RESUMEN
The discovery of novel antibiotic classes has not kept pace with the growing threat of bacterial resistance. Antibiotic candidates that act at new targets or via distinct mechanisms have the greatest potential to overcome resistance; however, novel approaches are also associated with higher attrition and longer timelines. This uncertainty has contributed to the withdrawal from antibiotic programs by many pharmaceutical companies. Genomic approaches have not yielded satisfactory results, in part due to nascent knowledge about unprecedented molecular targets, the challenge of achieving antibacterial activity by lead optimization of enzyme inhibitors, and the limitations of compound screening libraries for antibacterial discovery. Enhanced diversity of compound screening banks, entry into new chemical space, and new screening technologies are currently being exploited to improve hit rates for antibacterial discovery. Antibacterial compound lead optimization faces hurdles associated with the high plasma exposures required for efficacy. Lead optimization would be enhanced by the identification of new antibiotic classes with improved tractability and by expanding the predictability of in vitro safety assays. Implementing multiple screening and target identification strategies is recommended for improving the likelihood of discovering new antibacterial compounds that address unmet needs.
Asunto(s)
Antibacterianos/uso terapéutico , Infecciones Bacterianas/tratamiento farmacológico , Descubrimiento de Drogas/métodos , Farmacorresistencia Bacteriana/efectos de los fármacos , Animales , Diseño de Fármacos , Evaluación Preclínica de Medicamentos/métodos , Industria Farmacéutica/métodos , Humanos , Pruebas de Sensibilidad MicrobianaRESUMEN
We developed a homogenous microtiter based assay using the cationic dye 3, 3'-Diethyloxacarbocyanine iodide, DiOC2(3), to measure the effect of compounds on membrane potential in Staphylococcus aureus. In a screen of 372 compounds from a synthetic compound collection with anti-Escherichia coli activity due to unknown modes of action at least 17% demonstrated potent membrane activity, enabling rapid discrimination of nuisance compounds.
Asunto(s)
Antibacterianos/farmacología , Técnicas Bacteriológicas/métodos , Ensayos Analíticos de Alto Rendimiento/métodos , Potenciales de la Membrana/efectos de los fármacos , Staphylococcus aureus/efectos de los fármacos , Carbocianinas/metabolismo , Escherichia coli/efectos de los fármacos , Concentración 50 InhibidoraRESUMEN
Retapamulin MICs of > or =2 microg/ml were noted for 6 of 5,676 S. aureus recent clinical isolates evaluated. The ABC proteins VgaAv and VgaA were found to be responsible for the reduced susceptibility to pleuromutilins exhibited by these six isolates.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Antibacterianos/farmacología , Proteínas Bacterianas/genética , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Farmacorresistencia Bacteriana , Staphylococcus aureus/efectos de los fármacos , Diterpenos/farmacología , Farmacorresistencia Bacteriana/genética , Humanos , Pruebas de Sensibilidad Microbiana , Datos de Secuencia Molecular , Compuestos Policíclicos , Análisis de Secuencia de ADN , Staphylococcus aureus/genética , Staphylococcus aureus/aislamiento & purificación , PleuromutilinasRESUMEN
To assess their effects on susceptibility to retapamulin in Staphylococcus aureus, first-, second-, and third-step mutants with elevated MICs to tiamulin and other investigational pleuromutilin compounds were isolated and characterized through exposure to high drug concentrations. All first- and second-step mutations were in rplC, encoding ribosomal protein L3. Most third-step mutants acquired a third mutation in rplC. While first- and second-step mutations did cause an elevation in tiamulin and retapamulin MICs, a significant decrease in activity was not seen until a third mutation was acquired. All third-step mutants exhibited severe growth defects, and faster-growing variants arose at a high frequency from most isolates. These faster-growing variants were found to be more susceptible to pleuromutilins. In the case of a mutant with three alterations in rplC, the fast-growing variants acquired an additional mutation in rplC. In the case of fast-growing variants of isolates with two mutations in rplC and at least one mutation at an unmapped locus, one of the two rplC mutations reverted to wild type. These data indicate that mutations in rplC that lead to pleuromutilin resistance have a direct, negative effect on fitness. While reduction in activity of retapamulin against S. aureus can be seen through mutations in rplC, it is likely that target-specific resistance to retapamulin will be slow to emerge due to the need for three mutations for a significant effect on activity and the fitness cost of each mutational step.
Asunto(s)
Antibacterianos/farmacología , Farmacorresistencia Bacteriana/genética , Mutación , Proteínas Ribosómicas/genética , Staphylococcus aureus/efectos de los fármacos , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Diterpenos/farmacología , Humanos , Pruebas de Sensibilidad Microbiana , Compuestos Policíclicos , Proteína Ribosomal L3 , Staphylococcus aureus/genética , Staphylococcus aureus/metabolismo , PleuromutilinasRESUMEN
A novel benzylidenethiazolidinedione has been discovered with antimicrobial activity. Here, we present the results of a structure-activity study on this compound with respect to its antimicrobial activity.
Asunto(s)
Antibacterianos/síntesis química , Antibacterianos/farmacología , Compuestos de Bencilo/síntesis química , Compuestos de Bencilo/farmacología , Tiazolidinedionas/síntesis química , Tiazolidinedionas/farmacología , Enterococcus faecalis/efectos de los fármacos , Bacterias Gramnegativas/efectos de los fármacos , Indicadores y Reactivos , Pruebas de Sensibilidad Microbiana , Staphylococcus aureus/efectos de los fármacos , Streptococcus pneumoniae/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
Bacterial enoyl-ACP reductase (FabI) is responsible for catalyzing the final step of bacterial fatty acid biosynthesis and is an attractive target for the development of novel antibacterial agents. Previously we reported the development of FabI inhibitor 4 with narrow spectrum antimicrobial activity and in vivo efficacy against Staphylococcus aureus via intraperitoneal (ip) administration. Through iterative medicinal chemistry aided by X-ray crystal structure analysis, a new series of inhibitors has been developed with greatly increased potency against FabI-containing organisms. Several of these new inhibitors have potent antibacterial activity against multidrug resistant strains of S. aureus, and compound 30 demonstrates exceptional oral (po) in vivo efficacy in a S. aureus infection model in rats. While optimizing FabI inhibitory activity, compounds 29 and 30 were identified as having low micromolar FabK inhibitory activity, thereby increasing the antimicrobial spectrum of these compounds to include the FabK-containing pathogens Streptococcus pneumoniae and Enterococcus faecalis. The results described herein support the hypothesis that bacterial enoyl-ACP reductases are valid targets for antibacterial agents.
Asunto(s)
Acrilamidas/síntesis química , Antibacterianos/síntesis química , Inhibidores Enzimáticos/síntesis química , Ácido Graso Sintasas/antagonistas & inhibidores , Indoles/síntesis química , Naftiridinas/síntesis química , Oxidorreductasas/antagonistas & inhibidores , Absceso/tratamiento farmacológico , Acrilamidas/química , Acrilamidas/farmacología , Administración Oral , Animales , Antibacterianos/química , Antibacterianos/farmacología , Cristalografía por Rayos X , Farmacorresistencia Bacteriana , Enoil-ACP Reductasa (NADH) , Enterococcus faecalis/efectos de los fármacos , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Haemophilus influenzae/efectos de los fármacos , Indoles/química , Indoles/farmacología , Pruebas de Sensibilidad Microbiana , Modelos Moleculares , Naftiridinas/química , Naftiridinas/farmacología , Ratas , Staphylococcus aureus/efectos de los fármacos , Estereoisomerismo , Relación Estructura-Actividad , Triclosán/farmacologíaRESUMEN
The MICs of triclosan for 31 clinical isolates of Staphylococcus aureus were 0.016 micro g/ml (24 strains), 1 to 2 micro g/ml (6 strains), and 0.25 micro g/ml (1 strain). All the strains for which triclosan MICs were elevated (>0.016 micro g/ml) showed three- to fivefold increases in their levels of enoyl-acyl carrier protein (ACP) reductase (FabI) production. Furthermore, strains for which triclosan MICs were 1 to 2 micro g/ml overexpressed FabI with an F204C alteration. Binding studies with radiolabeled NAD(+) demonstrated that this change prevents the formation of the stable triclosan-NAD(+)-FabI complex, and both this alteration and its overexpression contributed to achieving MICs of 1 to 2 micro g/ml for these strains. Three novel, potent inhibitors of FabI (50% inhibitory concentrations, < or =64 nM) demonstrated up to 1,000-fold better activity than triclosan against the strains for which triclosan MICs were elevated. None of the compounds tested from this series formed a stable complex with NAD(+)-FabI. Consequently, although the overexpression of wild-type FabI gave rise to an increase in the MICs, as expected, overexpression of FabI with an F204C alteration did not cause an additional increase in resistance. Therefore, this work identifies the mechanisms of triclosan resistance in S. aureus, and we present three compounds from a novel chemical series of FabI inhibitors which have excellent activities against both triclosan-resistant and -sensitive clinical isolates of S. aureus.
Asunto(s)
Antiinfecciosos Locales/farmacología , Infecciones Estafilocócicas/microbiología , Staphylococcus aureus/efectos de los fármacos , Triclosán/farmacología , Antiinfecciosos Locales/metabolismo , Western Blotting , Cristalografía por Rayos X , Farmacorresistencia Bacteriana , Enoil-ACP Reductasa (NADH) , Humanos , Cinética , Pruebas de Sensibilidad Microbiana , Modelos Moleculares , Conformación Molecular , Oxidorreductasas/antagonistas & inhibidores , Oxidorreductasas/biosíntesis , Oxidorreductasas/aislamiento & purificación , Unión Proteica , Staphylococcus aureus/enzimología , Triclosán/metabolismoRESUMEN
Bacterial enoyl-acyl carrier protein (ACP) reductase (FabI) catalyzes the final step in each elongation cycle of bacterial fatty acid biosynthesis and is an attractive target for the development of new antibacterial agents. High-throughput screening of the Staphylococcus aureus FabI enzyme identified a novel, weak inhibitor with no detectable antibacterial activity against S. aureus. Iterative medicinal chemistry and X-ray crystal structure-based design led to the identification of compound 4 [(E)-N-methyl-N-(2-methyl-1H-indol-3-ylmethyl)-3-(7-oxo-5,6,7,8-tetrahydro-1,8-naphthyridin-3-yl)acrylamide], which is 350-fold more potent than the original lead compound obtained by high-throughput screening in the FabI inhibition assay. Compound 4 has exquisite antistaphylococci activity, achieving MICs at which 90% of isolates are inhibited more than 500 times lower than those of nine currently available antibiotics against a panel of multidrug-resistant strains of S. aureus and Staphylococcus epidermidis. Furthermore, compound 4 exhibits excellent in vivo efficacy in an S. aureus infection model in rats. Biochemical and genetic approaches have confirmed that the mode of antibacterial action of compound 4 and related compounds is via inhibition of FabI. Compound 4 also exhibits weak FabK inhibitory activity, which may explain its antibacterial activity against Streptococcus pneumoniae and Enterococcus faecalis, which depend on FabK and both FabK and FabI, respectively, for their enoyl-ACP reductase function. These results show that compound 4 is representative of a new, totally synthetic series of antibacterial agents that has the potential to provide novel alternatives for the treatment of S. aureus infections that are resistant to our present armory of antibiotics.
Asunto(s)
Antibacterianos , Inhibidores Enzimáticos , Oxidorreductasas/antagonistas & inhibidores , Animales , Antibacterianos/química , Antibacterianos/farmacocinética , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Farmacorresistencia Bacteriana Múltiple , Enoil-ACP Reductasa (NADH) , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacocinética , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Bacterias Gramnegativas/efectos de los fármacos , Bacterias Gramnegativas/enzimología , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Ratas , Ratas Sprague-Dawley , Infecciones Estafilocócicas/tratamiento farmacológico , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/enzimología , Streptococcus pneumoniae/efectos de los fármacos , Streptococcus pneumoniae/enzimología , Relación Estructura-ActividadRESUMEN
Bacterial enoyl-ACP reductase (FabI) catalyzes the final step in each cycle of bacterial fatty acid biosynthesis and is an attractive target for the development of new antibacterial agents. Our efforts to identify potent, selective FabI inhibitors began with screening of the GlaxoSmithKline proprietary compound collection, which identified several small-molecule inhibitors of Staphylococcus aureus FabI. Through a combination of iterative medicinal chemistry and X-ray crystal structure based design, one of these leads was developed into the novel aminopyridine derivative 9, a low micromolar inhibitor of FabI from S. aureus (IC(50) = 2.4 microM) and Haemophilus influenzae (IC(50) = 4.2 microM). Compound 9 has good in vitro antibacterial activity against several organisms, including S. aureus (MIC = 0.5 microg/mL), and is effective in vivo in a S. aureus groin abscess infection model in rats. Through FabI overexpressor and macromolecular synthesis studies, the mode of action of 9 has been confirmed to be inhibition of fatty acid biosynthesis via inhibition of FabI. Taken together, these results support FabI as a valid antibacterial target and demonstrate the potential of small-molecule FabI inhibitors for the treatment of bacterial infections.