RESUMEN
Aryl dihydrouracil derivatives were identified from high throughput screening as potent inhibitors of HCV NS5B polymerase. The aryl dihydrouracil derivatives were shown to be non-competitive with respect to template RNA and elongation nucleotide substrates. They demonstrated genotype 1 specific activity towards HCV NS5B polymerases. Structure activity relationships and genotype specific activities of aryl dihydrouracil derivatives suggested that they bind to the palm initiation nucleotide pocket, a hypothesis which was confirmed by studies with polymerases containing mutations in various inhibitor binding sites. Therefore, aryl dihydrouracil derivatives represent a novel class of palm initiation site inhibitors of HCV NS5B polymerase.
Asunto(s)
Inhibidores de Proteasas/química , Uracilo/análogos & derivados , Proteínas no Estructurales Virales/antagonistas & inhibidores , Sustitución de Aminoácidos , Genotipo , Hepacivirus/efectos de los fármacos , Hepacivirus/enzimología , Cinética , Inhibidores de Proteasas/síntesis química , Inhibidores de Proteasas/farmacología , Relación Estructura-Actividad , Sitio de Iniciación de la Transcripción , Uracilo/síntesis química , Uracilo/química , Uracilo/farmacología , Proteínas no Estructurales Virales/genética , Proteínas no Estructurales Virales/metabolismoRESUMEN
A series of quinoline derivatives was synthesized as potential bioisosteric replacements for the benzothiadiazine moiety of earlier Hepatitis C NS5B polymerase inhibitors. Several of these compounds exhibited potent activity in enzymatic and replicon assays.
Asunto(s)
Benzotiadiazinas/farmacología , Inhibidores de Proteasas/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Benzotiadiazinas/química , Hepacivirus/enzimología , Hepacivirus/fisiología , Inhibidores de Proteasas/química , Replicación ViralRESUMEN
Benzothiadiazine inhibitors of the HCV NS5B RNA-dependent RNA polymerase are an important class of non-nucleoside inhibitors that have received considerable attention in the search for novel HCV therapeutics. Research in our laboratories has identified a novel series of tetracyclic benzothiadiazine inhibitors of HCV polymerase bearing a benzylamino substituent on the B-ring. Compounds in this series exhibit low-nanomolar activities in both genotypes 1a and 1b polymerase inhibition assays and subgenomic replicon assays. Optimization of pharmacokinetic properties in rat led to compound 30, which has good oral bioavailability (F = 56%) and a favorable tissue distribution drug profile, with high liver to plasma ratios. Compound 30 is a potent inhibitor in replicon assays, with EC(50) values of 10 and 6 nM against genotypes 1a and 1b, respectively.
Asunto(s)
Benzotiadiazinas/síntesis química , Benzotiadiazinas/farmacología , Hepacivirus/enzimología , ARN Polimerasa Dependiente del ARN/antagonistas & inhibidores , Animales , Antivirales/síntesis química , Antivirales/farmacocinética , Antivirales/farmacología , Proteínas Bacterianas/antagonistas & inhibidores , Benzotiadiazinas/farmacocinética , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacocinética , Inhibidores Enzimáticos/farmacología , Genotipo , Hepacivirus/genética , Hígado/metabolismo , Pruebas de Sensibilidad Microbiana , Ratas , Relación Estructura-Actividad , Distribución TisularRESUMEN
The hepatitis C virus (HCV) NS5B polymerase is essential for viral replication and has been a prime target for drug discovery research. Our efforts directed toward the discovery of HCV polymerase inhibitors resulted in the identification of unsymmetrical dialkyl-hydroxynaphthalenoyl-benzothiadiazines 2 and 3. The most active compound displayed activity in genotypes 1a and 1b polymerase and replicon cell culture inhibition assays at subnanomolar and low nanomolar concentrations, respectively. It also displayed an excellent pharmacokinetic profile in rats, with a plasma elimination half-life after intravenous dosing of 4.5 h, oral bioavailability of 77%, and a peak liver concentration of 21.8 microg/mL.
Asunto(s)
Benzotiadiazinas/síntesis química , Benzotiadiazinas/farmacología , ARN Polimerasas Dirigidas por ADN/antagonistas & inhibidores , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Hepacivirus/enzimología , Animales , Benzotiadiazinas/farmacocinética , Disponibilidad Biológica , Inhibidores Enzimáticos/farmacocinética , Semivida , Humanos , Espectroscopía de Resonancia Magnética , Ratas , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
4,4-Dialkyl-1-hydroxy-3-oxo-3.4-dihydronaphthalene-3-yl benzothiadiazine derivatives were synthesized and evaluated as inhibitors of genotypes 1a and 1b HCV NS5B polymerase. A number of these compounds exhibited potent activity against genotypes 1a and 1b HCV polymerase in both enzymatic and cell culture activities. A representative compound also showed favorable pharmacokinetics in the rat.
Asunto(s)
Antivirales/síntesis química , Antivirales/farmacocinética , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacocinética , Proteínas no Estructurales Virales/antagonistas & inhibidores , Administración Oral , Animales , Área Bajo la Curva , Química Farmacéutica/métodos , Diseño de Fármacos , Genotipo , Infusiones Intravenosas , Concentración 50 Inhibidora , Modelos Químicos , Ratas , Proteínas no Estructurales Virales/genéticaRESUMEN
Halosalicylamide derivatives were identified from high-throughput screening as potent inhibitors of HCV NS5B polymerase. The subsequent structure and activity relationship revealed the absolute requirement of the salicylamide moiety for optimum activity. Methylation of either the hydroxyl group or the amide group of the salicylamide moiety abolished the activity while the substitutions on both phenyl rings are acceptable. The halosalicylamide derivatives were shown to be non-competitive with respect to elongation nucleotide and demonstrated broad genotype activity against genotype 1-3 HCV NS5B polymerases. Inhibitor competition studies indicated an additive binding mode to the initiation pocket that is occupied by the thiadiazine class of compounds and an additive binding mode to the elongation pocket that is occupied by diketoacids, but a mutually exclusive binding mode with respect to the allosteric thumb pocket that is occupied by the benzimidazole class of inhibitors. Therefore, halosalicylamides represent a novel class of allosteric inhibitors of HCV NS5B polymerase.
Asunto(s)
Antivirales/síntesis química , Antivirales/farmacología , Hepacivirus/efectos de los fármacos , Salicilamidas/síntesis química , Salicilamidas/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Antivirales/química , Técnicas Químicas Combinatorias , Diseño de Fármacos , Humanos , Estructura Molecular , Salicilamidas/química , Relación Estructura-ActividadRESUMEN
Substituted 1-hydroxy-4,4-dialkyl-3-oxo-3,4-dihydronaphthalene benzothiadiazine derivatives were investigated as inhibitors of genotype 1 HCV polymerase. Structure-activity relationship patterns for this class of compounds are discussed. It was found that the saturated alkane dialkyl units provided the most active analogs.
Asunto(s)
Antivirales/síntesis química , Benzotiadiazinas/síntesis química , Benzotiadiazinas/farmacología , Hepacivirus/efectos de los fármacos , Proteínas no Estructurales Virales/antagonistas & inhibidores , Alcanos , Antivirales/farmacología , Hepacivirus/enzimología , Concentración 50 Inhibidora , Replicón/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
Little is known about the mechanism of HCV polymerase-catalyzed nucleotide incorporation and the individual steps employed by this enzyme during a catalytic cycle. In this paper, we applied various biochemical tools and examined the mechanism of polymerase catalysis. We found that formation of a productive RNA-enzyme complex is the slowest step followed by RNA dissociation and initiation of primer strand synthesis. Various groups have reported several classes of small molecule inhibitors of hepatitis C virus NS5B polymerase; however, the mechanism of inhibition for many of these inhibitors is not clear. We undertook a series of detailed mechanistic studies to characterize the mechanisms of inhibition for these HCV polymerase inhibitors. We found that the diketoacid derivatives competitively bind to the elongation NTP pocket in the active site and inhibit both the initiation and elongation steps of polymerization. While both benzimidazoles and benzothiadiazines are noncompetitive with respect to the active site elongation NTP pocket, benzothiadiazine compounds competitively bind to the initiation pocket in the active site and inhibit only the initiation step of de novo RNA polymerization. The benzimidazoles bind to the thumb allosteric pocket and inhibit the conformational changes during RNA synthesis. We also observed a cross interaction between the thumb allosteric pocket and the initiation pocket using inhibitor-inhibitor cross competition studies. This information will be very important in designing combination therapies using two small molecule drugs to treat hepatitis C virus.
Asunto(s)
Bencimidazoles/farmacología , Benzotiadiazinas/farmacología , Polímeros/metabolismo , Proteínas no Estructurales Virales/antagonistas & inhibidores , Proteínas no Estructurales Virales/metabolismo , Bencimidazoles/química , Benzotiadiazinas/química , Electroforesis , Heparina/metabolismo , Humanos , Cinética , Unión Proteica , Estructura Secundaria de Proteína , ARN Viral/metabolismo , Especificidad por Sustrato , Moldes Genéticos , Factores de Tiempo , Proteínas no Estructurales Virales/químicaRESUMEN
A series of non-nucleoside HCV NS5B polymerase inhibitors based on the N-1-benzyl or N-1-[3-methylbutyl]-4-hydroxy-1,8-naphthyridon-3-yl benzothiadiazine core substituted in the D-ring aromatic moiety have been prepared and evaluated. Aromatic substituents extending from position 7 of the D-ring exhibited excellent potency against both genotypes 1a and 1b.
Asunto(s)
Benzotiadiazinas/farmacología , Inhibidores Enzimáticos/farmacología , ARN Polimerasa Dependiente del ARN/antagonistas & inhibidores , Proteínas no Estructurales Virales/antagonistas & inhibidores , Replicación Viral/efectos de los fármacos , Benzotiadiazinas/síntesis química , Inhibidores Enzimáticos/síntesis química , Genotipo , Hidrocarburos Aromáticos/química , Concentración 50 Inhibidora , Naftiridinas/química , Relación Estructura-ActividadRESUMEN
Substituted N-alkyl-4-hydroxyquinolon-3-yl-benzothiadiazine sulfamides were investigated as inhibitors of genotype 1 HCV polymerase. Structure-activity relationship patterns for this class of compounds are discussed.