheral blood mononuclear cells (PBMC), polymorphonuclear leukocytes (PMN), and erythrocytes (RBC). A triple quadrupole mass spectrometer in selected reaction monitoring mode was used with positive electrospray ionization. Cells and calibration standards were extracted with acetonitrile containing internal standard. Internal standards were a CSF derivate for echinocandins and itraconazole for triazoles. Chromatographic separation of the supernatant was achieved by a gradient method facilitating a BetaBasic C4 column. Analytes were quantified in a single 8-min run. Calibration curves were linear and fitted using least squares with a weighting factor of the reciprocal concentration. Limits of detection (ng/ml) were ANF, 8.3; CSF, 31.5; ISC, 1.5; MCF, 97.7; PSC, 3.3; and VRC, 1.4. The lower limits of quantitation (ng/ml) were ANF, 64; CSF, 108; ISC, 4.5; MCF, 160; PSC, 10; and VRC, 4.2. Intraday precisions ranged from 6.3% to 8.8% for azoles and 8.8% to 15.4% for echinocandins. Intraday and interday accuracies (percent bias) of all analytes were within 13.8%. The method was established as standard practice for the quantitation of intracellular antifungal concentrations and optimizes TDM by applying a rapid single method for 6 antifungals.

Anidulafungin is the most recently approved compound of the echinocandin antifungal class. Its mode of action is the noncompetitive inhibition of β-(1,3)-D-glucan synthesis. Potent fungicidal activity has been demonstrated against many Candida spp., including non-albicansCandida spp. and fluconazole-resistant strains, as well as fungistatic activity against Aspergillus spp. Owing to low oral bioavailability, it can only be administered intravenously. Anidulafungin is not metabolized by the liver and renal clearance is negligible, thus rendering dosage adjustments in patients with impaired hepatic or renal function unnecessary. Due to lack of interference with the cytochrome P450 pathway, it displays minimal drug-drug interaction. Anidulafungin has been approved by the US FDA for the treatment of esophageal and invasive candidiasis after clinical trials demonstrated its noninferiority to fluconazole. In September 2007, anidulafungin gained EMEA approval for the treatment of invasive candidiasis in adult non-neutropenic patients. For those with invasive or noninvasive candidiasis with resistance or intolerance to fluconazole in particular, as well as those requiring antifungal medication, that anidulafungin does not interact with concomitant medication means it may be regarded as a safe and efficacious treatment option. Promising results from animal models and experience with the other echinocandins indicate several potential lines of investigation: invasive aspergillosis, prophylaxis and treatment of transplant patients, and empirical treatment in patients with febrile neutropenia. Significant differences in clinical efficacy or safety favoring anidulafungin over the other echinocandins are yet to be discovered.

Posaconazole is a new drug in the triazole class that has recently been investigated in pivotal Phase III clinical trials. Its antifungal activity is based on the inhibition of the fungal ergosterol synthesis. As demonstrated in vitro, posaconazole exhibits fungicidal activity against Aspergillus spp., Candida spp. and zygomycetes. Currently, posaconazole is only available as an oral suspension. Food consumption affects the bioavailability of posaconazole, while the exposure to posaconazole increases in a dose-proportional manner with a saturation of absorption occurring with a daily dose over 800 mg. Posaconazole is well tolerated without an increase in risk of any treatment-related adverse events during prolonged treatment for 6 or more months (n = 108). Posaconazole has been recently approved by the US FDA and other regulatory bodies for the treatment of oropharyngeal candidiasis, and the prophylaxis of invasive Aspergillus and Candida infections in severely immunocompromised patients. As demonstrated in two pivotal Phase III trials, posaconazole prophylaxis of invasive fungal infection in patients severely immunocompromised by graft-versus-host disease (n = 600) or neutropenia (n = 602) is superior to fluconazole and/or itraconazole prophylaxis. Significantly more patients who received posaconazole, instead of fluconazole, as treatment for oropharyngeal candidiasis sustained clinical success after the treatment was stopped. Preliminary data from a subgroup analysis (n = 24) of two salvage therapy trials for invasive fungal infections, as well as single case reports and series and in vitro studies, suggest that posaconazole might be an attractive oral treatment alternative for zygomycosis.