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Objectives: Temocillin is a β-lactam antibiotic used for preventing or treating bacterial infections in liver-transplanted children. We characterized its pharmacokinetics in plasma and ascitic fluid and proposed dosing regimens that maximize the achievement of effective drug exposures in this patient group., Methods: Patients aged 6-36 months received 25 mg/kg/12 h (n = 14) or 25 mg/kg/8 h (n = 23). Total and unbound temocillin concentrations were measured in plasma and ascitic fluid. Drug safety was monitored. Non-compartmental and population pharmacokinetic analyses were performed, together with Monte Carlo simulations., Results: No safety concerns were reported. For 25 mg/kg/12 h, the unbound mean (±standard deviation) C max and C min were 38 ± 16 and 2 ± 1 mg/L, respectively. For the 25 mg/kg/8 h dose, the unbound C max remained similar although the mean C min increased to 5 ± 3 mg/L. Protein binding was saturable. Median penetration in ascitic fluid from plasma was 82% (min-max: 63-95%). A three-compartment model with first-order elimination best described unbound pharmacokinetic profiles in plasma and ascitic fluid, with body weight and estimated glomerular filtration rate (GFR) as significant covariates. Monte Carlo simulations suggested that 90% probability of target attainment was achieved in both fluids with 25 mg/kg/12 h for MICs ≤4 mg/L, estimated GFR ≤180 mL/min/1.73 m 2 or weight ≥6 kg, and with 25 mg/kg/8 h, for MICs ≤8 mg/L, GFR ≤120 mL/min/1.73 m 2 or weight ≥11 kg., Discussion: Although adequate in many instances, the current dosing regimen is likely inadequate for patients with low body weight, high renal function, or bacteria with high MIC, emphasizing the need for patient-specific factors to be considered in dose selection. These data support the importance of paediatric pharmacokinetic studies to optimize drug dosing regimens., (Copyright © 2024 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Background: Biofilm-associated pulmonary infections pose therapeutic challenges in cystic fibrosis patients, especially when involving multiple bacterial species. Enzymatic degradation of the biofilm matrix may offer a potential solution to enhance antibiotic efficacy. This study investigated the repurposing of DNase I, commonly used for its mucolytic activity in cystic fibrosis, to target extracellular DNA within biofilms, as well as potential synergies with alginate lyase and broad-spectrum antibiotics in dual-species biofilms of Pseudomonas aeruginosa and Staphylococcus aureus., Methods: Dual-species biofilms were grown in artificial sputum medium using S. aureus and P. aeruginosa isolated by pairs from the same patients and exposed to various combinations of enzymes, meropenem, or tobramycin. Activity was assessed by measuring biofilm biomass and viable counts. Matrix degradation and decrease in bacterial load were visualized using confocal microscopy. Biofilm viscoelasticity was estimated by rheology., Results: Nearly complete destruction of the biofilms was achieved only if combining the enzymatic cocktail with the two antibiotics, and if using supratherapeutic levels of DNase I and high concentrations of alginate lyase. Biofilms containing non-pigmented mucoid P. aeruginosa required higher antibiotic concentrations, despite low viscoelasticity. In contrast, for biofilms with pigmented mucoid P. aeruginosa, a correlation was observed between the efficacy of different treatments and the reduction they caused in elasticity and viscosity of the biofilm., Conclusions: In this complex, highly drug-tolerant biofilm model, enzymes prove useful adjuvants to enhance antibiotic activity. However, the necessity for high enzyme concentrations emphasizes the need for thorough concentration-response evaluations and safety assessments before considering clinical applications., Competing Interests: Declaration of competing interest The authors have no conflict of interest to declare., (Copyright © 2024 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.)

Background: In lung transplant patients, direct oral anticoagulants are often taken in combination with immunosuppressive drugs such as tacrolimus. Since tacrolimus is a substrate and inhibitor of the efflux protein ABCB1, also transporting direct oral anticoagulants, a possible drug-drug interaction mediated by competition for this transporter needs to be investigated., Objectives: To determine the in vitro effect of tacrolimus on ABCB1-mediated rivaroxaban transport in order to support clinician practice., Methods: Recombinant cell line models, based on human embryonic kidney 293 cells, were generated by a stable transfection process to overexpress ABCB1 or not (control cells). The impact of tacrolimus on ABCB1-mediated rivaroxaban transport was assessed by accumulation experiments., Results: ABCB1 expression decreased the cellular accumulation of rivaroxaban and tacrolimus at their respective clinically relevant concentrations when compared with control cells. This confirms the involvement of ABCB1 in the active transport of tacrolimus and rivaroxaban. However, tacrolimus had no significant influence on rivaroxaban disposition at those clinically relevant concentrations., Conclusion: Our study does not provide evidence for a possible interaction between tacrolimus and rivaroxaban when used together in practice., (© 2024 The Authors.)

Background: Increasing multidrug resistance rates in Neisseria gonorrhoeae have raised concerns and an urgent call for new antibiotics for treatment of gonorrhoea. Several decades of subdued drug development in this field and the recent failures of two new antibiotics to show non-inferiority compared with the current first-line antibiotics ceftriaxone plus azithromycin highlight the need for improved preclinical tools to predict clinical outcome of new drugs in the development process., Objectives: To summarize current pharmacokinetic/pharmacodynamic (PK/PD) knowledge and dose-finding strategies for antibiotics against gonorrhoea., Sources: Literature review of published papers and discussions by global experts at a special workshop on this topic., Content: We review current knowledge of gonococcal specific PK/PD principles and provide an update on new in vitro and in vivo models to correlate drug exposure with clinical outcome, and identify challenges and gaps in gonococcal therapeutic research., Implications: Identifying the ideal antimicrobial agent and dose for treating uncomplicated urogenital and pharyngeal gonococcal disease requires appropriate validated non-clinical PK/PD models. Recent advances in adapting in vitro and in vivo models for use in gonorrhoea are an important step for enabling the development of new drugs with reduced risk of failure in Phase 3 clinical development and diminish the risk of emergence of resistance., (Copyright © 2020. Published by Elsevier Ltd.)

Objectives: Burkholderia pseudomallei, Yersinia pestis and Francisella tularensis are facultative intracellular bacteria causing life-threatening infections. We have (a) compared the activity of finafloxacin (a fluoroquinolone in development showing improved activity at acidic pH) with that of ciprofloxacin, levofloxacin and imipenem against the extracellular and intracellular (THP-1 monocytes) forms of infection by attenuated surrogates of these species (B. thailandensis, Y. pseudotuberculosis, F. philomiragia) and (b) assessed finafloxacin cellular pharmacokinetics (accumulation, distribution, efflux)., Methods: Bacteria in broth or in infected monocytes were exposed to antibiotics at pH 7.4 or 5.5 for 24 hr. Maximal relative efficacies (E max ) and static concentrations (C s ) were calculated using the Hill equation (concentration-response curves). Finafloxacin pharmacokinetics in cells at pH 7.4 or 5.5 was investigated using 14 C-labelled drug., Results: Extracellularly, all drugs sterilized the cultures, with finafloxacin being two to six times more potent at acidic pH. Intracellularly, E max reached the limit of detection (4-5 log 10  cfu decrease) for finafloxacin against all species, but only against B. thailandensis and F. philomiragia for ciprofloxacin and levofloxacin, while imipenem caused less than 2 log 10  cfu decrease for all species. At acid pH, C s shifted to two to five times lower values for finafloxacin and to one to four times higher values for the other drugs. Finafloxacin accumulated in THP-1 cells by approximately fivefold at pH 7.4 but up to 20-fold at pH 5.5, and distributed in the cytosol., Conclusions: Fluoroquinolones have proven to be effective in reducing the intracellular reservoirs of B. thailandensis, Y. pseudotuberculosis and F. philomiragia, with finafloxacin demonstrating an additional advantage in acidic environments., (Copyright © 2019 European Society of Clinical Microbiology and Infectious Diseases. All rights reserved.)

Long-term exposure to pharmacological agents can select for cells that overexpress efflux transporters. We previously showed that mouse J774 macrophages cultivated for a prolonged period of time with toxic concentrations of the fluoroquinolone ciprofloxacin overexpress the efflux transporter Mrp4 and display a reduced accumulation of this antibiotic, but no change in the accumulation of moxifloxacin, a closely related molecule (Antimicrob. Agents Chemother. [2006] 50, 1689-1695 and [2009] 53, 2410-2416). Because of this striking difference between the two fluoroquinolones, we have now examined the modifications in the expression of ABC efflux transporters induced by the prolonged exposure of J774 macrophages to high concentrations of moxifloxacin. The resulting cell line showed (i) no difference in the accumulation of moxifloxacin but an increased accumulation and decreased efflux of ciprofloxacin; (ii) an overexpression of the multidrug transporters Abcb1a (P-gp), Abcc2 (Mrp2) and Abcg2 (Bcrp1), and a decreased expression of Abcc4 (Mrp4). While P-gp and Bcrp1 were functional, they did not modify the cellular accumulation of fluoroquinolones. The data show that exposing cells to high concentrations of a drug that is not affected by active efflux can trigger a pleiotropic response leading to a modulation in the expression of several transporters. These changes, however, are not sufficient to protect cells against the toxicity that fluoroquinolones may exert at large concentrations. They could also cause unanticipated drug interactions in vivo, should the drug exposure grossly exceed what is anticipated from its current registered use., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

The rational selection of antibiotics for the treatment of meningitis must take into account several criteria, among which their intrinsic activity against the causative bacteria, and their pharmacokinetic and pharmacodynamic properties. The intrinsic activity is evaluated by the Minimal Inhibitory Concentration (MIC), which, however, does not give any information on the bactericidal potency of the drug (important property for infections localized in compartments with low immune defense such as the CSF). The capacity of the antibiotic to reach the infected compartment depends on its physicochemical properties (molecular weight, lipophilicity) and its protein binding capacity, but also on the properties of the blood-CSF barrier (permeability modulated by inflammation and activity of active transporters). Pharmacodynamics correlate intrinsic activity to pharmacokinetics by determining the optimal value of the ratio between MIC and time of exposure, area under the curve, or peak concentration. On these bases, beta-lactams appear as first-line antibiotics, if used with large and repeated doses (or even as a continuous infusion), because of their time-dependent activity. The choice of the molecule is based on the susceptibility of the bacterium. Potential alternatives include chloramphenicol (limited however by its toxicity), moxifloxacin (showing high bactericidal effect, a low MIC, and appropriate penetration) but little clinically documented, linezolid and vancomycin for Methicillin-Resistant Staphylococcus aureus (MRSA), and vancomycin for penicillin non-susceptible pneumococci. Other molecules in clinical development are being evaluated for this indication.

Decreased susceptibility of Staphylococcus aureus to antistaphylococcal agents may be associated with inability to eradicate intracellular forms, which could explain therapeutic failures. This hypothesis was tested using clinical isolates obtained from a patient with persistent staphylococcal bacteraemia under therapy. Four isogenic isolates (three from tissue, one from blood) with increased MICs for vancomycin (1-4 mg/L) and for daptomycin (1-4 mg/L) were collected after an initial 16-day treatment with vancomycin-rifampicin-gentamicin, followed by 13-20 days of treatment with daptomycin-rifampicin-gentamicin. Isolates were tested for MICs and for: (i) vancomycin (BODIPY-FL-vancomycin) and daptomycin binding; (ii) cell wall turnover (loss of N-acetyl-d-[1-(14)C]glucosamine in 30 min after 1 h of labelling); and (iii) Triton X-100-induced autolysis. Extracellular (broth) and intracellular (THP-1 macrophages) activities of rifampicin, linezolid and fusidic acid at C(max), and of vancomycin, daptomycin, quinupristin-dalfopristin and oritavancin over a wide range of extracellular concentrations (with pharmacological modelling to determine E(max)), were measured at 24 h. Increases in vancomycin MICs correlated with increased drug binding, and decreased cell wall turnover and detergent-induced autolysis. Increases in daptomycin MICs correlated with decreased daptomycin binding. Intracellular activity was weak (E(max) <1 log(10) CFU decrease) for vancomycin against all isolates, and for daptomycin against isolates with MICs >1 mg/L. Among all antibiotics tested, only quinupristin-dalfopristin and oritavancin provided close to bactericidal intracellular activities (1.6-2.5 log(10) CFU decreases at C(max)). Determination of the intracellular susceptibility of S. aureus, combined with improved methods of diagnosis, could be useful when dealing with persistent staphylococcal infections and could improve therapy.

Pseudomonas aeruginosa is a major cause of nosocomial infections. This organism shows a remarkable capacity to resist antibiotics, either intrinsically (because of constitutive expression of beta-lactamases and efflux pumps, combined with low permeability of the outer-membrane) or following acquisition of resistance genes (e.g., genes for beta-lactamases, or enzymes inactivating aminoglycosides or modifying their target), over-expression of efflux pumps, decreased expression of porins, or mutations in quinolone targets. Worryingly, these mechanisms are often present simultaneously, thereby conferring multiresistant phenotypes. Susceptibility testing is therefore crucial in clinical practice. Empirical treatment usually involves combination therapy, selected on the basis of known local epidemiology (usually a beta-lactam plus an aminoglycoside or a fluoroquinolone). However, therapy should be simplified as soon as possible, based on susceptibility data and the patient's clinical evolution. Alternative drugs (e.g., colistin) have proven useful against multiresistant strains, but innovative therapeutic options for the future remain scarce, while attempts to develop vaccines have been unsuccessful to date. Among broad-spectrum antibiotics in development, ceftobiprole, sitafloxacin and doripenem show interesting in-vitro activity, although the first two molecules have been evaluated in clinics only against Gram-positive organisms. Doripenem has received a fast track designation from the US Food and Drug Administration for the treatment of nosocomial pneumonia. Pump inhibitors are undergoing phase I trials in cystic fibrosis patients. Therefore, selecting appropriate antibiotics and optimising their use on the basis of pharmacodynamic concepts currently remains the best way of coping with pseudomonal infections.

Quinolones are one of the largest classes of antimicrobial agents used worldwide. This review considers the quinolones that are available currently and used widely in Europe (norfoxacin, ciprofloxacin, ofloxacin, levofloxacin and moxifloxacin) within their historical perspective, while trying to position them in the context of recent and possible future advances based on an understanding of: (1) their chemical structures and how these impact on activity and toxicity; (2) resistance mechanisms (mutations in target genes, efflux pumps); (3) their pharmacodynamic properties (AUC/MIC and Cmax/MIC ratios; mutant prevention concentration and mutant selection window); and (4) epidemiological considerations (risk of emergence of resistance, clonal spread). Their main indications are examined in relation to their advantages and drawbacks. Overall, it is concluded that these important agents should be used in an educated fashion, based on a careful balance between their ease of use and efficacy vs. the risk of emerging resistance and toxicity. However, there is now substantial evidence to support use of the most potent drug at the appropriate dose whenever this is required.

The dicationic macrolide antibiotic azithromycin inhibits the uptake of horseradish peroxidase (HRP) by fluid-phase pinocytosis in fibroblasts in a time- and concentration-dependent fashion without affecting its decay (regurgitation and/or degradation). The azithromycin effect is additive to that of nocodazole, known to impair endocytic uptake and transport of solutes along the endocytic pathway. Cytochemistry (light and electron microscopy) shows a major reduction by azithromycin in the number of HRP-labeled endocytic vesicles at 5 min (endosomes) and 2 h (lysosomes). Within 3 h of exposure, azithromycin also causes the appearance of large and light-lucentlelectron-lucent vacuoles, most of which can be labeled by lucifer yellow when this tracer is added to culture prior to azithromycin exposure. Three days of treatment with azithromycin result in the accumulation of very large vesicles filled with pleiomorphic content, consistent with phospholipidosis. These vesicles are accessible to fluorescein-labeled bovine serum albumin (FITC-BSA) and intensively stained with filipin, indicating a mixed storage with cholesterol. The impairment of HRP pinocytosis directly correlates with the amount of azithromycin accumulated by the cells, but not with the phospholipidosis induced by the drug. The proton ionophore monensin, which completely suppresses azithromycin accumulation, also prevents inhibition of HRP uptake. Erythromycylamine, another dicationic macrolide, also inhibits HRP pinocytosis in direct correlation with its cellular accumulation and is as potent as azithromycin at equimolar cellular concentrations. We suggest that dicationic macrolides inhibit fluid-phase pinocytosis by impairing the formation of pinocytic vacuoles and endosomes.