RESUMEN
Hepatitis C virus (HCV) NS5B RNA-dependent RNA polymerase (RdRp) plays a central role in virus replication. NS5B has no functional equivalent in mammalian cells, and as a consequence is an attractive target for selective inhibition. This paper describes the discovery of a novel family of HCV NS5B non-nucleoside inhibitors inspired by the bioisosterism between sulfonamide and phosphonamide. Systematic structural optimization in this new series led to the identification of IDX375, a potent non-nucleoside inhibitor that is selective for genotypes 1a and 1b. The structure and binding domain of IDX375 were confirmed by X-ray co-crystalisation study.
Asunto(s)
Antivirales/química , Hepacivirus/enzimología , Lactamas/química , Compuestos Organofosforados/química , Proteínas no Estructurales Virales/antagonistas & inhibidores , Regulación Alostérica , Animales , Antivirales/síntesis química , Antivirales/metabolismo , Antivirales/farmacología , Sitios de Unión , Cristalografía por Rayos X , Genotipo , Semivida , Haplorrinos , Hepacivirus/genética , Hepacivirus/fisiología , Humanos , Lactamas/farmacología , Ratones , Simulación de Dinámica Molecular , Compuestos Organofosforados/farmacología , Estructura Terciaria de Proteína , Ratas , Relación Estructura-Actividad , Sulfonamidas/química , Proteínas no Estructurales Virales/metabolismo , Replicación Viral/efectos de los fármacosRESUMEN
The hepatitis C virus (HCV) NS5B RNA-dependent RNA polymerase (RdRp) plays a central role in virus replication. NS5B has no functional equivalent in mammalian cells and, as a consequence, is an attractive target for selective inhibition. This Letter describes the discovery of a new family of HCV NS5B non-nucleoside inhibitors, based on the bioisosterism between amide and phosphonamidate functions. As part of this program, SAR in this new series led to the identification of IDX17119, a potent non-nucleoside inhibitor, active on the genotypes 1b, 2a, 3a and 4a. The structure and binding domain of IDX17119 were confirmed by X-ray co-crystallization study.
Asunto(s)
Antivirales/farmacología , Genotipo , Hepacivirus/efectos de los fármacos , Proteínas no Estructurales Virales/antagonistas & inhibidores , Sitio Alostérico , Antivirales/química , Antivirales/metabolismo , Cristalografía por Rayos X , Relación Estructura-Actividad , Proteínas no Estructurales Virales/metabolismoRESUMEN
Here, we describe the design, synthesis, biological evaluation, and identification of a clinical candidate non-nucleoside reverse transcriptase inhibitors (NNRTIs) with a novel aryl-phospho-indole (APhI) scaffold. NNRTIs are recommended components of highly active antiretroviral therapy (HAART) for the treatment of HIV-1. Since a major problem associated with NNRTI treatment is the emergence of drug resistant virus, this work focused on optimization of the APhI against clinically relevant HIV-1 Y181C and K103N mutants and the Y181C/K103N double mutant. Optimization of the phosphinate aryl substituent led to the discovery of the 3-Me,5-acrylonitrile-phenyl analogue RP-13s (IDX899) having an EC50 of 11 nM against the Y181C/K103N double mutant.
Asunto(s)
Fármacos Anti-VIH/farmacología , Descubrimiento de Drogas , Transcriptasa Inversa del VIH/antagonistas & inhibidores , VIH-1/efectos de los fármacos , VIH-1/enzimología , Indoles/farmacología , Ácidos Fosfínicos/farmacología , Inhibidores de la Transcriptasa Inversa/farmacología , Animales , Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/química , Línea Celular , Cristalografía por Rayos X , Perros , Relación Dosis-Respuesta a Droga , Transcriptasa Inversa del VIH/metabolismo , Hepatocitos/química , Hepatocitos/metabolismo , Humanos , Indoles/síntesis química , Indoles/química , Macaca fascicularis , Masculino , Modelos Moleculares , Estructura Molecular , Ácidos Fosfínicos/síntesis química , Ácidos Fosfínicos/química , Ratas , Ratas Sprague-Dawley , Inhibidores de la Transcriptasa Inversa/síntesis química , Inhibidores de la Transcriptasa Inversa/química , Relación Estructura-ActividadRESUMEN
BACKGROUND: Ribonucleoside analogs possessing a ß-methyl substituent at the 2'-position of the d-ribose moiety have been previously discovered to be potent and selective inhibitors of hepatitis C virus (HCV) replication, their triphosphates acting as alternative substrate inhibitors of the HCV RdRp NS5B. Results/methodology: In this article, the authors detail the synthesis, anti-HCV evaluation in cell-based replicon assays and structure-activity relationships of several phosphoramidate diester derivatives of 2'-C-methylguanosine (2'-MeG). CONCLUSION: The most promising compound, namely the O-[S-(hydroxyl)pivaloyl-2-thioethyl]{abbreviated as O-[(HO)tBuSATE)]} N-benzylamine phosphoramidate diester derivative (IDX184), was selected for further in vivo studies, and was the first clinical pronucleotide evaluated for the treatment of chronic hepatitis C up to Phase II trials.
Asunto(s)
Antivirales/síntesis química , Antivirales/farmacología , Descubrimiento de Drogas , Guanosina Monofosfato/análogos & derivados , Hepacivirus/efectos de los fármacos , Hepatitis C/tratamiento farmacológico , Guanosina Monofosfato/síntesis química , Guanosina Monofosfato/farmacología , Humanos , Relación Estructura-ActividadRESUMEN
Exploration of the P2 region by mimicking the proline motif found in BILN2061 resulted in the discovery of two series of potent HCV NS3/4A protease inhibitors. X-ray crystal structure of the ligand in contact with the NS3/4A protein and modulation of the quinoline heterocyclic region by structure based design and modeling allowed for the optimization of enzyme potency and cellular activity. This research led to the selection of clinical candidate IDX320 having good genotype coverage and pharmacokinetic properties in various species.
Asunto(s)
Descubrimiento de Drogas , Hepacivirus/efectos de los fármacos , Compuestos Macrocíclicos/química , Compuestos Macrocíclicos/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Animales , Haplorrinos , Hepatocitos/enzimología , Humanos , Concentración 50 Inhibidora , Ratones , Microsomas Hepáticos/enzimología , Estructura Molecular , Ratas , Ratas Sprague-Dawley , Inhibidores de Serina Proteinasa/síntesis química , Inhibidores de Serina Proteinasa/química , Inhibidores de Serina Proteinasa/farmacología , Proteínas no Estructurales Virales/químicaRESUMEN
We disclose here the synthesis of a series of macrocyclic HCV protease inhibitors, where the homoserine linked together the quinoline P2' motif and the macrocyclic moiety. These compounds exhibit potent inhibitory activity against HCV NS3/4A protease and replicon cell based assay. Their enzymatic and antiviral activities are modulated by substitutions on the quinoline P2' at position 8 by methyl and halogens and by small heterocycles at position 2. The in vitro structure activity relationship (SAR) studies and in vivo pharmacokinetic (PK) evaluations of selected compounds are described herein.
Asunto(s)
Antivirales/síntesis química , Antivirales/farmacología , Hepacivirus/efectos de los fármacos , Homoserina/farmacología , Inhibidores de Serina Proteinasa/síntesis química , Inhibidores de Serina Proteinasa/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Antivirales/química , Relación Dosis-Respuesta a Droga , Hepacivirus/enzimología , Homoserina/síntesis química , Homoserina/química , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Inhibidores de Serina Proteinasa/química , Relación Estructura-Actividad , Proteínas no Estructurales Virales/metabolismoRESUMEN
We identified a novel class of 2-((phenylsulfonyl)methyl)-thieno[3,2-d]pyrimidine compounds as potent HIV-1 replication inhibitors serendipitously during the process of evaluation of triazolothienopyrimidine (TTPM) compounds. Herein, we report synthesis and biological evaluation of 2-((phenylsulfonyl)methyl)-thieno[3,2-d]pyrimidine compounds using a cell-based full replication assay to identify thienopyrimidines 6 and 30, which could be further utilized as viable lead compounds.
Asunto(s)
VIH-1/efectos de los fármacos , Pirimidinas/química , Descubrimiento de Drogas , Humanos , Relación Estructura-ActividadRESUMEN
Structural homology between thrombin inhibitors and the early tetrapeptide HCV protease inhibitor led to the bioisosteric replacement of the P2 proline by a 2,4-disubstituted azetidine within the macrocyclic ß-strand mimic. Molecular modeling guided the design of the series. This approach was validated by the excellent activity and selectivity in biochemical and cell based assays of this novel series and confirmed by the co-crystal structure of the inhibitor with the NS3/4A protein (PDB code: 4TYD).
Asunto(s)
Azetidinas/farmacología , Diseño de Fármacos , Inhibidores de Serina Proteinasa/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Azetidinas/síntesis química , Azetidinas/química , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Modelos Moleculares , Estructura Molecular , Inhibidores de Serina Proteinasa/síntesis química , Inhibidores de Serina Proteinasa/química , Relación Estructura-Actividad , Proteínas no Estructurales Virales/metabolismoRESUMEN
New antimalarial agents that exhibit multistage activities against drug-resistant strains of malaria parasites represent good starting points for developing next-generation antimalarial therapies. To facilitate the progression of such agents into the development phase, we developed an image-based parasitological screening method for defining drug effects on different asexual life cycle stages of Plasmodium falciparum. High-throughput screening of a newly assembled diversity-oriented synthetic library using this approach led to the identification of carbohybrid-based 2-aminopyrimidine compounds with fast-acting growth inhibitory activities against three laboratory strains of multidrug-resistant P. falciparum. Our structure-activity relationship study led to the identification of two derivatives (8aA and 11aA) as the most promising antimalarial candidates (mean EC50 of 0.130 and 0.096 µM against all three P. falciparum strains, selectivity indices >600, microsomal stabilities >80%, and mouse malaria ED50 values of 0.32 and 0.12 mg/kg/day, respectively), targeting all major blood stages of multidrug-resistant P. falciparum parasites.
Asunto(s)
Antimaláricos/farmacología , Estadios del Ciclo de Vida/efectos de los fármacos , Plasmodium falciparum/efectos de los fármacos , Pirimidinas/farmacología , Animales , Antimaláricos/química , Antimaláricos/farmacocinética , Área Bajo la Curva , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Descubrimiento de Drogas , Evaluación Preclínica de Medicamentos , Células Hep G2 , Interacciones Huésped-Parásitos/efectos de los fármacos , Humanos , Malaria/parasitología , Malaria/prevención & control , Masculino , Tasa de Depuración Metabólica , Ratones , Ratones Endogámicos BALB C , Modelos Químicos , Estructura Molecular , Plasmodium chabaudi/efectos de los fármacos , Plasmodium chabaudi/fisiología , Plasmodium falciparum/crecimiento & desarrollo , Pirimidinas/química , Pirimidinas/farmacocinética , Ratas , Ratas Sprague-Dawley , Relación Estructura-ActividadRESUMEN
Nefang, a polyherbal product composed of Mangifera indica (bark and leaf), Psidium guajava, Carica papaya, Cymbopogon citratus, Citrus sinensis, and Ocimum gratissimum (leaves), is a potential therapy against P. falciparum malaria. In vitro antiplasmodial activities of its constituent solvent extracts were analyzed on CQ-sensitive (3D7) and multidrug resistant (Dd2) P. falciparum strains. The interactions involving the differential solvent extracts were further analyzed using a variable potency ratio drug combination approach. Effective concentration 50 (EC50) values were determined by nonlinear regression curve-fitting of the dose-response data and used in calculating the fractional inhibitory concentration 50 (FIC50) and combination indices (CI) for each pair. The derived EC50 values (3D7/Dd2, µ g/mL) are Nefang-96.96/55.08, MiB-65.33/34.58, MiL-82.56/40.04, Pg-47.02/25.79, Cp-1188/317.5, Cc-723.3/141, Cs-184.4/105.1, and Og-778.5/118.9. Synergism was obtained with MiB/Pg (CI = 0.351), MiL/Pg (0.358), MiB/Cs (0.366), MiL/Cs (0.482), Pg/Cs (0.483), and Cs/Og (0.414) when analyzed at equipotency ratios. Cytotoxicity testing of Nefang and the solvent extracts on two human cell lines (Hep G2 and U2OS) revealed no significant toxicity relative to their antiplasmodial activities (SI > 20). Taken together, our data confirm the antimalarial activities of Nefang and its constituent plant extracts and identified extract pairs with promising synergistic interactions for exploitation towards a rational phytotherapeutic and evidence-based antimalarial drug discovery.
Asunto(s)
Antimaláricos , Malaria Falciparum/tratamiento farmacológico , Extractos Vegetales , Plasmodium falciparum , Solventes/química , Antimaláricos/química , Antimaláricos/farmacología , Evaluación Preclínica de Medicamentos , Células Hep G2 , Humanos , Extractos Vegetales/química , Extractos Vegetales/farmacologíaRESUMEN
Dengue virus is a mosquito-borne flavivirus that has a large impact in global health. It is considered as one of the medically important arboviruses, and developing a preventive or therapeutic solution remains a top priority in the medical and scientific community. Drug discovery programs for potential dengue antivirals have increased dramatically over the last decade, largely in part to the introduction of high-throughput assays. In this study, we have developed an image-based dengue high-throughput/high-content assay (HT/HCA) using an innovative computer vision approach to screen a kinase-focused library for anti-dengue compounds. Using this dengue HT/HCA, we identified a group of compounds with a 4-(1-aminoethyl)-N-methylthiazol-2-amine as a common core structure that inhibits dengue viral infection in a human liver-derived cell line (Huh-7.5 cells). Compounds CND1201, CND1203 and CND1243 exhibited strong antiviral activities against all four dengue serotypes. Plaque reduction and time-of-addition assays suggests that these compounds interfere with the late stage of viral infection cycle. These findings demonstrate that our image-based dengue HT/HCA is a reliable tool that can be used to screen various chemical libraries for potential dengue antiviral candidates.
Asunto(s)
Antivirales/aislamiento & purificación , Antivirales/farmacología , Virus del Dengue/efectos de los fármacos , Descubrimiento de Drogas/métodos , Ensayos Analíticos de Alto Rendimiento , Antivirales/química , Línea Celular , Hepatocitos/virología , Humanos , Pruebas de Sensibilidad MicrobianaRESUMEN
Microarray technology has become a very popular approach in cases where multiple experiments need to be conducted repeatedly or done with a variety of samples. In our lab, we are applying our high density spots microarray approach to microscopy visualization of the effects of transiently introduced siRNA or cDNA on cellular morphology or phenotype. In this publication, we are discussing the possibility of using this micro-scale high throughput process to study the role of microRNAs in the biology of selected cellular models. After reverse-transfection of microRNAs and siRNA, the cellular phenotype generated by microRNAs regulated NF-κB expression comparably to the siRNA. The ability to print microRNA molecules for reverse transfection into cells is opening up the wide horizon for the phenotypic high content screening of microRNA libraries using cellular disease models.
RESUMEN
We identified a novel class of triazolothienopyrimidine (TTPM) compounds as potent HIV-1 replication inhibitors during a high-throughput screening campaign that evaluated more than 200,000 compounds using a cell-based full replication assay. Herein, we report the optimization of the antiviral activity in a cell-based assay system leading to the discovery of aryl-substituted TTPM derivatives (38, 44, and 45), which exhibited significant inhibition of HIV-1 replication with acceptable safety margins. These novel and potent TTPMs could serve as leads for further development.
Asunto(s)
Fármacos Anti-VIH/síntesis química , VIH-1/metabolismo , Pirimidinas/química , Triazoles/química , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Línea Celular , Evaluación Preclínica de Medicamentos , VIH-1/efectos de los fármacos , Ensayos Analíticos de Alto Rendimiento , Humanos , Pirimidinas/síntesis química , Pirimidinas/farmacología , Relación Estructura-Actividad , Replicación Viral/efectos de los fármacosRESUMEN
We describe a novel 7-aminopyrazolo[1,5-a]pyrimidine (7-APP) derivative as a potent hepatitis C virus (HCV) inhibitor. A series of 7-APPs was synthesized and evaluated for inhibitory activity against HCV in different cell culture systems. The synthesis and preliminary structure-activity relationship study of 7-APP are reported.
Asunto(s)
Antivirales/farmacología , Descubrimiento de Drogas , Hepacivirus/efectos de los fármacos , Pirazoles/farmacología , Pirimidinas/farmacología , Animales , Antivirales/química , Antivirales/metabolismo , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Estabilidad de Medicamentos , Humanos , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Pirazoles/química , Pirazoles/metabolismo , Pirimidinas/química , Pirimidinas/metabolismo , Ratas , Relación Estructura-ActividadRESUMEN
Following the previous SAR of a novel dihydropyrimidinone scaffold as HIV-1 replication inhibitors a detailed study directed towards optimizing the metabolic stability of the ester functional group in the dihydropyrimidinone (DHPM) scaffold is described. Replacement of the ester moiety by thiazole ring significantly improved the metabolic stability while retaining antiviral activity against HIV-1 replication. These novel and potent DHPMs with bioisosteres could serve as advanced leads for further optimization.
Asunto(s)
Transcriptasa Inversa del VIH/antagonistas & inhibidores , VIH-1/efectos de los fármacos , Pirimidinonas/síntesis química , Inhibidores de la Transcriptasa Inversa/síntesis química , Replicación Viral/efectos de los fármacos , Animales , Línea Celular Tumoral , Estabilidad de Medicamentos , VIH-1/fisiología , Humanos , Microsomas Hepáticos/metabolismo , Modelos Moleculares , Nevirapina/farmacología , Pirimidinonas/farmacología , Ratas , Inhibidores de la Transcriptasa Inversa/farmacología , Relación Estructura-Actividad , Tiazoles/químicaRESUMEN
We identified a novel class of aryl-substituted triazine compounds as potent non-nucleoside reverse transcriptase inhibitors (NNRTIs) during a high-throughput screening campaign that evaluated more than 200000 compounds for antihuman immunodeficiency virus (HIV) activity using a cell-based full replication assay. Herein, we disclose the optimization of the antiviral activity in a cell-based assay system leading to the discovery of compound 27, which possessed excellent potency against wild-type HIV-1 (EC50 = 0.2 nM) as well as viruses bearing Y181C and K103N resistance mutations in the reverse transcriptase gene. The X-ray crystal structure of compound 27 complexed with wild-type reverse transcriptase confirmed the mode of action of this novel class of NNRTIs. Introduction of a chloro functional group in the pyrazole moiety dramatically improved hERG and CYP inhibition profiles, yielding highly promising leads for further development.
RESUMEN
A novel series of 3-aryl-phospho-indole (API) non-nucleoside reverse transcriptase inhibitors of HIV-1 was developed. Chemical variation in the phosphorus linker led to the discovery of 3-phenyl-methyl-phosphinate-2-carboxamide 14, which possessed excellent potency against wild-type HIV-1 as well as viruses bearing K103N and Y181C single mutants in the reverse transcriptase gene. Chiral separation of the enantiomers showed that only R enantiomer retained the activity. The pharmacokinetic, solubility, and metabolic properties of 14 were assessed.
Asunto(s)
Fármacos Anti-VIH/síntesis química , Transcriptasa Inversa del VIH/metabolismo , Indoles/síntesis química , Ácidos Fosfínicos/síntesis química , Inhibidores de la Transcriptasa Inversa/síntesis química , Animales , Fármacos Anti-VIH/farmacocinética , Fármacos Anti-VIH/farmacología , Línea Celular , Perros , Farmacorresistencia Viral , Transcriptasa Inversa del VIH/genética , Haplorrinos , Hepatocitos/metabolismo , Humanos , Indoles/farmacocinética , Indoles/farmacología , Modelos Moleculares , Mutación , Ácidos Fosfínicos/farmacocinética , Ácidos Fosfínicos/farmacología , Ratas , Inhibidores de la Transcriptasa Inversa/farmacocinética , Inhibidores de la Transcriptasa Inversa/farmacología , Solubilidad , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
An SAR study that identified a series of thienopyridine-based potent IkappaB Kinase beta (IKKbeta) inhibitors is described. With focuses on the structural optimization at C4 and C6 of structure 1 (Fig. 1), the study reveals that small alkyl and certain aromatic groups are preferred at C4, whereas polar groups with proper orientation at C6 efficiently enhance compound potency. The most potent analogues inhibit IKKbeta with IC50s as low as 40 nM, suppress LPS-induced TNF-alpha production in vitro and in vivo, display good kinase selectivity profiles, and are active in a HeLa cell NF-kappaB reporter gene assay, demonstrating that they directly interfere with the NF-kappaB signaling pathway.
Asunto(s)
Quinasa I-kappa B/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Piridinas/química , Animales , Descubrimiento de Drogas , Células HeLa , Humanos , Quinasa I-kappa B/metabolismo , Lipopolisacáridos/farmacología , Ratones , FN-kappa B/metabolismo , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/síntesis química , Piridinas/farmacología , Transducción de Señal , Relación Estructura-Actividad , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
A functional screen of an adenovirus-delivered shRNA library that targets approximately 4500 host genes was performed to identify cellular factors that regulate hepatitis C virus (HCV) sub-genomic RNA replication. Seventy-three hits were further examined by siRNA oligonucleotide-directed knockdown, and silencing of the PI4KA gene was demonstrated to have a significant effect on the replication of a HCV genotype 1b replicon. Using transient siRNA oligonucleotide transfections and stable shRNA knockdown clones in HuH-7 cells, the PI4KA gene was shown to be essential for the replication of all HCV genotypes tested (1a, 1b and 2a) but not required for bovine viral diarrhea virus (BVDV) RNA replication.
Asunto(s)
Hepacivirus/fisiología , Hepatitis C/enzimología , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , ARN Viral/genética , Replicación Viral/genética , Adenoviridae/genética , Adenoviridae/metabolismo , Adenoviridae/fisiología , Línea Celular Tumoral , Regulación Viral de la Expresión Génica , Técnicas de Silenciamiento del Gen , Biblioteca de Genes , Genoma Viral , Hepacivirus/genética , Hepacivirus/crecimiento & desarrollo , Hepacivirus/metabolismo , Humanos , Antígenos de Histocompatibilidad Menor , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , ARN Interferente Pequeño/metabolismo , Reproducibilidad de los ResultadosRESUMEN
Several thieno[3,4-d]pyrimidine derivatives, including four hitherto unknown 2',3'-dideoxy- and 2',3'-dideoxy-2',3'-didehydro-C-nucleoside analogues of adenosine and inosine have been synthesized. When evaluated in cell culture experiments against human immunodeficiency virus, none of the tested compounds exhibited any significant antiviral effect, while two of them showed some cytotoxicity.