Asunto(s)

Infecciones Bacterianas , Cefalosporinas , Infecciones Bacterianas/tratamiento farmacológico , Cefalosporinas/uso terapéutico , Humanos , Resultado del Tratamiento , Cefiderocol

RESUMEN

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 (Emax) and static concentrations (Cs) were calculated using the Hill equation (concentration-response curves). Finafloxacin pharmacokinetics in cells at pH 7.4 or 5.5 was investigated using 14C-labelled drug. RESULTS: Extracellularly, all drugs sterilized the cultures, with finafloxacin being two to six times more potent at acidic pH. Intracellularly, Emax reached the limit of detection (4-5 log10 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 log10 cfu decrease for all species. At acid pH, Cs 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.

Asunto(s)

Antibacterianos/farmacología , Burkholderia/efectos de los fármacos , Fluoroquinolonas/farmacología , Francisella/efectos de los fármacos , Yersinia pseudotuberculosis/efectos de los fármacos , Humanos , Concentración de Iones de Hidrógeno , Imipenem/farmacología , Levofloxacino/farmacología , Pruebas de Sensibilidad Microbiana , Monocitos , Células THP-1

RESUMEN

Objectives: When tested in broth, avibactam reverses ceftazidime resistance in many Pseudomonas aeruginosa that express ESBLs. We examined whether similar reversal is observed against intracellular forms of P. aeruginosa . Methods: Strains: reference strains; two engineered strains with basal non-inducible expression of AmpC and their isogenic mutants with stably derepressed AmpC; and clinical isolates with complete, partial or no resistance to reversion with avibactam. Pharmacodynamic model: 24 h concentration-response to ceftazidime [0.01-200 mg/L alone or with avibactam (4 mg/L)] of bacteria in broth or bacteria phagocytosed by THP-1 monocytes, with calculation of ceftazidime relative potency ( C s : concentration yielding a static effect) and maximal relative effect [ E max : cfu decrease at infinitely large antibiotic concentrations (efficacy in the model)] using the Hill equation. Cellular content of avibactam: quantification by LC-MS/MS. Results: For both extracellular and intracellular bacteria, ceftazidime C s was always close to its MIC. For ceftazidime-resistant strains, avibactam addition shifted ceftazidime C s to values close to the MIC of the combination in broth. E max was systematically below the detection limit (-5 log 10 ) for extracellular bacteria, but limited to -1.3 log 10 for intracellular bacteria (except for two isolates) with no effect of avibactam. The cellular concentration of avibactam reflected extracellular concentration and was not influenced by ceftazidime (0-160 mg/L). Conclusions: The potential for avibactam to inhibit ß-lactamases does not differ for extracellular and intracellular forms of P. aeruginosa , denoting an unhindered access to its target in both situations. The loss of maximal relative efficacy of ceftazidime against intracellular P. aeruginosa was unrelated to resistance via avibactam-inhibitable ß-lactamases.

Asunto(s)

Antibacterianos/farmacología , Compuestos de Azabiciclo/farmacología , Ceftazidima/farmacología , Leucocitos Mononucleares/microbiología , Pseudomonas aeruginosa/efectos de los fármacos , Inhibidores de beta-Lactamasas/farmacología , Citoplasma/efectos de los fármacos , Citoplasma/microbiología , Combinación de Medicamentos , Humanos , Cinética , Leucocitos Mononucleares/efectos de los fármacos , Pruebas de Sensibilidad Microbiana , Pseudomonas aeruginosa/genética , Espectrometría de Masas en Tándem

RESUMEN

INTRODUCTION: Discharge from the hospital is a period at risk for the continuity of patient's medication (seamless pharmaceutical care). The community pharmacist is often the first health care professional seen by the patient after hospital discharge. The clinical pharmacist has potentially a key role in establishing an efficient information transfer from the hospital to the community pharmacy. OBJECTIVE: (1) To develop and, (2) to evaluate the impact of a structured discharge medication form prepared at hospital discharge by the clinical pharmacist and containing information items related to the medication regimen for the community pharmacist, and (3) to survey the information needs of the Belgian community pharmacists to ensure continuity of care after hospitalization. METHODS: (1) A structured discharge medication form has been developed based on a Literature review and on opinions expressed by community and clinical pharmacists, members of the Belgian Pharmaceutical Union (Association Pharmaceutique Belge) and an ethical committee. (2) A prospective study has been conducted with patients from geriatrics and orthopaedics wards of the University Hospital Dinant-Godinne returning home after hospital discharge with the discharge medication form to be given to their commuiity pharmacist; its use, the reasons for non-use, the perceived impact and the satisfaction of the community pharmacist have been assessed. (3) An on-line survey addressed to all Belgian community pharmacists evaluated their information needs. RESULTS: (1) The final version of the discharge medication form included key information items concerning the hospital, the patient, the discharge treatment (including the type of modifications made as compared to medications taken before admission), and on medication management at home. Some items were excluded because of Lack of perceived utility by pharmacists, confidentiality issues, and respect of patient's freedom of choice. (2) From the 71 medication forms given to patients, 48 were received by the community pharmacist. One quarter of respondents stated that they did not use the form, the main reason being that it was received after dispensing of the discharge treatment (n=6/11). The majority of the community pharmacists considered most of the information items as useful and the discharge medication form as being valuable for continuity of care. Requests for additional information were made (e.g., reason of admission and of treatment modifications, etc.). (3) The utility, benefits, and need for additional information items beyond what was included in the discharge medication form were highlighted by the respondents (n=309) of the national survey. Most of these respondents confirmed the value of the different information items included in the discharge medication form. CONCLUSION: The transmission of a structured medication form containing information about the medication regimen upon hospital discharge is of real interest and value for the community pharmacist because it goes beyond what is usually provided on a medical prescription. However, this discharge medication form should include more information items for effective pharmaceutical care.

Asunto(s)

Servicios Comunitarios de Farmacia/organización & administración , Continuidad de la Atención al Paciente , Bélgica , Encuestas de Atención de la Salud , Hospitales , Humanos , Conciliación de Medicamentos , Alta del Paciente , Farmacéuticos , Estudios Prospectivos

RESUMEN

Continuous nationwide surveillance of invasive pneumococcal disease (IPD) was conducted in Germany. From July 1, 1997, to June 30, 2013, data on penicillin susceptibility were available for 20,437 isolates. 2,790 of these isolates (13.7 %) originate from patients with meningitis and 17,647 isolates (86.3 %) are from non-meningitis cases. A slight decline in isolates susceptible at 0.06 and 0.12 µg/ml can be noticed over the years. Overall, 89.1 % of the isolates had minimum inhibitory concentrations (MICs) of ≤0.015 µg/ml. In 2012/2013, the first three isolates of Streptococcus pneumoniae with MICs of 8 µg/ml were found. The application of different guidelines with other MIC breakpoints for the interpretation of penicillin resistance leads to differences in susceptibility categorisation. According to the pre-2008 Clinical and Laboratory Standards Institute (CLSI) interpretive criteria, 5.3 % of isolates overall were intermediate and 1.4 % were resistant to penicillin. Application of the 2008-2014 CLSI interpretive criteria resulted in 7.6 % resistance among meningitis cases and 0.5 % intermediate resistance in non-meningitis cases. Referring to the 2009-2014 European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints, 7.6 % of the isolates in the meningitis group were resistant to penicillin. In the non-meningitis group, 6.1 % of the isolates were intermediate and 0.5 % were resistant. These differences should be kept in mind when surveillance studies on pneumococcal penicillin resistance are compared.

Asunto(s)

Antibacterianos/farmacología , Penicilinas/farmacología , Infecciones Neumocócicas/microbiología , Streptococcus pneumoniae/efectos de los fármacos , Farmacorresistencia Bacteriana , Alemania , Humanos , Pruebas de Sensibilidad Microbiana/normas , Prevalencia , Streptococcus pneumoniae/aislamiento & purificación

RESUMEN

Extended and continuous infusions with beta-lactam antibiotics have been suggested as a means of pharmacokinetic and pharmacodynamic optimisation of antimicrobial therapy. Vancomycin is also frequently administered in continuous infusion, although more for practical reasons. A survey was undertaken to investigate the recommendations by the local antibiotic management teams (AMTs) in Belgian acute hospitals concerning the administration (intermittent, extended or continuous infusion) and therapeutic drug monitoring of four beta-lactam antibiotics (ceftazidime, cefepime, piperacillin-tazobactam, meropenem) and vancomycin for adult patients with a normal kidney function. A structured questionnaire survey comprising three domains was developed and approved by the members of the Belgian Antibiotic Policy Coordination Committee (BAPCOC). The questionnaire was sent by e-mail to the official AMT correspondents of 105 Belgian hospitals, followed by two reminders. The response rate was 32 %, with 94 %, 59 %, 100 %, 100 % and 100 % of the participating Belgian hospitals using ceftazidime, cefepime, piperacillin-tazobactam, meropenem and vancomycin, respectively. Comparing intensive care unit (ICU) with non-ICU wards showed a higher implementation of extended or continuous infusions for ceftazidime (81 % vs. 41 %), cefepime (35 % vs. 10 %), piperacillin-tazobactam (38 % vs. 12 %), meropenem (68 % vs. 35 %) and vancomycin (79 % vs. 44 %) on the ICU wards. A majority of the hospitals recommended a loading dose prior to the first dose. For vancomycin, the loading dose and the trough target concentration were too low based on the current literature. This survey shows that extended and continuous infusions with beta-lactams and vancomycin are widely implemented in Belgian hospitals.

Asunto(s)

Antibacterianos/administración & dosificación , Unidades de Cuidados Intensivos , Habitaciones de Pacientes , Vancomicina/administración & dosificación , beta-Lactamas/administración & dosificación , Bélgica , Encuestas de Atención de la Salud , Hospitales , Humanos , Encuestas y Cuestionarios

RESUMEN

Staphylococcus aureus small-colony variants (SCVs) persist intracellularly, which may contribute to persistence/recurrence of infections and antibiotic failure. We have studied the intracellular fate of menD and hemB mutants (corresponding to menadione- and hemin-dependent SCVs, respectively) of the COL methicillin-resistant S. aureus (MRSA) strain and the antibiotic pharmacodynamic profile against extracellular (broth) and intracellular (human THP-1 monocytes) bacteria. Compared to the parental strain, SCVs showed slower extracellular growth (restored upon medium supplementation with menadione or hemin), reduced phagocytosis, and, for the menD SCV, lower intracellular counts at 24 h postinfection. Against extracellular bacteria, daptomycin, gentamicin, rifampin, moxifloxacin, and oritavancin showed similar profiles of activity against all strains, with a static effect obtained at concentrations close to their MICs and complete eradication as maximal effect. In contrast, vancomycin was not bactericidal against SCVs. Against intracellular bacteria, concentration-effect curves fitted sigmoidal regressions for vancomycin, daptomycin, gentamicin, and rifampin (with maximal effects lower than a 2-log decrease in CFU) but biphasic regressions (with a maximal effect greater than a 3-log decrease in CFU) for moxifloxacin and oritavancin, suggesting a dual mode of action against intracellular bacteria. For all antibiotics, these curves were indistinguishable between the strains investigated, except for the menD mutant, which systematically showed a lower amplitude of the concentration-effect response, with markedly reduced minimal efficacy (due to slower growth) but no change in maximal efficacy. The data therefore show that the maximal efficacies of antibiotics are similar against normal-phenotype and menadione- and hemin-dependent strains despite their different intracellular fates, with oritavancin, and to some extent moxifloxacin, being the most effective.

Asunto(s)

Antibacterianos/farmacología , Hemina/metabolismo , Monocitos/microbiología , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/metabolismo , Vitamina K 3/metabolismo , Antibacterianos/farmacocinética , Línea Celular , Daptomicina/farmacocinética , Daptomicina/farmacología , Gentamicinas/farmacocinética , Gentamicinas/farmacología , Glicopéptidos/farmacocinética , Glicopéptidos/farmacología , Humanos , Lipoglucopéptidos , Pruebas de Sensibilidad Microbiana , Rifampin/farmacocinética , Rifampin/farmacología , Infecciones Estafilocócicas , Vancomicina/farmacocinética , Vancomicina/farmacología

RESUMEN

DD-ligases catalyze the synthesis of the D-Ala-D-Ala and D-Ala-D-Ser dipeptides or the D Ala-D-Lac depsipeptide in an early step of peptidoglycan synthesis. Their function is essential for bacterial growth and specific to bacteria, making them attractive targets for the development of novel antibiotics. This review examines the biochemical and structural features of these enzymes and presents the main families of inhibitors described so far. Over the last 20 years, 7 structures of DD-ligases have been solved by X-ray crystallography, giving a detailed view of the general topology of the active site and of the residues in the catalytic pocket that play a central role in substrate recognition. This has paved the way to the rational design of inhibitors, which can be classified as (i) analogues of substrates, (ii) analogues of the product of the reaction, (iii) analogues of the transition state, and (iv) original scaffolds discovered by screening or by rational computer-aided design. The three first strategies have led to molecules that are polar by nature and have therefore poor access to their cytosolic target. The fourth one is potentially most promising as it yields more diverse structures. The most active molecules show affinity constants in the microM range, but microbiological evaluation remains scarce (typical MIC 1-8 mg/L for the tested compounds). These data strongly suggest targeting DD-ligases is a promising approach for discovery of new antibiotics. Future research should, however, aim at finding more potent inhibitors endowed with the appropriate pharmacokinetic properties that ensure access to their intracellular target.

Asunto(s)

Antibacterianos/química , Proteínas Bacterianas/química , Dipéptidos/química , Ligasas/química , Secuencia de Aminoácidos , Antibacterianos/farmacología , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Dipéptidos/metabolismo , Descubrimiento de Drogas/métodos , Descubrimiento de Drogas/tendencias , Ligasas/antagonistas & inhibidores , Ligasas/metabolismo , Datos de Secuencia Molecular , Estructura Molecular , Estructura Terciaria de Proteína , Homología de Secuencia de Aminoácido

RESUMEN

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.

Asunto(s)

Antibacterianos/líquido cefalorraquídeo , Antibacterianos/farmacocinética , Meningitis Bacterianas/tratamiento farmacológico , Acetamidas/uso terapéutico , Antibacterianos/uso terapéutico , Transporte Biológico , Humanos , Linezolid , Meningitis Bacterianas/líquido cefalorraquídeo , Resistencia a la Meticilina , Oxazolidinonas/uso terapéutico , Sensibilidad y Especificidad , Solubilidad , Infecciones Estafilocócicas/líquido cefalorraquídeo , Infecciones Estafilocócicas/tratamiento farmacológico , Staphylococcus aureus/efectos de los fármacos , Relación Estructura-Actividad , Vancomicina/uso terapéutico , beta-Lactamas/líquido cefalorraquídeo , beta-Lactamas/farmacocinética , beta-Lactamas/uso terapéutico

RESUMEN

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.

Asunto(s)

Antibacterianos/farmacología , Bacteriemia/microbiología , Endocarditis Bacteriana/microbiología , Macrófagos/microbiología , Resistencia a la Meticilina , Staphylococcus aureus/efectos de los fármacos , Anciano , Daptomicina/farmacología , Farmacorresistencia Bacteriana , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Infecciones Estafilocócicas/microbiología , Staphylococcus aureus/aislamiento & purificación , Vancomicina/farmacología

RESUMEN

Apoptosis plays a central role not only in the physiological processes of kidney growth and remodeling, but also in various human renal diseases and drug-induced nephrotoxicity. We present in a synthetic fashion the main molecular and cellular pathways leading to drug-induced apoptosis in kidney and the mechanisms regulating it. We illustrate them using three main nephrotoxic drugs (cisplatin, gentamicin, and cyclosporine A). We discuss the main regulators and effectors that have emerged as key targets for the design of therapeutic strategies. Novel approaches using gene therapy, antisense strategies, recombinant proteins, or compounds obtained from both classical organic and combinatorial chemistry are examined. Finally, key issues that need to be addressed for the success of apoptosis-based therapies are underlined.

Asunto(s)

Proteínas Reguladoras de la Apoptosis/metabolismo , Apoptosis/efectos de los fármacos , Caspasas/metabolismo , Cisplatino/toxicidad , Ciclosporina/toxicidad , Gentamicinas/toxicidad , Riñón/efectos de los fármacos , Animales , Cisplatino/metabolismo , Ciclosporina/metabolismo , Gentamicinas/metabolismo , Humanos , Riñón/citología , Redes y Vías Metabólicas , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Insuficiencia Renal/metabolismo

RESUMEN

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.

Asunto(s)

Antibacterianos/uso terapéutico , Farmacorresistencia Bacteriana Múltiple , Infecciones por Pseudomonas/tratamiento farmacológico , Pseudomonas aeruginosa/efectos de los fármacos , Antibacterianos/farmacología , Infección Hospitalaria/tratamiento farmacológico , Humanos

RESUMEN

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.

Asunto(s)

Antiinfecciosos/uso terapéutico , Quinolonas/uso terapéutico , Antiinfecciosos/efectos adversos , Antiinfecciosos/química , Antiinfecciosos/farmacología , Bacterias/efectos de los fármacos , Infecciones Bacterianas/tratamiento farmacológico , Infecciones Bacterianas/epidemiología , Farmacorresistencia Bacteriana/efectos de los fármacos , Humanos , Pruebas de Sensibilidad Microbiana , Quinolonas/efectos adversos , Quinolonas/química , Quinolonas/farmacología , Relación Estructura-Actividad

RESUMEN

OBJECTIVES: Quinolones accumulate in eukaryotic cells and show activity against a large array of intracellular organisms, but systematic studies aimed at examining their pharmacodynamic profile against intracellular bacteria are scarce. The present work aims at comparing intracellular-to-extracellular activities in this context. METHODS: We assessed the activities of ciprofloxacin, levofloxacin, moxifloxacin and garenoxacin against the extracellular (broth) and intracellular (infected J774 macrophages) forms of Listeria monocytogenes (cytosolic infection) and Staphylococcus aureus (phagolysosomal infection) using a range of clinically meaningful extracellular concentrations (0.06-4 mg/L). RESULTS: All four quinolones displayed concentration-dependent bactericidal activity against extracellular and intracellular L. monocytogenes and S. aureus for extracellular concentrations in the range 1-4-fold their MIC. Compared at equipotent extracellular concentrations, intracellular activities against L. monocytogenes were roughly equal to those that were extracellular, but were 50-100 times lower against S. aureus. Because quinolones accumulate in cells (ciprofloxacin, approximately 3 times; levofloxacin, approximately 5 times; garenoxacin, approximately 10 times, moxifloxacin, approximately 13 times), these data show that, intracellularly, quinolones are 5-10 times less potent against L. monocytogenes (P=0.065 [ANCOVA]), and at least 100 times less potent (P < 0.0001) against S. aureus. Because of their lower MICs and higher accumulation levels, garenoxacin and moxifloxacin were, however, more active than ciprofloxacin and levofloxacin when compared at similar extracellular concentrations. CONCLUSIONS: Quinolone activity is reduced intracellulary. This suggests that either only a fraction of cell-associated quinolones exert an antibacterial effect, or that intracellular activity is defeated by the local environment, or that intracellular bacteria only poorly respond to the action of quinolones.

Asunto(s)

Antibacterianos/farmacología , Listeria monocytogenes/efectos de los fármacos , Staphylococcus aureus/efectos de los fármacos , Animales , Compuestos Aza/farmacología , Línea Celular Tumoral , Ciprofloxacina/farmacología , Fluoroquinolonas/farmacología , Levofloxacino , Listeria monocytogenes/fisiología , Macrófagos/microbiología , Pruebas de Sensibilidad Microbiana , Moxifloxacino , Ofloxacino/farmacología , Quinolinas/farmacología , Staphylococcus aureus/fisiología , Factores de Tiempo

RESUMEN

The macrolide antibiotic azithromycin was shown to markedly inhibit endocytosis. Here we investigate the interaction of azithromycin with biomembranes and its effects on membrane biophysics in relation to endocytosis. Equilibrium dialysis and 31P NMR revealed that azithromycin binds to lipidic model membranes and decreases the mobility of phospholipid phosphate heads. In contrast, azithromycin had no effect deeper in the bilayer, based on fluorescence polarization of TMA-DPH and DPH, compounds that, respectively, explore the interfacial and hydrophobic domains of bilayers, and it did not induce membrane fusion, a key event of vesicular trafficking. Atomic force microscopy showed that azithromycin perturbed lateral phase separation in Langmuir-Blodgett monolayers, indicating a perturbation of membrane organization in lateral domains. The consequence of azithromycin/ phospholipid interaction on membrane endocytosis was next evaluated in J774 macrophages by using three tracers with different insertion preferences inside the biological membranes and intracellular trafficking: C6-NBD-SM, TMA-DPH and N-Rh-PE. Azithromycin differentially altered their insertion into the plasma membrane, slowed down membrane trafficking towards lysosomes, as evaluated by the rate of N-Rh-PE self-quenching relief, but did not affect bulk membrane internalization of C6-NBD-SM and TMA-DPH. Azithromycin also decreased plasma membrane fluidity, as shown by TMA-DPH fluorescence polarization and confocal microscopy after labeling by fluorescent concanavalin A. We conclude that azithromycin directly interacts with phospholipids, modifies biophysical properties of membrane and affects membrane dynamics in living cells. This antibiotic may therefore help to elucidate the physico-chemical properties underlying endocytosis.

Asunto(s)

1,2-Dipalmitoilfosfatidilcolina/química , Azitromicina/química , Azitromicina/farmacología , Membrana Celular/metabolismo , Membrana Celular/ultraestructura , Liposomas/química , Fluidez de la Membrana/efectos de los fármacos , Antibacterianos/química , Antibacterianos/farmacología , Línea Celular , Membrana Celular/química , Endocitosis/efectos de los fármacos , Endocitosis/fisiología , Concentración de Iones de Hidrógeno , Lípidos/química , Sustancias Macromoleculares , Macrófagos/química , Macrófagos/citología , Macrófagos/metabolismo , Temperatura

Asunto(s)

Antibacterianos/metabolismo , Antibacterianos/farmacología , Proteínas Portadoras/metabolismo , Células Eucariotas/metabolismo , Animales , Antibacterianos/farmacocinética , Antibacterianos/toxicidad , Humanos , Absorción Intestinal

Asunto(s)

Antibacterianos/metabolismo , Antibacterianos/uso terapéutico , Proteínas Portadoras/metabolismo , Farmacorresistencia Bacteriana , Células Procariotas/metabolismo , Animales , Humanos

RESUMEN

Three pharmacokinetic/pharmacodynamic parameters--(i) the peak concentration to the minimum inhibitory concentration ratio (C(max)/MIC); (ii) the area under the concentration-time curve to MIC ratio (AUC(24h)/MIC); and (iii) the time the concentration exceeds the MIC (T>MIC)--are important predictors of the clinical efficacy of antibiotics. For antibiotics with pronounced concentration-dependent killing, such as the fluoroquinolones or the aminoglycosides, C(max)/MIC and AUC(24)/MIC are the main factors that establish efficacy. Antibiotics with a weak, or no, concentration dependency generally have their efficacy linked to T>MIC, and these include the beta-lactams and the conventional macrolides. Antibiotics with weak concentration-dependent effects, but with prolonged persistent effects, such as tetracyclines and azithromycin, have their activity mostly related to the AUC(24)/MIC. By applying these concepts to current antibiotics, and also to the development of novel agents, it is possible to optimise their dosages and administration schedules. This will maximise therapeutic efficacy, may prevent or delay the emergence of bacterial resistance to antibiotics, and can certainly minimise side-effects.

Asunto(s)

Antibacterianos/farmacocinética , Aminoglicósidos/administración & dosificación , Aminoglicósidos/farmacocinética , Aminoglicósidos/farmacología , Antibacterianos/administración & dosificación , Antibacterianos/farmacología , Bacterias/efectos de los fármacos , Bacterias/metabolismo , Relación Dosis-Respuesta a Droga , Farmacorresistencia Bacteriana , Humanos , Cinética , Lactamas , Macrólidos , Pruebas de Sensibilidad Microbiana , Quinolonas/administración & dosificación , Quinolonas/farmacocinética , Quinolonas/farmacología

RESUMEN

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.

Asunto(s)

Antibacterianos/farmacología , Azitromicina/farmacología , Eritromicina/análogos & derivados , Lisosomas/metabolismo , Pinocitosis/efectos de los fármacos , Adenosina Trifosfato/metabolismo , Animales , Antineoplásicos/farmacología , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Membrana Celular/ultraestructura , Células Cultivadas , Colorantes , ADN/biosíntesis , Eritromicina/farmacología , Feto/citología , Fibroblastos/citología , Peroxidasa de Rábano Silvestre/farmacocinética , Humanos , Ionóforos/farmacología , Lisosomas/efectos de los fármacos , Lisosomas/ultraestructura , Microscopía Electrónica , Monensina/farmacología , Nocodazol/farmacología , Fosfolípidos/metabolismo , Unión Proteica/efectos de los fármacos , Ratas , Ratas Wistar , Cloruro de Tolonio , Transferrina/metabolismo , Vesículas Transportadoras/efectos de los fármacos , Vesículas Transportadoras/metabolismo , Vesículas Transportadoras/ultraestructura , Vacuolas/efectos de los fármacos , Vacuolas/metabolismo , Vacuolas/ultraestructura

RESUMEN

The stability and compatibility of ceftazidime have been examined in the context of its potential use in concentrated solutions for continuous infusion in patients suffering from severe nosocomial pneumonia and receiving other intravenous medications by the same route. Ceftazidime stability in 4 to 12% solutions was found satisfactory (<10% degradation) for 24 h if kept at a temperature of 25 degrees C (77 degrees F) maximum. Studies mimicking the simultaneous administration of ceftazidime and other drugs as done in clinics showed physical incompatibilities with vancomycin, nicardipine, midazolam, and propofol and a chemical incompatibility with N-acetylcystein. Concentrated solutions (50 mg/ml) of erythromycin or clarithromycin caused the appearance of a precipitate, whereas gentamicin, tobramycin, amikacin, isepamicin, fluconazole, ketamine, sufentanil, valproic acid, furosemide, uradipil, and a standard amino acid solution were physically and chemically compatible.

Asunto(s)

Ceftazidima/administración & dosificación , Cefalosporinas/administración & dosificación , Ceftazidima/efectos adversos , Ceftazidima/metabolismo , Ceftazidima/uso terapéutico , Cefalosporinas/efectos adversos , Cefalosporinas/metabolismo , Cefalosporinas/uso terapéutico , Cuidados Críticos , Infección Hospitalaria/tratamiento farmacológico , Infección Hospitalaria/metabolismo , Incompatibilidad de Medicamentos , Estabilidad de Medicamentos , Quimioterapia Combinada , Humanos , Infusiones Intravenosas , Midazolam/farmacología , Nicardipino/farmacología , Neumonía Bacteriana/tratamiento farmacológico , Neumonía Bacteriana/metabolismo , Propofol/farmacología , Temperatura , Vancomicina/farmacología