RESUMEN
Data from both our own and literature studies of the biochemistry and inhibition of influenza virus endonuclease was combined with data on the mechanism of action and the likely active site mechanism to propose a pharmacophore. The pharmacophore was used to design a novel structural class of inhibitors, some of which were found to have activities similar to that of known influenza endonuclease inhibitors and were also antiviral in cell culture.
Asunto(s)
Antivirales/síntesis química , Endonucleasas/antagonistas & inhibidores , Inhibidores Enzimáticos/síntesis química , Animales , Antivirales/química , Antivirales/farmacología , Sitios de Unión , Butiratos/síntesis química , Butiratos/química , Butiratos/farmacología , Línea Celular , Técnicas Químicas Combinatorias , ARN Polimerasas Dirigidas por ADN/química , Bases de Datos Factuales , Perros , Endonucleasas/química , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Compuestos Heterocíclicos con 3 Anillos/síntesis química , Compuestos Heterocíclicos con 3 Anillos/química , Compuestos Heterocíclicos con 3 Anillos/farmacología , Compuestos Heterocíclicos de 4 o más Anillos/síntesis química , Compuestos Heterocíclicos de 4 o más Anillos/química , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Virus de la Influenza A/efectos de los fármacos , Virus de la Influenza A/enzimología , Virus de la Influenza B/efectos de los fármacos , Virus de la Influenza B/enzimología , Cetoácidos/síntesis química , Cetoácidos/química , Cetoácidos/farmacología , Modelos Moleculares , Ftalimidas/síntesis química , Ftalimidas/química , Ftalimidas/farmacología , Unión Proteica , Relación Estructura-ActividadRESUMEN
A series of 4'-substituted ribonucleoside derivatives has been prepared and evaluated for inhibition of hepatitis C virus (HCV) RNA replication in cell culture. The most potent and non-cytotoxic derivative was compound 28 (4'-azidocytidine, R1479) with an IC(50) of 1.28 microM in the HCV replicon system. The triphosphate of compound 28 was prepared and shown to be an inhibitor of RNA synthesis mediated by NS5B (IC(50)=320 nM), the RNA polymerase encoded by HCV. Data on related analogues have been used to generate some preliminary requirements for activity within this series of nucleosides.
Asunto(s)
Antivirales/química , Química Farmacéutica/métodos , Citidina/análogos & derivados , Hepacivirus/genética , Ribonucleósidos/química , Replicación Viral/efectos de los fármacos , Citidina/farmacología , Diseño de Fármacos , Evaluación Preclínica de Medicamentos , Concentración 50 Inhibidora , Modelos Químicos , Conformación Molecular , Nucleósidos/química , ARN/química , UridinaRESUMEN
Hepatitis C virus (HCV) polymerase activity is essential for HCV replication. Targeted screening of nucleoside analogs identified R1479 (4'-azidocytidine) as a specific inhibitor of HCV replication in the HCV subgenomic replicon system (IC(50) = 1.28 microM) with similar potency compared with 2'-C-methylcytidine (IC(50) = 1.13 microM). R1479 showed no effect on cell viability or proliferation of HCV replicon or Huh-7 cells at concentrations up to 2 mM. HCV replicon RNA could be fully cleared from replicon cells after prolonged incubation with R1479. The corresponding 5'-triphosphate derivative (R1479-TP) is a potent inhibitor of native HCV replicase isolated from replicon cells and of recombinant HCV polymerase (NS5B)-mediated RNA synthesis activity. R1479-TP inhibited RNA synthesis as a CTP-competitive inhibitor with a K(i) of 40 nM. On an HCV RNA-derived template substrate (complementary internal ribosome entry site), R1479-TP showed similar potency of NS5B inhibition compared with 3'-dCTP. R1479-TP was incorporated into nascent RNA by HCV polymerase and reduced further elongation with similar efficiency compared with 3'-dCTP under the reaction conditions. The S282T point mutation in the coding sequence of NS5B confers resistance to inhibition by 2'-C-MeATP and other 2'-methyl-nucleotides. In contrast, the S282T mutation did not confer cross-resistance to R1479.