Impact of nutritional factors on in vitro PK/PD modelling of polymyxin B against various strains of Acinetobacter baumannii.

The main objective of this study was to assess the effect of rich artificial cation-adjusted Mueller-Hinton broth (CAMHB) on the growth of three strains of Acinetobacter baumannii (ATCC 19606 and two clinical strains), either susceptible or resistant to polymyxin B (PMB), and on PMB bactericidal activity. A pharmacokinetic (PK)/pharmacodynamic (PD) modelling approach was used to characterize the effect of PMB in various conditions. Time-kill experiments were performed using undiluted CAMHB or CAMHB diluted to 50%, 25% and 10%, with or without Ca2+ and Mg2+ compensation (known to affect PMB activity), and with PMB concentrations ranging from 0.25 to 256 mg/L based on the strain's MIC. For each strain, time-kill replicates were modelled using NONMEM. Unexpectedly, dilution of CAMHB by up to 10-fold did not affect the growth rate of any of the three strains in the absence of PMB. However, the bactericidal activity of PMB increased with medium dilution, resulting in a reduction in the apparent bacterial regrowth of the various strains observed after a few hours. Data for each strain were well characterized by a PK/PD model, with two bacterial subpopulations with different susceptibility to PMB (more susceptible and less susceptible). The impact of medium dilution and cation compensation showed relatively high, unexplained between-strain variability. Further studies are needed to characterize the mechanism underlying the medium dilution effect.

Characterization of Pseudomonas aeruginosa resistance to ceftolozane-tazobactam due to ampC and/or ampD mutations observed during treatment using semi-mechanistic PKPD modeling.

A double ampC (AmpCG183D) and ampD (AmpDH157Y) genes mutations have been identified by whole genome sequencing in a Pseudomonas aeruginosa (PaS) that became resistant (PaR) in a patient treated by ceftolozane/tazobactam (C/T). To precisely characterize the respective contributions of these mutations on the decreased susceptibility to C/T and on the parallel increased susceptibility to imipenem (IMI), mutants were generated by homologous recombination in PAO1 reference strain (PAO1- AmpCG183D, PAO1-AmpDH157Y, PAO1-AmpCG183D/AmpDH157Y) and in PaR (PaR-AmpCPaS/AmpDPaS). Sequential time-kill curve experiments were conducted on all strains and analyzed by semi-mechanistic PKPD modeling. A PKPD model with adaptation successfully described the data, allowing discrimination between initial and time-related (adaptive resistance) effects of mutations. With PAO1 and mutant-derived strains, initial EC50 values increased by 1.4, 4.1, and 29-fold after AmpCG183D , AmpDH157Y and AmpCG183D/AmpDH157Y mutations, respectively. EC50 values were increased by 320, 12.4, and 55-fold at the end of the 2 nd experiment. EC50 of PAO1-AmpCG183D/AmpDH157Y was higher than that of single mutants at any time of the experiments. Within the PaR clinical background, reversal of AmpCG183D, and AmpDH157Y mutations led to an important decrease of EC50 value, from 80.5 mg/L to 6.77 mg/L for PaR and PaR-AmpCPaS/AmpDPaS, respectively. The effect of mutations on IMI susceptibility mainly showed that the AmpCG183D mutation prevented the emergence of adaptive resistance. The model successfully described the separate and combined effect of AmpCG183D and AmpDH157Y mutations against C/T and IMI, allowing discrimination and quantification of the initial and time-related effects of mutations. This method could be reproduced in clinical strains to decipher complex resistance mechanisms.

Pharmacokinetic and pharmacodynamic properties of polymyxin B in Escherichia coli and Klebsiella pneumoniae murine infection models.

BACKGROUND: Although polymyxin B has been in use since the late 1950s, there have been limited studies done to unravel its pharmacokinetics (PK) and pharmacodynamics (PD) index. METHODS: We determined, in neutropenic infected mice, the PK, plasma protein binding and PK/PD index best correlating with efficacy for Escherichia coli and Klebsiella pneumoniae strains. RESULTS: The pharmacokinetic profile showed non-linear PK; dose was significantly correlated with absorption rate and clearance. The inhibitory sigmoid dose-effect model for the fCmax/MIC index of E. coli fitted best, but was only modestly higher than the R2 of %fT>MIC and fAUC/MIC (R2 0.91-0.93). For K. pneumoniae the fAUC/MIC index had the best fit, which was slightly higher than the R2 of %fT>MIC and fCmax/MIC (R2 0.85-0.91). Static targets of polymyxin B fAUC/MIC were 27.5-102.6 (median 63.5) and 5.9-60.5 (median 11.6) in E. coli and in K. pneumoniae isolates, respectively. A 1 log kill effect was only reached in two E. coli isolates and one K. pneumoniae. The PTA with the standard dosing was low for isolates with MIC >0.25 mg/L. CONCLUSIONS: This study confirms that fAUC/MIC can describe the exposure-response relationship for polymyxin B. The 1 log kill effect was achieved in the minority of the isolates whereas polymyxin B PK/PD targets cannot be attained for the majority of clinical isolates with the standard dosing regimen, indicating that polymyxin B may be not effective against serious infections as monotherapy.

A Minimal Physiologically Based Pharmacokinetic Model to Characterize CNS Distribution of Metronidazole in Neuro Care ICU Patients.

Understanding antibiotic concentration-time profiles in the central nervous system (CNS) is crucial to treat severe life-threatening CNS infections, such as nosocomial ventriculitis or meningitis. Yet CNS distribution is likely to be altered in patients with brain damage and infection/inflammation. Our objective was to develop a physiologically based pharmacokinetic (PBPK) model to predict brain concentration-time profiles of antibiotics and to simulate the impact of pathophysiological changes on CNS profiles. A minimal PBPK model consisting of three physiological brain compartments was developed from metronidazole concentrations previously measured in plasma, brain extracellular fluid (ECF) and cerebrospinal fluid (CSF) of eight brain-injured patients. Volumes and blood flows were fixed to their physiological value obtained from the literature. Diffusion clearances characterizing transport across the blood-brain barrier and blood-CSF barrier were estimated from system- and drug-specific parameters and were confirmed from a Caco-2 model. The model described well unbound metronidazole pharmacokinetic profiles in plasma, ECF and CSF. Simulations showed that with metronidazole, an antibiotic with extensive CNS distribution simply governed by passive diffusion, pathophysiological alterations of membrane permeability, brain ECF volume or cerebral blood flow would have no effect on ECF or CSF pharmacokinetic profiles. This work will serve as a starting point for the development of a new PBPK model to describe the CNS distribution of antibiotics with more limited permeability for which pathophysiological conditions are expected to have a greater effect.

Population pharmacokinetic/pharmacodynamic study suggests continuous infusion of ceftaroline daily dose in ventilated critical care patients with early-onset pneumonia and augmented renal clearance.

OBJECTIVES: Ceftaroline could be suitable to treat early-onset ventilator-associated pneumonia (VAP) because of its antibacterial spectrum. However, augmented renal clearance (ARC) is frequent in ICU patients and may affect ceftaroline pharmacokinetics and efficacy. The objective of the study was to explore the impact of ARC on ceftaroline pharmacokinetics and evaluate whether the currently recommended dosing regimen (600 mg every 12 h) is appropriate to treat VAP in ICU patients. METHODS: A population pharmacokinetic model was developed using pharmacokinetic data from 18 patients with measured creatinine clearance (CLCR) ranging between 83 and 309 mL/min. Monte Carlo simulations were conducted to determine the PTA and the cumulative fraction of response (CFR) against Streptococcus pneumoniae and MRSA for five dosing regimens. Study registered at ClinicalTrials.gov (NCT03025841). RESULTS: Ceftaroline clearance increased non-linearly with CLCR, with lower concentrations and lower probability of reaching pharmacokinetic/pharmacodynamic targets when CLCR increases. For the currently recommended dosing regimen, the probability of having unbound ceftaroline concentrations above the MIC over the entire dose range is greater than 90% for MICs below 0.125 mg/L. Considering the distribution of MICs, this regimen would not be effective against MRSA infections (CFR between 21% and 67% depending on CLCR), but would be effective against S. pneumoniae infections (CFR >86%). CONCLUSIONS: The recommended dosing regimen of ceftaroline seems sufficient for covering S. pneumoniae in ICU patients with ARC, but not for MRSA. Among the dosing regimens tested it appears that a constant infusion (50 mg/h) after a loading dose of 600 mg could be more appropriate for MRSA infections.

Optimized Rhombic Experimental Dynamic Checkerboard Designs to Elucidate Pharmacodynamic Drug Interactions of Antibiotics.

PURPOSE: Quantification of pharmacodynamic interactions is key in combination therapies, yet conventional checkerboard experiments with up to 10 by 10 combinations are labor-intensive. Therefore, this study provides optimized experimental rhombic checkerboard designs to enable an efficient interaction screening with significantly reduced experimental workload. METHODS: Based on the general pharmacodynamic interaction (GPDI) model implemented in Bliss Independence, a novel rhombic 'dynamic' checkerboard design with quantification of bacteria instead of turbidity as endpoint was developed. In stochastic simulations and estimations (SSE), the precision and accuracy of interaction parameter estimations and classification rates of conventional reference designs and the newly proposed rhombic designs based on effective concentrations (EC) were compared. RESULTS: Although a conventional rich design with 20-times as many combination scenarios provided estimates of interaction parameters with higher accuracy, precision and classification rates, the optimized rhombic designs with one natural growth scenario, three monotherapy scenarios per combination partner and only four combination scenarios were still superior to conventional reduced designs with twice as many combination scenarios. Additionally, the rhombic designs were able to identify whether an interaction occurred as a shift on maximum effect or EC50 with > 98%. Overall, effective concentration-based designs were found to be superior to traditional standard concentrations, but were more challenged by strong interaction sizes exceeding their adaptive concentration ranges. CONCLUSION: The rhombic designs proposed in this study enable a reduction of resources and labor and can be a tool to streamline higher throughput in drug interaction screening.

Activation of class 1 integron integrase is promoted in the intestinal environment.

Class 1 integrons are widespread genetic elements playing a major role in the dissemination of antibiotic resistance. They allow bacteria to capture, express and exchange antibiotic resistance genes embedded within gene cassettes. Acquisition of gene cassettes is catalysed by the class 1 integron integrase, a site-specific recombinase playing a key role in the integron system. In in vitro planktonic culture, expression of intI1 is controlled by the SOS response, a regulatory network which mediates the repair of DNA damage caused by a wide range of bacterial stress, including antibiotics. However, in vitro experimental conditions are far from the real lifestyle of bacteria in natural environments such as the intestinal tract which is known to be a reservoir of integrons. In this study, we developed an in vivo model of intestinal colonization in gnotobiotic mice and used a recombination assay and quantitative real-time PCR, to investigate the induction of the SOS response and expression and activity of the class 1 integron integrase, IntI1. We found that the basal activity of IntI1 was higher in vivo than in vitro. In addition, we demonstrated that administration of a subinhibitory concentration of ciprofloxacin rapidly induced both the SOS response and intI1 expression that was correlated with an increase of the activity of IntI1. Our findings show that the gut is an environment in which the class 1 integron integrase is induced and active, and they highlight the potential role of integrons in the acquisition and/or expression of resistance genes in the gut, particularly during antibiotic therapy.

PKPD Modeling of the Inoculum Effect of Acinetobacter baumannii on Polymyxin B in vivo.

The reduction in antimicrobial activity at high bacterial counts is a microbiological phenomenon known as the inoculum effect (IE). In a previous in vitro study, a significant IE was observed for polymyxin B (PMB) against a clinical isolate of Acinetobacter baumannii, and well described by a new pharmacokinetic-pharmacodynamic model. Few in vivo studies have investigated the impact of inoculum size on survival or antibiotic efficacy. Therefore, our objective was to confirm the influence of inoculum size of this A. baumannii clinical isolate on PMB in vivo effect over time. Pharmacokinetics and pharmacodynamics of PMB after a single subcutaneous administration (1, 15 and 40 mg/kg) were studied in a neutropenic murine thigh infection model. The impact of A. baumannii inoculum size (105, 106 and 107 CFU/thigh) on PMB efficacy was also evaluated. In vivo PMB PK was well described by a two-compartment model including saturable absorption from the subcutaneous injection site and linear elimination. The previous in vitro PD model was modified to adequately describe the decrease of PMB efficacy with increased inoculum size in infected mice. The IE was modeled as a decrease of 32% in the in vivo PMB bactericidal effect when the starting inoculum increases from 105 to 107 CFU/thigh. Although not as important as previously characterized in vitro an IE was confirmed in vivo.

A physiologically based pharmacokinetic (PBPK) model exploring the blood-milk barrier in lactating species - A case study with oxytetracycline administered to dairy cows and goats.

Antibiotic excretion into milk depends on several factors such as the compound's physicochemical properties, the animal physiology, and the milk composition. The objective of this study was to develop a physiologically based pharmacokinetic (PBPK) model describing the passage of drugs into the milk of lactating species. The udder is described as a permeability limited compartment, divided into vascular, extracellular water (EW), intracellular water (IW) and milk, which was stored in alveolar and cistern compartments. The pH and ionization in each compartment and the binding to IW components and to milk fat, casein, whey protein, calcium, and magnesium were considered. Bidirectional passive diffusion across the blood-milk barrier was implemented, based on in vitro permeability studies. The model application used to predict the distribution of oxytetracycline in cow and goat milk, after different doses and routes of administration, was successful. By integrating inter-individual variability and uncertainty, the model also allowed a suitable estimation of the withdrawal periods. Further work is in progress to evaluate the predictive ability of the PBPK model for compounds with different physico-chemical properties that are potentially actively transported in order to extrapolate the excretion of xenobiotics in milk of various animal species including humans.

A New Pharmacokinetic-Pharmacodynamic Model To Characterize the Inoculum Effect of Acinetobacter baumannii on Polymyxin B In Vitro.

The inoculum effect (i.e., reduction in antimicrobial activity at large starting inoculum) is a phenomenon described for various pathogens. Given that limited data exist regarding inoculum effect of Acinetobacter baumannii, we evaluated killing of A. baumannii by polymyxin B, a last-resort antibiotic, at several starting inocula and developed a pharmacokinetic-pharmacodynamic (PKPD) model to capture this phenomenon. In vitro static time-kill experiments were performed using polymyxin B at concentrations ranging from 0.125 to 128 mg/L against a clinical A. baumannii isolate at four starting inocula from 105 to 108 CFU/mL. Samples were collected up to 30 h to quantify the viable bacterial burden and were simultaneously modeled in the NONMEM software program. The expression of polymyxin B resistance genes (lpxACD, pmrCAB, and wzc), and genetic modifications were studied by RT-qPCR and DNA sequencing experiments, respectively. The PKPD model included a single homogeneous bacterial population with adaptive resistance. Polymyxin B effect was modeled as a sigmoidal Emax model and the inoculum effect as an increase of polymyxin B EC50 with increasing starting inoculum using a power function. Polymyxin B displayed a reduced activity as the starting inoculum increased: a 20-fold increase of polymyxin B EC50 was observed between the lowest and the highest inoculum. No effects of polymyxin B and inoculum size were observed on the studied genes. The proposed PKPD model successfully described and predicted the pronounced in vitro inoculum effect of A. baumannii on polymyxin B activity. These results should be further validated using other bacteria/antibiotic combinations and in vivo models.

Lipid Nanoparticles Loaded with Farnesol or Geraniol to Enhance the Susceptibility of E. coli MCR-1 to Colistin.

Resistance to colistin, one of the antibiotics of last resort against multidrug-resistant Gram-negative bacteria, is increasingly reported. Notably, MCR plasmids discovered in 2015 have now been reported worldwide in humans. To keep this antibiotic of last resort efficient, a way to tackle this mechanism seems essential. Terpene alcohols such as farnesol have been shown to improve the efficacy of some antibiotics. However, their high lipophilicity makes them difficult to use. This problem can be solved by encapsulating them in water-dispersible lipid nanoparticles (LNPs). The aim of this study was to discover, using checkerboard tests and time-kill curve experiments, an association between colistin and farnesol or geraniol loaded in LNPs, which would improve the efficacy of colistin against E. coli and, in particular, MCR-1 transconjugants. Then, the effect of the combination on E. coli inner membrane permeabilisation was evaluated using propidium iodide (PI) uptake and compared to human red blood cells plasma membrane permeabilisation. Both terpene alcohols were able to restore the susceptibility of E. coli J53 MCR-1 to colistin with the same efficacy (Emax = 16, i.e., colistin MIC was decreased from 8 to 0.5 mg/L). However, with an EC50 of 2.69 mg/L, farnesol was more potent than geraniol (EC50 = 39.49 mg/L). Time-kill studies showed a bactericidal effect on MCR-1 transconjugant 6 h after incubation, with no regrowth up to 30 h in the presence of 1 mg/L colistin (1/8 MIC) and 60 mg/L or 200 mg/L farnesol or geraniol, respectively. Colistin alone was more potent in increasing PI uptake rate in the susceptible strain (EC50 = 0.86 ± 0.08 mg/L) than in the MCR-1 one (EC50 = 7.38 ± 0.85 mg/L). Against the MCR-1 strain, farnesol-loaded LNP at 60 mg/L enhanced the colistin-induced inner membrane permeabilization effect up to 5-fold and also increased its potency as shown by the decrease in its EC50 from 7.38 ± 0.85 mg/L to 2.69 ± 0.25 mg/L. Importantly, no hemolysis was observed for LNPs loaded with farnesol or geraniol, alone or in combination with colistin, at the concentrations showing the maximum decrease in colistin MICs. The results presented here indicate that farnesol-loaded LNPs should be studied as combination therapy with colistin to prevent the development of resistance to this antibiotic of last resort.

Clinical Pharmacokinetics of Daptomycin.

Due to the low level of resistance observed with daptomycin, this antibiotic has an important place in the treatment of severe Gram-positive infections. It is the first-in-class of the group of calcium-dependent, membrane-binding lipopeptides, and is a cyclic peptide constituted of 13 amino acids and an n-decanoyl fatty acid chain. The antibacterial action of daptomycin requires its complexation with calcium. Daptomycin is not absorbed from the gastrointestinal tract and needs to be administered parenterally. The distribution of daptomycin is limited (volume of distribution of 0.1 L/kg in healthy volunteers) due to its negative charge at physiological pH and its high binding to plasma proteins (about 90%). Its elimination is mainly renal, with about 50% of the dose excreted unchanged in the urine, justifying dosage adjustment for patients with renal insufficiency. The pharmacokinetics of daptomycin are altered under certain pathophysiological conditions, resulting in high interindividual variability. As a result, therapeutic drug monitoring of daptomycin may be of interest for certain patients, such as intensive care unit patients, patients with renal or hepatic insufficiency, dialysis patients, obese patients, or children. A target for the ratio of the area under the curve to the minimum inhibitory concentration > 666 is usually recommended for clinical efficacy, whereas in order to limit the risk of undesirable muscular effects the residual concentration should not exceed 24.3 mg/L.

Comparative pharmacokinetics of the three echinocandins in ICU patients.

BACKGROUND: We conducted a prospective study in ICU patients of two tertiary hospitals in order to determine basic pharmacokinetic (PK) parameters, associated variation and target attainment rates for anidulafungin, micafungin and caspofungin. METHODS: Serum samples from patients treated for 7 days with the standard doses of anidulafungin (N = 13), micafungin (N = 14) or caspofungin (N = 7) were analysed by validated chromatographic methods. PK parameters determined with non-compartmental analysis were correlated with demographic, laboratory and disease severity characteristics. The percentages of patients attaining drug exposures described in the summary of product characteristics (SmPC) documents and preclinical PK/PD targets for stasis were estimated. RESULTS: The median (range) AUC24 was 101.46 (54.95-274.15) mg·h/L for anidulafungin, 79.35 (28.00-149.30) mg·h/L for micafungin and 48.46 (19.44-103.69) mg·h/L for caspofungin. The interindividual variability of anidulafungin, micafungin and caspofungin AUC24 was 46%-58%, attributed mainly to variability in volume of distribution (V), clearance (CL) and in both V and CL, respectively. Significant correlations were found between anidulafungin AUC24 and BMI (rs = -0.670, P = 0.012) and liver enzymes (rs = 0.572-0.665, P = 0.013-0.041) and between caspofungin Cmin and transaminase levels (rs = -0.775 to -0.786, P = 0.036-0.041). Less than 50% of our patients attained the corresponding SmPC median AUC24s and none of the patients attained the PK/PD targets for Candida albicans and Candida parapsilosis. CONCLUSIONS: Anidulafungin exposure in ICU patients was comparable with that reported in non-ICU patients and in healthy volunteers. Micafungin exposure was comparable to that of other patients but ∼30% lower than that in healthy volunteers, whereas caspofungin exposure was rather low (∼50% lower than in healthy volunteers). Larger interindividual variability (50%-60%) was recorded in ICU patients compared with other groups for all three echinocandins.

Improved antibacterial efficiency of inhaled thiamphenicol dry powders: Mathematical modelling of in vitro dissolution kinetic and in vitro antibacterial efficacy.

Thiamphenicol (TAP) is reported to be effective against many respiratory pathogens including methicillin-resistant Staphylococcus aureus (MRSA). However, its poor solubility in water remains as one of the obstacles hindering the preparation of inhalable TAP formulations. The aim of this study was to improve the dissolution rate of TAP by micronization, and investigate whether variations in the dissolution rates of TAP would affect its in vitro antibacterial activity. Inhalable dry powders composed of TAP microcrystals (MDP) or nanocrystals (NDP) were prepared by using a wet ball milling method followed by spray drying. The morphology, solid state and in vitro dissolution of these dry powders were characterized. In vitro antibacterial activities of the inhalable TAP dry powders against a MRSA strain were evaluated. A dissolution-efficacy model relating antibacterial activity with time and dissolution rate was established via modified time-kill assays. Upon being spray dried, the volumetric mean diameters of MDP and NDP were found to be around 5 µm. Solid state analyses showed that MDP and NDP possess the same crystalline form as the raw materials. NDP exhibited faster in vitro dissolution rate as compared to MDP. The in vitro antibacterial efficiency of NDP and MDP were superior to raw TAP when the test was performed at a TAP concentration of 32 mg/L. Simulated colony forming units predictions were consistent with the result measured in the time-kill experiments with Raw TAP, MDP and NDP. This study characterized the effect of the dissolution rate of TAP dry powders on in vitro antibacterial activity against MRSA, and an enhanced antibacterial activity of TAP was observed with an increase in the dissolution rate of TAP from the dry powders at certain concentration ranges.

Pharmacokinetics of colistin after nebulization or intravenous administration of colistin methanesulphonate (Colimycin®) to cystic fibrosis patients.

OBJECTIVES: Colistin, administered as the prodrug colistin methanesulphonate (CMS), is an antibiotic frequently administered as aerosol in cystic fibrosis (CF) patient. Our aim was to assess the plasma PK of colistin in CF patients treated with CMS administered intravenously or as aerosol and to compare these results with those previously reported in healthy volunteers. METHODS: Six CF patients were included, CMS and colistin concentrations were measured in plasma, urine and sputum. Either after single intravenous administration of 2 Million International Unit (MIU) or after repeated nebulizations of 2 MIU of CMS. PK of CMS and colistin were assessed by a mixed effect modeling approach. RESULTS: Renal clearance of CMS was lower in CF patients compared to that previously reported in healthy volunteers (64.3 mL/min (RSE = 15%) vs. 103 mL/min (RSE = 8%)). However, apparent clearance of colistin was higher in CF patients compared to healthy volunteers (124 mL/min (RSE = 13%) vs. 48.7 mL/min (RSE = 15%)), resulting in reduced systemic exposure to colistin (dose normalized AUC (2 MIU) of 7.4 h.mg/L/MIU vs. 11.2 h.mg/L/MIU). After repeated nebulizations, colistin concentrations were very low in plasma (<0.21 mg/L). CONCLUSIONS: Although our study suggests a lower median dose normalized colistin plasma concentrations in CF patients compared with healthy controls, this difference was not significant and a larger study is needed to substantiate this.

Pharmacokinetics of Polymyxins in Animals.

All of the small number of studies conducted during the second half of last century to investigate the pharmacokinetics of polymyxins in animals used microbiological methods to quantify the compounds in biological fluids. Those methods generally lacked the accuracy and precision required for such investigations and, in the case of studies involving administration of colistin methanesulfonate (CMS), ongoing conversion to colistin during microbiological incubation of collected samples artifactually elevated the measured concentration of colistin. The pharmacokinetic studies reviewed in this chapter involved use of more accurate, precise and specific methods for the measurement of the relevant compounds in biological matrices. The studies have been conducted in a number of pre-clinical animal species following administration via various routes (e.g. intravenous, intrapulmonary), and have provided important insights into not only the global pharmacokinetics as viewed from plasma but also the tissue distribution and handling by key organs particularly the kidneys.

In vitro evaluation of Pseudomonas aeruginosa chronic lung infection models: Are agar and calcium-alginate beads interchangeable?

Animal models of chronic lung infection with P. aeruginosa (PA) are useful tools to improve antibiotic (ATB) treatment. Two main models based on the pulmonary instillation of PA embedded in agar or calcium-alginate beads are currently used. However, these two polymers used to prepare the beads have different properties; for example, agar is a neutral polysaccharide while alginate is anionic. We hypothesized that the effect of an ATB on PA entrapped in agar or calcium-alginate beads depends on its physicochemical properties, including charge, and concentration. To test this hypothesis, PAs were entrapped in agar or calcium-alginate beads dispersed in a growth medium containing either tobramycin (TOB), selected as a cationic ATB, or ciprofloxacin (CIP) selected as a neutral zwitterionic ATB. In vitro, time-kill curves evaluating the efficacy of ATBs over time were performed by measuring the light emitted by a bioluminescent PA for 42 h in the presence of ATB concentrations ranging from 0 to 100 times the MIC. In the presence of CIP, time-kill curves obtained with PA trapped in agar or calcium-alginate beads were comparable, whatever the CIP concentration used. In the presence of TOB, a clear difference was observed between the kill curves obtained with PA embedded in agar or calcium-alginate beads. While PA trapped within agar displayed the same susceptibility than the planktonic one, it was unresponsive to TOB for concentrations up to 1-fold MIC when trapped in calcium-alginate. At 10-fold the TOB's MIC, the luminescence emitted by PA01 in the agar beads was reduced by 95% after 40 h, whereas it returned to the same initial value for PA01 trapped in alginate-based beads. The reduction in TOB efficiency was even greater when alginate-based beads were dispersed in a mucus-simulating medium. These results show that the agar and alginate beads models can be interchangeable only for uncharged ATB, such as CIP, but not for cationic ATB, like TOB. In vitro experiments performed in this study could be a quick way to evaluate the effect of each model on a given ATB before performing animal experiments.

Semimechanistic Pharmacodynamic Modeling of Aztreonam-Avibactam Combination to Understand Its Antimicrobial Activity Against Multidrug-Resistant Gram-Negative Bacteria.

Aztreonam-avibactam (ATM-AVI) is a promising combination to treat serious infections caused by multidrug-resistant (MDR) pathogens. Three distinct mechanisms of action have been previously characterized for AVI: inhibition of ATM degradation by ß-lactamases, proper bactericidal effect, and enhancement of ATM bactericidal activity. The aim of this study was to quantify the individual contribution of each of the three AVI effects. In vitro static time-kill studies were performed on four MDR Enterobacteriaceae with different ß-lactamase profiles. ß-Lactamase activity was characterized by measuring ATM concentrations over 27 hours. Data were analyzed by a semimechanistic pharmacodynamics modeling approach. Surprisingly, even though AVI prevented ATM degradation, the combined bactericidal activity was mostly explained by the enhancement of ATM effect within clinical range of ATM (5-125 mg/L) and AVI concentrations (0.9-22.5 mg/L). Therefore, when selecting a ß-lactamase inhibitor for combination with a ß-lactam, its capability to enhance the ß-lactam activity should be considered in addition to the spectrum of ß-lactamases inhibited.

Reassessing the dosing of cefoxitin prophylaxis during major abdominal surgery: insights from microdialysis and population pharmacokinetic modelling.

OBJECTIVES: Cefoxitin is frequently used for surgical antibiotic prophylaxis (SAP). Using microdialysis, we evaluated whether the currently recommended dosing regimen is appropriate to maintain cefoxitin subcutaneous tissue concentrations above the MIC for pathogens involved in abdominal surgical site infection. METHODS: Data from eight patients undergoing major abdominal surgery were analysed using population pharmacokinetic modelling, and Monte Carlo simulations were conducted to determine the PTA for aerobic and anaerobic pathogens. ClinicalTrials.gov: NCT02703857. RESULTS: Only 2.3% and 47.4% of the simulated patients maintained cefoxitin subcutaneous concentrations above the MIC breakpoint for anaerobic (MIC = 16 mg/L) and aerobic (MIC = 8 mg/L) pathogens, respectively. New simulations with administration of a loading dose followed by a constant infusion of cefoxitin were conducted and demonstrate that, notwithstanding using the same total dose per unit of time, continuous infusion of cefoxitin can cover aerobes in 96.6% of the simulated patients, but remains insufficient for anaerobic bacteria. CONCLUSIONS: The recommended dosing regimen of cefoxitin is insufficient for covering the usual bacteria during abdominal surgery. Administration of a loading dose followed by a constant infusion should be considered for aerobic bacteria and cefoxitin should be avoided as SAP for anaerobic bacteria.

Preclinical Pharmacokinetic and Pharmacodynamic Data To Support Cefoxitin Nebulization for the Treatment of Mycobacterium abscessus.

Mycobacterium abscessus is responsible for difficult-to-treat chronic pulmonary infections in humans. Current regimens, including parenteral administrations of cefoxitin (FOX) in combination with amikacin and clarithromycin, raise compliance problems and are frequently associated with high failure and development of resistance. Aerosol delivery of FOX could be an interesting alternative. FOX was administered to healthy rats by intravenous bolus or intratracheal nebulization, and concentrations were determined in plasma and epithelial lining fluid (ELF) by liquid chromatography-tandem mass spectrometry. After intrapulmonary administration, the FOX area under the curve within ELF was 1,147 times higher than that in plasma, indicating that this route of administration offers a biopharmaceutical advantage over intravenous administration. FOX antimicrobial activity was investigated using time-kill curves combined with a pharmacokinetic/pharmacodynamic (PK/PD) type modeling approach in order to account for its in vitro instability that precludes precise determination of MIC. Time-kill data were adequately described by a model including in vitro degradation, a sensitive (S) and a resistant (R) bacteria subpopulation, logistic growth, and a maximal inhibition-type growth inhibition effect of FOX. Median inhibitory concentrations were estimated at 16.2 and 252 mg/liter for the S and R subpopulations, respectively. These findings suggest that parenteral FOX dosing regimens used in patients for the treatment of M. abscessus are not sufficient to reduce the bacterial burden and that FOX nebulization offers a potential advantage that needs to be further investigated.