RESUMO
Selection of resistant strains in Streptococcus pneumoniae was studied in vitro with nemonoxacin, a novel nonfluorinated quinolone (NFQ), in comparison with quinolone benchmarks, ciprofloxacin, garenoxacin, and gatifloxacin. In stepwise resistance selection studies, a 256-fold loss of potency was observed after three to four steps of exposure to ciprofloxacin or garenoxacin. In contrast, the loss of potency was limited to eightfold after three steps of exposure to nemonoxacin and repeated attempts to isolate highly resistant organisms after four steps of exposure yielded isolates that could not be subcultured in liquid medium. The quinolone resistance-determining regions of the target genes, parC, parE, gyrA, and gyrB, were analyzed through DNA sequencing. Known mutations, especially in the hotspots of parC and gyrA, were selected with exposure to garenoxacin, ciprofloxacin, and gatifloxacin. In contrast, mutations selected with nemonoxacin were limited to GyrA, GyrB, and ParE, sparing ParC, which is known as a key driver of resistance in clinical isolates of S. pneumoniae. This observation is consistent with previous data using other NFQs, which showed no loss of potency due to ParC mutations in clinical isolates. This apparently unique feature of nemonoxacin is potentially attributable to the structural uniqueness of the NFQs, distinguishing them from the fluoroquinolones that are commonly prescribed for infections by S. pneumoniae.
Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Mutação , Quinolonas/farmacologia , Streptococcus pneumoniae/efeitos dos fármacos , Ciprofloxacina/farmacologia , Meios de Cultura/química , DNA Girase/genética , DNA Topoisomerase IV/genética , Farmacorresistência Bacteriana/genética , Fluoroquinolonas/farmacologia , Gatifloxacina , Expressão Gênica , Testes de Sensibilidade Microbiana , Análise de Sequência de DNA , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/crescimento & desenvolvimento , Relação Estrutura-AtividadeRESUMO
In this article we describe the application of structural biology methods to the discovery of novel potent inhibitors of methionine aminopeptidases. These enzymes are employed by the cells to cleave the N-terminal methionine from nascent peptides and proteins. As this is one of the critical steps in protein maturation, it is very likely that inhibitors of these enzymes may prove useful as novel antibacterial agents. Involvement of crystallography at the very early stages of the inhibitor design process resulted in serendipitous discovery of a new inhibitor class, the pyrazole-diamines. Atomic-resolution structures of several inhibitors bound to the enzyme illuminate a new mode of inhibitor binding.
Assuntos
Bactérias/enzimologia , Inibidores de Proteases/farmacologia , Aminopeptidases/química , Aminopeptidases/isolamento & purificação , Bactérias/efeitos dos fármacos , Proteínas de Bactérias/farmacologia , Cristalização , Cristalografia por Raios X , Cinética , Espectroscopia de Ressonância Magnética , Metionil Aminopeptidases , Modelos Moleculares , Inibidores de Proteases/química , Conformação Proteica , Teoria QuânticaRESUMO
A high-throughput screen (HTS) was developed and used to identify inhibitors of bacterial DNA gyrase. Among the validated hits were 53 compounds that also inhibited mammalian topoisomerase II with IC(50) values of <12.5 micro g/mL for 51 of them. Using computational methods, these compounds were subjected to cluster analysis to categorize them according to their chemical and structural properties. Nine compounds from different clusters were tested for their whole-cell inhibitory activity against 3 cancer cell lines-NCI-H460 (lung), MCF7 (breast), and SF-268 (CNS)-at a concentration of 100 micro M. Five compounds inhibited cell growth by >50% for all 3 cell lines tested. These compounds were tested further against a panel of 53 to 57 cell lines representing leukemia, melanoma, colon, CNS, ovarian, renal, prostate, breast, and non-small cell lung cancers. In this assay, PGE-7143417 was found to be the most potent compound, which inhibited the growth of all the cell lines by 50% at a concentration range of 0.31 to 2.58 micro M, with an average of 1.21 micro M. An additional 17 compounds were also tested separately against a panel of 10 cell lines representing melanoma, colon, lung, mammary, ovarian, prostate, and renal cancers. In this assay, 4 compounds-PGE-3782569, PGE-7411516, PGE-2908955, and PGE-3521917-were found to have activity with concentrations for 50% cell growth inhibition in the 0.59 to 3.33, 22.5 to 59.1, 7.1 to >100, and 24.7 to >100 micro M range.
Assuntos
Antineoplásicos/metabolismo , Proteínas de Bactérias/metabolismo , Bioensaio/métodos , DNA Girase/metabolismo , DNA Topoisomerases Tipo II/metabolismo , Inibidores da Topoisomerase II , Animais , Anti-Infecciosos/metabolismo , Antineoplásicos/química , Linhagem Celular Tumoral , Ciprofloxacina/metabolismo , Desenho de Fármacos , Humanos , Estrutura MolecularRESUMO
Quinolones without the usual 6-fluorine substituent have been recently described as potent antibacterial agents. A series of non-fluorinated analogues of the antibacterial quinolone Levofloxacin were synthesized and tested.