RESUMEN
A novel method for applying high-throughput docking to challenging metalloenzyme targets is described. The method utilizes information-based virtual transformation of library carboxylates to hydroxamic acids prior to docking, followed by compound acquisition, one-pot (two steps) chemical synthesis and in vitro screening. In two experiments targeting the botulinum neurotoxin serotype A metalloprotease light chain, hit rates of 32% and 18% were observed.
Asunto(s)
Toxinas Botulínicas Tipo A/antagonistas & inhibidores , Inhibidores de Proteasas/química , Sitios de Unión , Toxinas Botulínicas Tipo A/metabolismo , Ácidos Carboxílicos/química , Dominio Catalítico , Química Farmacéutica , Diseño de Fármacos , Evaluación Preclínica de Medicamentos , Ácidos Hidroxámicos/química , Simulación del Acoplamiento Molecular , Inhibidores de Proteasas/síntesis química , Inhibidores de Proteasas/metabolismo , Unión Proteica , Serotipificación , Bibliotecas de Moléculas Pequeñas/síntesis química , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/metabolismo , Relación Estructura-ActividadRESUMEN
Four core structures capable of providing sub-nanomolar inhibitors of anthrax lethal factor (LF) were evaluated by comparing the potential for toxicity, physicochemical properties, in vitro ADME profiles, and relative efficacy in a rat lethal toxin (LT) model of LF intoxication. Poor efficacy in the rat LT model exhibited by the phenoxyacetic acid series (3) correlated with low rat microsome and plasma stability. Specific molecular interactions contributing to the high affinity of inhibitors with a secondary amine in the C2-side chain were revealed by X-ray crystallography.
Asunto(s)
Acetatos/síntesis química , Carbunco/tratamiento farmacológico , Antídotos/síntesis química , Bacillus anthracis/efectos de los fármacos , Toxinas Bacterianas/antagonistas & inhibidores , Acetatos/farmacocinética , Acetatos/farmacología , Animales , Antídotos/farmacocinética , Antídotos/farmacología , Antígenos Bacterianos , Bacillus anthracis/fisiología , Cristalografía por Rayos X , Citocromo P-450 CYP2D6/metabolismo , Inhibidores del Citocromo P-450 CYP2D6 , Microsomas Hepáticos/enzimología , Modelos Moleculares , Conejos , RatasRESUMEN
BACKGROUND: Dengue fever, dengue haemorrhagic fever, and dengue shock syndrome are caused by infections with any of the four serotypes of the dengue virus (DENV), and are an increasing global health risk. The related West Nile virus (WNV) causes significant morbidity and mortality as well, and continues to be a threat in endemic areas. Currently no FDA-approved vaccines or therapeutics are available to prevent or treat any of these infections. Like the other members of Flaviviridae, DENV and WNV encode a protease (NS3) which is essential for viral replication and therefore is a promising target for developing therapies to treat dengue and West Nile infections. METHODS: Flaviviral protease inhibitors were identified and biologically characterized for mechanism of inhibition and DENV antiviral activity. RESULTS: A guanidinylated 2,5-dideoxystreptamine class of compounds was identified that competitively inhibited the NS3 protease from DENV(1-4) and WNV with 50% inhibitory concentration values in the 1-70 µM range. Cytotoxicity was low; however, antiviral activity versus DENV-2 on VERO cells was not detectable. CONCLUSIONS: This class of compounds is the first to demonstrate competitive pan-dengue and WNV NS3 protease inhibition and, given the sequence conservation among flavivirus NS3 proteins, suggests that developing a pan-dengue or possibly pan-flavivirus therapeutic is feasible.
Asunto(s)
Antivirales/farmacología , Virus del Dengue/efectos de los fármacos , Inhibidores de Proteasas/farmacología , Serina Endopeptidasas/metabolismo , Proteínas no Estructurales Virales/antagonistas & inhibidores , Virus del Nilo Occidental/efectos de los fármacos , Animales , Antivirales/química , Chlorocebus aethiops , Virus del Dengue/enzimología , Virus del Dengue/aislamiento & purificación , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Peso Molecular , Inhibidores de Proteasas/química , ARN Helicasas/antagonistas & inhibidores , ARN Helicasas/metabolismo , Relación Estructura-Actividad , Células Vero , Proteínas no Estructurales Virales/metabolismo , Virus del Nilo Occidental/enzimología , Virus del Nilo Occidental/aislamiento & purificaciónRESUMEN
Neurotoxins synthesized by Clostridium botulinum bacteria (BoNT), the etiological agent of human botulism, are extremely toxic proteins making them high-risk agents for bioterrorism. Small molecule inhibitor development has been focused on the light chain zinc-dependent metalloprotease domain of the neurotoxin, an effort that has been hampered by its relatively flexible active site. Developed in concert with structure--activity relationship studies, the X-ray crystal structures of the complex of BoNT serotype A light chain (BoNT/A LC) with three different micromolar-potency hydroxamate-based inhibitors are reported here. Comparison with an unliganded BoNT/A LC structure reveals significant changes in the active site as a result of binding by the unique inhibitor scaffolds. The 60/70 loop at the opening of the active site pocket undergoes the largest conformational change, presumably through an induced-fit mechanism, resulting in the most compact catalytic pocket observed in all known BoNT/A LC structures.
Asunto(s)
Toxinas Botulínicas Tipo A/química , Quelantes/química , Clostridium botulinum/enzimología , Ácidos Hidroxámicos/química , Metaloproteasas/química , Neurotoxinas/química , Zinc/química , Sitios de Unión , Toxinas Botulínicas Tipo A/antagonistas & inhibidores , Toxinas Botulínicas Tipo A/metabolismo , Quelantes/metabolismo , Cristalografía por Rayos X , Ácidos Hidroxámicos/metabolismo , Ligandos , Metaloproteasas/antagonistas & inhibidores , Metaloproteasas/metabolismo , Neurotoxinas/antagonistas & inhibidores , Neurotoxinas/metabolismo , Unión Proteica , Conformación Proteica , Estructura Terciaria de Proteína , Zinc/metabolismoRESUMEN
New anthrax lethal factor inhibitors (LFIs) were designed based upon previously identified potent inhibitors 1a and 2. Combining the new core structures with modifications to the C2-side chain yielded analogs with improved efficacy in the rat lethal toxin model.
Asunto(s)
Antídotos/uso terapéutico , Antígenos Bacterianos/envenenamiento , Toxinas Bacterianas/envenenamiento , Animales , Estructura Molecular , Intoxicación/tratamiento farmacológico , RatasRESUMEN
Sub-nanomolar small molecule inhibitors of anthrax lethal factor have been identified using SAR and Merck L915 (4) as a model compound. One of these compounds (16) provided 100% protection in a rat lethal toxin model of anthrax disease.