RESUMEN
During 2015-2016, we evaluated the performance of whole-genome sequencing (WGS) as a routine typing tool. Its added value for microbiological and epidemiologic surveillance of listeriosis was compared with that for pulsed-field gel electrophoresis (PFGE), the current standard method. A total of 2,743 Listeria monocytogenes isolates collected as part of routine surveillance were characterized in parallel by PFGE and core genome multilocus sequence typing (cgMLST) extracted from WGS. We investigated PFGE and cgMLST clusters containing human isolates. Discrimination of isolates was significantly higher by cgMLST than by PFGE (p<0.001). cgMLST discriminated unrelated isolates that shared identical PFGE profiles and phylogenetically closely related isolates with distinct PFGE profiles. This procedure also refined epidemiologic investigations to include only phylogenetically closely related isolates, improved source identification, and facilitated epidemiologic investigations, enabling identification of more outbreaks at earlier stages. WGS-based typing should replace PFGE as the primary typing method for L. monocytogenes.
Asunto(s)
Genoma Bacteriano , Listeria monocytogenes/genética , Secuenciación Completa del Genoma/métodos , Brotes de Enfermedades , Monitoreo Epidemiológico , Microbiología de Alimentos , Francia/epidemiología , Humanos , Listeria monocytogenes/clasificación , Listeria monocytogenes/aislamiento & purificación , Listeriosis/epidemiología , Listeriosis/microbiología , Tipificación Molecular/métodosRESUMEN
High-throughput screening against the human sirtuin SIRT1 led to the discovery of a series of indoles as potent inhibitors that are selective for SIRT1 over other deacetylases and NAD-processing enzymes. The most potent compounds described herein inhibit SIRT1 with IC50 values of 60-100 nM, representing a 500-fold improvement over previously reported SIRT inhibitors. Preparation of enantiomerically pure indole derivatives allowed for their characterization in vitro and in vivo. Kinetic analyses suggest that these inhibitors bind after the release of nicotinamide from the enzyme and prevent the release of deacetylated peptide and O-acetyl-ADP-ribose, the products of enzyme-catalyzed deacetylation. These SIRT1 inhibitors are low molecular weight, cell-permeable, orally bioavailable, and metabolically stable. These compounds provide chemical tools to study the biology of SIRT1 and to explore therapeutic uses for SIRT1 inhibitors.
Asunto(s)
Carbazoles/síntesis química , Inhibidores de Histona Desacetilasas , Indoles/síntesis química , Sirtuinas/antagonistas & inhibidores , Animales , Disponibilidad Biológica , Células CHO , Carbazoles/química , Carbazoles/farmacología , Permeabilidad de la Membrana Celular , Cricetinae , Cricetulus , Estabilidad de Medicamentos , Fluorometría , Histona Desacetilasas/química , Humanos , Técnicas In Vitro , Indoles/química , Indoles/farmacología , Cinética , Ratones , Ratones Endogámicos C57BL , Microsomas Hepáticos/metabolismo , NAD/química , NAD+ Nucleosidasa/química , Niacinamida/química , Ratas , Proteínas Recombinantes/antagonistas & inhibidores , Proteínas Recombinantes/química , Sirtuina 1 , Sirtuinas/química , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
Heat shock protein 90 (Hsp90) plays a key role in stress response and protection of the cell against the effects of mutation. Herein we report the identification of an Hsp90 inhibitor identified by fragment screening using a high-concentration biochemical assay, as well as its optimisation by in silico searching coupled with a structure-based drug design (SBDD) approach.