Manuel Haschke

Efficacy of Daptomycin in Implant-Associated Infection Due to Methicillin-Resistant Staphylococcus aureus: Importance of Combination with Rifampin

ABSTRACT Limited treatment options are available for implant-associated infections caused by methicillin (meticillin)-resistant Staphylococcus aureus (MRSA). We compared the activity of daptomycin (alone and with rifampin [rifampicin]) with the activities of other antimicrobial regimens against MRSA ATCC 43300 in the guinea pig foreign-body infection model. The daptomycin MIC and the minimum bactericidal concentration in logarithmic phase and stationary growth phase of MRSA were 0.625, 0.625, and 20 μg/ml, respectively. In time-kill studies, daptomycin showed rapid and concentration-dependent killing of MRSA in stationary growth phase. At concentrations above 20 μg/ml, daptomycin reduced the counts by >3 log10 CFU/ml in 2 to 4 h. In sterile cage fluid, daptomycin peak concentrations of 23.1, 46.3, and 53.7 μg/ml were reached 4 to 6 h after the administration of single intraperitoneal doses of 20, 30, and 40 mg/kg of body weight, respectively. In treatment studies, daptomycin alone reduced the planktonic MRSA counts by 0.3 log10 CFU/ml, whereas in combination with rifampin, a reduction in the counts of >6 log10 CFU/ml was observed. Vancomycin and daptomycin (at both doses) were unable to cure any cage-associated infection when they were given as monotherapy, whereas rifampin alone cured the infections in 33% of the cages. In combination with rifampin, daptomycin showed cure rates of 25% (at 20 mg/kg) and 67% (at 30 mg/kg), vancomycin showed a cure rate of 8%, linezolid showed a cure rate of 0%, and levofloxacin showed a cure rate of 58%. In addition, daptomycin at a high dose (30 mg/kg) completely prevented the emergence of rifampin resistance in planktonic and adherent MRSA cells. Daptomycin at a high dose, corresponding to 6 mg/kg in humans, in combination with rifampin showed the highest activity against planktonic and adherent MRSA. Daptomycin plus rifampin is a promising treatment option for implant-associated MRSA infections.

Linezolid Alone or Combined with Rifampin against Methicillin-Resistant Staphylococcus aureus in Experimental Foreign-Body Infection

ABSTRACT We investigated the activity of linezolid, alone and in combination with rifampin (rifampicin), against a methicillin-resistant Staphylococcus aureus (MRSA) strain in vitro and in a guinea pig model of foreign-body infection. The MIC, minimal bactericidal concentration (MBC) in logarithmic phase, and MBC in stationary growth phase were 2.5, >20, and >20 μg/ml, respectively, for linezolid; 0.01, 0.08, and 2.5 μg/ml, respectively, for rifampin; and 0.16, 0.63, >20 μg/ml, respectively, for levofloxacin. In time-kill studies, bacterial regrowth and the development of rifampin resistance were observed after 24 h with rifampin alone at 1× or 4× the MIC and were prevented by the addition of linezolid. After the administration of single intraperitoneal doses of 25, 50, and 75 mg/kg of body weight, linezolid peak concentrations of 6.8, 12.7, and 18.1 μg/ml, respectively, were achieved in sterile cage fluid at ≈3 h. The linezolid concentration remained above the MIC of the test organism for 12 h with all doses. Antimicrobial treatments of animals with cage implant infections were given twice daily for 4 days. Linezolid alone at 25, 50, and 75 mg/kg reduced the planktonic bacteria in cage fluid during treatment by 1.2 to 1.7 log10 CFU/ml; only linezolid at 75 mg/kg prevented bacterial regrowth 5 days after the end of treatment. Linezolid used in combination with rifampin (12.5 mg/kg) was more effective than linezolid used as monotherapy, reducing the planktonic bacteria by ≥3 log10 CFU (P < 0.05). Efficacy in the eradication of cage-associated infection was achieved only when linezolid was combined with rifampin, with cure rates being between 50% and 60%, whereas the levofloxacin-rifampin combination demonstrated the highest cure rate (91%) against the strain tested. The linezolid-rifampin combination is a treatment option for implant-associated infections caused by quinolone-resistant MRSA.

Pharmacokinetics and Pharmacodynamics of Recombinant Human Angiotensin-Converting Enzyme 2 in Healthy Human Subjects

Background and ObjectivesAngiotensin-converting enzyme 2 (ACE2) converts angiotensin II (Ang1-8) to angiotensin 1-7 (Ang1-7), a functional antagonist of Ang1-8, with vasodilatory, antiproliferative, antiangiogenic, and anti-inflammatory properties. In conditions with an unbalanced renin–angiotensin–aldosterone system with elevated Ang1-8, administration of ACE2 has shown promising effects in a variety of animal models. Enhancing ACE2 activity by exogenous administration of ACE2 might also be beneficial in human diseases with pathologically elevated Ang1-8. As a first step we performed a first-in-man study to determine pharmacokinetics, pharmacodynamics, safety, and tolerability of recombinant ACE2 in healthy volunteers.MethodsRecombinant human ACE2 (rhACE2) was administered intravenously to healthy human subjects in a randomized, double-blind, placebo-controlled, single-dose, dose-escalation study followed by an open-label multiple-dose study. ACE2 concentrations were determined by quantifying ACE2 activity and ACE2 content in plasma samples. Concentrations of the angiotensin system effector peptides Ang1-8, Ang1-7, and Ang1-5 were determined using a liquid chromatography–tandem mass spectrometry method.ResultsSingle rhACE2 doses of 100–1,200xa0μg/kg caused a dose-dependent increase of systemic exposure with biphasic elimination and a dose-independent terminal half-life of 10xa0h. In all single-dose cohorts, Ang1-8 decreased within 30xa0min postinfusion, angiotensin 1-7 (Ang1-7) either increased (100 and 200xa0μg/kg doses), decreased, or remained unchanged (400–1,200xa0μg/kg doses), whereas angiotensin 1-5 (Ang1-5) transiently increased for all doses investigated. With the exception of the lowest rhACE2 dose, the decrease in Ang1-8 levels lasted for at least 24xa0h. Repeated dosing (400xa0μg/kg for 3 or 6xa0days) caused only minimal accumulation of ACE2, and Ang1-8 levels were suppressed over the whole application period.ConclusionsAdministration of rhACE2 was well tolerated by healthy human subjects. Exposure was dose dependent with a dose-independent terminal elimination half-life in the range of 10xa0h. Despite marked changes in angiotensin system peptide concentrations, cardiovascular effects were absent, suggesting the presence of effective compensatory mechanisms in healthy volunteers.

Pharmacokinetics and concentration-effect relationship of oral LSD in humans

Background: The pharmacokinetics of oral lysergic acid diethylamide are unknown despite its common recreational use and renewed interest in its use in psychiatric research and practice. Methods: We characterized the pharmacokinetic profile, pharmacokinetic-pharmacodynamic relationship, and urine recovery of lysergic acid diethylamide and its main metabolite after administration of a single oral dose of lysergic acid diethylamide (200 μg) in 8 male and 8 female healthy subjects. Results: Plasma lysergic acid diethylamide concentrations were quantifiable (>0.1ng/mL) in all the subjects up to 12 hours after administration. Maximal concentrations of lysergic acid diethylamide (mean±SD: 4.5±1.4ng/mL) were reached (median, range) 1.5 (0.5–4) hours after administration. Concentrations then decreased following first-order kinetics with a half-life of 3.6±0.9 hours up to 12 hours and slower elimination thereafter with a terminal half-life of 8.9±5.9 hours. One percent of the orally administered lysergic acid diethylamide was eliminated in urine as lysergic acid diethylamide, and 13% was eliminated as 2-oxo-3-hydroxy-lysergic acid diethylamide within 24 hours. No sex differences were observed in the pharmacokinetic profiles of lysergic acid diethylamide. The acute subjective and sympathomimetic responses to lysergic acid diethylamide lasted up to 12 hours and were closely associated with the concentrations in plasma over time and exhibited no acute tolerance. Conclusions: These first data on the pharmacokinetics and concentration-effect relationship of oral lysergic acid diethylamide are relevant for further clinical studies and serve as a reference for the assessment of intoxication with lysergic acid diethylamide.

Activity of dalbavancin, alone and in combination with rifampicin, against meticillin-resistant Staphylococcus aureus in a foreign-body infection model

The activity of dalbavancin, a representative of the lipoglycopeptide antibiotics, alone and in combination with rifampicin, was investigated against meticillin-resistant Staphylococcus aureus (MRSA) in a foreign-body infection model in guinea pigs. The MIC, MBC and time-kill profile of dalbavancin were determined for MRSA ATCC 43300 in the logarithmic (MBC(log)) and stationary (MBC(stat)) growth phases. The pharmacokinetic profile of dalbavancin was determined in sterile cage fluid in guinea pigs. The activity of intraperitoneal dalbavancin (40, 60 or 80 mg/kg as a single dose), rifampicin (12.5 mg/kg/12 h for 4 days) and their combination was assessed against planktonic and biofilm MRSA. The MIC of dalbavancin was 0.078 mg/L; MBC(log) and MBC(stat) were both >128× MIC. In time-kill studies, bacterial reduction of 3log(10)CFU/mL was achieved after 48 h at ≥32× MIC (logarithmic growth) and at ≥1× MIC (stationary growth). Dalbavancin was neither synergistic nor antagonistic with rifampicin, and prevented the emergence of rifampicin resistance in vitro. The half-life of dalbavancin in cage fluid was 35.8-45.4 h and the concentration remained above the MIC of MRSA during 7 days after a single dose. Dalbavancin reduced planktonic MRSA in cage fluid at high dose (60 mg/kg and 80 mg/kg) but failed to eradicate biofilm MRSA from cages. In combination with rifampicin, dalbavancin at 80 mg/kg cured 36% of infected cages, and emergence of rifampicin resistance was completely prevented. Dalbavancin at 80 mg/kg and in combination with rifampicin eradicated approximately one-third of cage-associated MRSA infections and prevented emergence of rifampicin resistance.

The Basel Cocktail for Simultaneous Phenotyping of Human Cytochrome P450 Isoforms in Plasma, Saliva and Dried Blood Spots

Background and ObjectivePhenotyping cocktails use a combination of cytochrome P450 (CYP)-specific probe drugs to simultaneously assess the activity of different CYP isoforms. To improve the clinical applicability of CYP phenotyping, the main objectives of this study were to develop a new cocktail based on probe drugs that are widely used in clinical practice and to test whether alternative sampling methods such as collection of dried blood spots (DBS) or saliva could be used to simplify the sampling process.MethodsIn a randomized crossover study, a new combination of commercially available probe drugs (the Basel cocktail) was tested for simultaneous phenotyping of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6 and CYP3A4. Sixteen subjects received low doses of caffeine, efavirenz, losartan, omeprazole, metoprolol and midazolam in different combinations. All subjects were genotyped, and full pharmacokinetic profiles of the probe drugs and their main metabolites were determined in plasma, dried blood spots and saliva samples.ResultsThe Basel cocktail was well tolerated, and bioequivalence tests showed no evidence of mutual interactions between the probe drugs. In plasma, single timepoint metabolic ratios at 2xa0h (for CYP2C19 and CYP3A4) or at 8xa0h (for the other isoforms) after dosing showed high correlations with corresponding area under the concentration–time curve (AUC) ratios (AUC0–24hxa0parent/AUC0–24hxa0metabolite) and are proposed as simple phenotyping metrics. Metabolic ratios in dried blood spots (for CYP1A2 and CYP2C19) or in saliva samples (for CYP1A2) were comparable to plasma ratios and offer the option of minimally invasive or non-invasive phenotyping of these isoforms.ConclusionsThis new combination of phenotyping probe drugs can be used without mutual interactions. The proposed sampling timepoints have the potential to facilitate clinical application of phenotyping but require further validation in conditions of altered CYP activity. The use of DBS or saliva samples seems feasible for phenotyping of the selected CYP isoforms.

Comparison Of Liver Cell Models Using The Basel Phenotyping Cocktail

Currently used hepatocyte cell systems for in vitro assessment of drug metabolism include hepatoma cell lines and primary human hepatocyte (PHH) cultures. We investigated the suitability of the validated in vivo Basel phenotyping cocktail (caffeine [CYP1A2], efavirenz [CYP2B6], losartan [CYP2C9], omeprazole [CYP2C19], metoprolol [CYP2D6], midazolam [CYP3A4]) in vitro and characterized four hepatocyte cell systems (HepG2 cells, HepaRG cells, and primary cryopreserved human hepatocytes in 2-dimensional [2D] culture or in 3D-spheroid co-culture) regarding basal metabolism and CYP inducibility. Under non-induced conditions, all CYP activities could be determined in 3D-PHH, CYP2B6, CYP2C19, CYP2D6, and CYP3A4 in 2D-PHH and HepaRG, and CYP2C19 and CYP3A4 in HepG2 cells. The highest non-induced CYP activities were observed in 3D-PHH and HepaRG cells. mRNA expression was at least four-fold higher for all CYPs in 3D-PHH compared to the other cell systems. After treatment with 20 μM rifampicin, mRNA increased 3- to 50-fold for all CYPs except CYP1A2 and 2D6 for HepaRG and 3D-PHH, 4-fold (CYP2B6) and 17-fold (CYP3A4) for 2D-PHH and four-fold (CYP3A4) for HepG2. In 3D-PHH at least a two-fold increase in CYP activity was observed for all inducible CYP isoforms while CYP1A2 and CYP2C9 activity did not increase in 2D-PHH and HepaRG. CYP inducibility assessed in vivo using the same phenotyping probes was also best reflected by the 3D-PHH model. Our studies show that 3D-PHH and (with some limitations) HepaRG are suitable cell systems for assessing drug metabolism and CYP induction in vitro. HepG2 cells are less suited to assess CYP induction of the 2C and 3A family. The Basel phenotyping cocktail is suitable for the assessment of CYP activity and induction also in vitro.

Efficacy, safety and pharmacokinetics of 1,2,4-trioxolane OZ78 against an experimental infection with Fasciola hepatica in sheep

The synthetic peroxide OZ78 is an effective flukicide in the rodent model, but the potential of OZ78 in target animals has not been studied to date. In the present study, OZ78 was administered at 50mg/kg orally and subcutaneously to sheep harbouring an experimental Fasciola hepatica infection and the efficacy, tolerability and pharmacokinetic profiles were monitored. OZ78 given orally or subcutaneously revealed no effect neither on faecal egg counts nor on worm burdens. Apart from significant subcutaneous swelling at the injection sites of most of the treated animals, no other treatment related adverse events occurred. OZ78 had no significant effect on any haematological, coagulation or clinical chemistry variables tested. Following oral administration, a mean C(max) of 45.8±13 μg/ml was reached after 1h. An estimated elimination half-life of 1.0 h and a mean AUC of 116.2±47 μg min/ml was calculated for the oral administration. Following subcutaneous treatment with OZ78 C(max) and t(max) were 13.7±6.1μg/ml and 0.9±0.4h, respectively. The α and β half-lives were 4.5±4.3 h and 56.5±36 h, respectively and the mean AUC was 219.1±74 μg min/ml. Further studies are needed to determine whether the excellent activity observed with OZ78 in the rat model can be translated into efficacy in larger mammals.

Quantification of plasma carnitine and acylcarnitines by high-performance liquid chromatography-tandem mass spectrometry using online solid-phase extraction

AbstractCarnitine is an amino acid derivative that plays a key role in energy metabolism. Endogenous carnitine is found in its free form or esterified with acyl groups of several chain lengths. Quantification of carnitine and acylcarnitines is of particular interest for screening for research and metabolic disorders. We developed a method with online solid-phase extraction coupled to high-performance liquid chromatography and tandem mass spectrometry to quantify carnitine and three acylcarnitines with different polarity (acetylcarnitine, octanoylcarnitine, and palmitoylcarnitine). Plasma samples were deproteinized with methanol, loaded on a cation exchange trapping column and separated on a reversed-phase C8 column using heptafluorobutyric acid as an ion-pairing reagent. Considering the endogenous nature of the analytes, we quantified with the standard addition method and with external deuterated standards. Solid-phase extraction and separation were achieved within 8xa0min. Recoveries of carnitine and acylcarnitines were between 98 and 105xa0%. Both quantification methods were equally accurate (all values within 84 to 116xa0% of target concentrations) and precise (day-to-day variation of less than 18xa0%) for all carnitine species and concentrations analyzed. The method was used successfully for determination of carnitine and acylcarnitines in different human samples. In conclusion, we present a method for simultaneous quantification of carnitine and acylcarnitines with a rapid sample work-up. This approach requires small sample volumes and a short analysis time, and it can be applied for the determination of other acylcarnitines than the acylcarnitines tested. The method is useful for applications in research and clinical routine.n FigureA method is presented for the analysis of carnitine and acylcarnitines in urine which includes a precipitation step, on-column extraction and LC-MS/MS. The run time is 8 minutes and the method was validated for carnitine, acetylcarnitine, octanoylcarnitine and palmitoylcarnitine. Analysis of a patient sample with medium-chain acyl-CoA dehydrogenase deficiency is shown.

Therapeutic drug monitoring of once daily aminoglycoside dosing : comparison of two methods and investigation of the optimal blood sampling strategy

PurposeTherapeutic drug monitoring of patients receiving once daily aminoglycoside therapy can be performed using pharmacokinetic (PK) formulas or Bayesian calculations. While these methods produced comparable results, their performance has never been checked against full PK profiles. We performed a PK study in order to compare both methods and to determine the best time-points to estimate AUC0-24 and peak concentrations (Cmax).MethodsWe obtained full PK profiles in 14 patients receiving a once daily aminoglycoside therapy. PK parameters were calculated with PKSolver using non-compartmental methods. The calculated PK parameters were then compared with parameters estimated using an algorithm based on two serum concentrations (two-point method) or the software TCIWorks (Bayesian method).ResultsFor tobramycin and gentamicin, AUC0-24 and Cmax could be reliably estimated using a first serum concentration obtained at 1xa0h and a second one between 8 and 10xa0h after start of the infusion. The two-point and the Bayesian method produced similar results. For amikacin, AUC0-24 could reliably be estimated by both methods. Cmax was underestimated by 10–20xa0% by the two-point method and by up to 30xa0% with a large variation by the Bayesian method.ConclusionsThe ideal time-points for therapeutic drug monitoring of once daily administered aminoglycosides are 1xa0h after start of a 30-min infusion for the first time-point and 8–10xa0h after start of the infusion for the second time-point. Duration of the infusion and accurate registration of the time-points of blood drawing are essential for obtaining precise predictions.