RESUMEN
An assay recapitulating the 3' processing activity of HIV-1 integrase (IN) was used to screen the Boehringer Ingelheim compound collection. Hit-to-lead and lead optimization beginning with compound 1 established the importance of the C3 and C4 substituent to antiviral potency against viruses with different aa124/aa125 variants of IN. The importance of the C7 position on the serum shifted potency was established. Introduction of a quinoline substituent at the C4 position provided a balance of potency and metabolic stability. Combination of these findings ultimately led to the discovery of compound 26 (BI 224436), the first NCINI to advance into a phase Ia clinical trial.
RESUMEN
BI 224436 is an HIV-1 integrase inhibitor with effective antiviral activity that acts through a mechanism that is distinct from that of integrase strand transfer inhibitors (INSTIs). This 3-quinolineacetic acid derivative series was identified using an enzymatic integrase long terminal repeat (LTR) DNA 3'-processing assay. A combination of medicinal chemistry, parallel synthesis, and structure-guided drug design led to the identification of BI 224436 as a candidate for preclinical profiling. It has antiviral 50% effective concentrations (EC50s) of <15 nM against different HIV-1 laboratory strains and cellular cytotoxicity of >90 µM. BI 224436 also has a low, â¼2.1-fold decrease in antiviral potency in the presence of 50% human serum and, by virtue of a steep dose-response curve slope, exhibits serum-shifted EC95 values ranging between 22 and 75 nM. Passage of virus in the presence of inhibitor selected for either A128T, A128N, or L102F primary resistance substitutions, all mapping to a conserved allosteric pocket on the catalytic core of integrase. BI 224436 also retains full antiviral activity against recombinant viruses encoding INSTI resistance substitutions N155S, Q148H, and E92Q. In drug combination studies performed in cellular antiviral assays, BI 224436 displays an additive effect in combination with most approved antiretrovirals, including INSTIs. BI 224436 has drug-like in vitro absorption, distribution, metabolism, and excretion (ADME) properties, including Caco-2 cell permeability, solubility, and low cytochrome P450 inhibition. It exhibited excellent pharmacokinetic profiles in rat (clearance as a percentage of hepatic flow [CL], 0.7%; bioavailability [F], 54%), monkey (CL, 23%; F, 82%), and dog (CL, 8%; F, 81%). Based on the excellent biological and pharmacokinetic profile, BI 224436 was advanced into phase 1 clinical trials.
Asunto(s)
Inhibidores de Integrasa VIH/farmacología , VIH-1/efectos de los fármacos , VIH-1/enzimología , Sustitución de Aminoácidos/genética , Sustitución de Aminoácidos/fisiología , Animales , Fármacos Anti-VIH/farmacología , Células CACO-2 , Clonación Molecular , Inhibidores Enzimáticos del Citocromo P-450/farmacología , ADN Viral/efectos de los fármacos , Farmacorresistencia Viral , Integrasa de VIH/biosíntesis , Integrasa de VIH/genética , Integrasa de VIH/metabolismo , Inhibidores de Integrasa VIH/metabolismo , Inhibidores de Integrasa VIH/farmacocinética , Hepatocitos/metabolismo , Humanos , Ratones , Ratas , Suero/virología , Replicación Viral/efectos de los fármacosRESUMEN
Despite truly impressive achievements in the global battle against HIV there remains a need for new drugs directed against novel targets, and the viral capsid protein (CA) may represent one such target. Intense structural characterization of CA over the last two decades has provided unprecedented insight into the structure and assembly of this key viral protein. Furthermore, several inhibitor-binding sites that elicit antiviral activity have been reported on CA, two of which are located on its N-terminal domain (CANTD). In this work, the binding of a novel capsid-assembly inhibitor that targets a unique inhibitory site on CANTD is reported. Moreover, whereas cocrystallization of CANTD in complex with ligands has proven to be challenging in the past, the use of this inhibitor as a tool compound is shown to vastly facilitate ternary cocrystallizations with CANTD. This improvement in crystallization is likely to be achieved through the formation of a compound-mediated homodimer, the intrinsic symmetry of which greatly increases the prospect of generating a crystal lattice. While protein engineering has been used in the literature to support a link between the inherent symmetry of a macromolecule and its propensity to crystallize, to our knowledge this work represents the first use of a synthetic ligand for this purpose.
Asunto(s)
Antivirales/química , Proteínas de la Cápside/química , Cápside/química , VIH-1/química , Antivirales/metabolismo , Sitios de Unión , Cápside/metabolismo , Proteínas de la Cápside/antagonistas & inhibidores , Proteínas de la Cápside/metabolismo , Cristalización , VIH-1/metabolismo , Modelos Moleculares , Difracción de Rayos XRESUMEN
The HIV-1 capsid (CA) protein, a domain of Gag, which participates in formation of both the mature and immature capsid, represents a potential target for anti-viral drug development. Characterization of hits obtained via high-throughput screening of an in vitro capsid assembly assay led to multiple compounds having this potential. We previously presented the characterization of two inhibitor series that bind the N-terminal domain of the capsid (CA(NTD)), at a site located at the bottom of its helical bundle, often referred to as the CAP-1 binding site. In this work we characterize a novel series of benzimidazole hits. Initial optimization of this series led to compounds with improved in vitro assembly and anti-viral activity. Using NMR spectroscopy we found that this series binds to a unique site on CA(NTD), located at the apex of the helical bundle, well removed from previously characterized binding sites for CA inhibitors. 2D (1)H-(15)N HSQC and (19)F NMR showed that binding of the benzimidazoles to this distinct site does not affect the binding of either cyclophilin A (CypA) to the CypA-binding loop or a benzodiazepine-based CA assembly inhibitor to the CAP-1 site. Unfortunately, while compounds of this series achieved promising in vitro assembly and anti-viral effects, they also were found to be quite sensitive to a number of naturally occurring CA(NTD) polymorphisms observed among clinical isolates. Despite the negative impact of this finding for drug development, the discovery of multiple inhibitor binding sites on CA(NTD) shows that capsid assembly is much more complex than previously realized.
Asunto(s)
Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Proteínas de la Cápside/antagonistas & inhibidores , Proteínas de la Cápside/química , Proteínas de la Cápside/metabolismo , VIH-1 , Fármacos Anti-VIH/metabolismo , Bencimidazoles/química , Sitios de Unión , Unión Competitiva , Cristalografía por Rayos X , Ciclofilina A/metabolismo , Ciclofilina A/farmacología , VIH-1/genética , VIH-1/aislamiento & purificación , Espectroscopía de Resonancia Magnética , Polimorfismo Genético , Conformación Proteica , Relación Estructura-ActividadRESUMEN
The nucleocapsid (NC) protein is an essential factor with multiple functions within the human immunodeficiency virus type 1 (HIV-1) replication cycle. In this study, we describe the discovery of a novel series of inhibitors that targets HIV-1 NC protein by blocking its interaction with nucleic acids. This series was identified using a previously described capsid (CA) assembly assay, employing a recombinant HIV-1 CA-NC protein and immobilized TG-rich deoxyoligonucleotides. Using visible absorption spectroscopy, we were able to demonstrate that this new inhibitor series binds specifically and reversibly to the NC with a peculiar 2:1 stoichiometry. A fluorescence-polarization-based binding assay was also developed in order to monitor the inhibitory activities of this series of inhibitors. To better characterize the structural aspect of inhibitor binding onto NC, we performed NMR studies using unlabeled and (13)C,(15)N-double-labeled NC(1-55) protein constructs. This allowed the determination of the solution structure of a ternary complex characterized by two inhibitor molecules binding to the two zinc knuckles of the NC protein. To the best of our knowledge, this represents the first report of a high-resolution structure of a small-molecule inhibitor bound to NC, demonstrating sub-micromolar potency and moderate antiviral potency with one analogue of the series. This structure was compared with available NC/oligonucleotide complex structures and further underlined the high flexibility of the NC protein, allowing it to adopt many conformations in order to bind its different oligonucleotide/nucleomimetic targets. In addition, analysis of the interaction details between the inhibitor molecules and NC demonstrated how this novel inhibitor series is mimicking the guanosine nucleobases found in many reported complex structures.
Asunto(s)
Fármacos Anti-VIH/aislamiento & purificación , Fármacos Anti-VIH/metabolismo , VIH-1/efectos de los fármacos , Productos del Gen gag del Virus de la Inmunodeficiencia Humana/química , Productos del Gen gag del Virus de la Inmunodeficiencia Humana/metabolismo , Fármacos Anti-VIH/química , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Estructura Molecular , Unión Proteica , Conformación Proteica , Productos del Gen gag del Virus de la Inmunodeficiencia Humana/antagonistas & inhibidoresRESUMEN
Screening of our sample collection led to the identification of a set of benzofurano[3,2-d]pyrimidine-2-one hits acting as nucleotide-competing HIV-1 reverse transcriptase inhibitiors (NcRTI). Significant improvement in antiviral potency was achieved when substituents were introduced at positions N1, C4, C7 and C8 on the benzofuranopyrimidone scaffold. The series was optimized from low micromolar enzymatic activity against HIV-1 RT and no antiviral activity to low nanomolar antiviral potency. Further profiling of inhibitor 30 showed promising overall in vitro properties and also demonstrated that its potency was maintained against viruses resistant to the other major classes of HIV-1 RT inhibitors.
Asunto(s)
Benzofuranos/química , Transcriptasa Inversa del VIH/antagonistas & inhibidores , Nucleótidos/química , Pirimidinonas/química , Inhibidores de la Transcriptasa Inversa/química , Animales , Transcriptasa Inversa del VIH/metabolismo , VIH-1/efectos de los fármacos , VIH-1/enzimología , Humanos , Microsomas Hepáticos/metabolismo , Nucleótidos/metabolismo , Unión Proteica , Pirimidinonas/síntesis química , Pirimidinonas/farmacología , Ratas , Inhibidores de la Transcriptasa Inversa/síntesis química , Inhibidores de la Transcriptasa Inversa/farmacología , Relación Estructura-ActividadRESUMEN
The emergence of resistance to existing classes of antiretroviral drugs underlines the need to find novel human immunodeficiency virus (HIV)-1 targets for drug discovery. The viral capsid protein (CA) represents one such potential target. Recently, a series of benzodiazepine inhibitors was identified via high-throughput screening using an in vitro capsid assembly assay (CAA). Here, we demonstrate how a combination of NMR and X-ray co-crystallography allowed for the rapid characterization of the early hits from this inhibitor series. Ligand-based (19)F NMR was used to confirm inhibitor binding specificity and reversibility as well as to identify the N-terminal domain of the capsid (CA(NTD)) as its molecular target. Protein-based NMR ((1)H and (15)N chemical shift perturbation analysis) identified key residues within the CA(NTD) involved in inhibitor binding, while X-ray co-crystallography confirmed the inhibitor binding site and its binding mode. Based on these results, two conformationally restricted cyclic inhibitors were designed to further validate the possible binding modes. These studies were crucial to early hit confirmation and subsequent lead optimization.
Asunto(s)
Benzodiazepinas/metabolismo , Proteínas de la Cápside/metabolismo , VIH-1/metabolismo , Benzodiazepinas/química , Sitios de Unión , Proteínas de la Cápside/química , Cristalografía por Rayos X , Flúor/química , Humanos , Ligandos , Espectroscopía de Resonancia Magnética , Isótopos de Nitrógeno/química , Unión Proteica , Estructura Terciaria de ProteínaRESUMEN
Phosphatidylinositol-4-kinase IIIα (PI4KIIIα) is an essential host cell factor for hepatitis C virus (HCV) replication. An N-terminally truncated 130-kDa form was used to reconstitute an in vitro biochemical lipid kinase assay that was optimized for small-molecule compound screening and identified potent and specific inhibitors. Cell culture studies with PI4KIIIα inhibitors demonstrated that the kinase activity was essential for HCV RNA replication. Two PI4KIIIα inhibitors were used to select cell lines harboring HCV replicon mutants with a 20-fold loss in sensitivity to the compounds. Reverse genetic mapping isolated an NS4B-NS5A segment that rescued HCV RNA replication in PIK4IIIα-deficient cells. HCV RNA replication occurs on specialized membranous webs, and this study with PIK4IIIα inhibitor-resistant mutants provides a genetic link between NS4B/NS5A functions and PI4-phosphate lipid metabolism. A comprehensive assessment of PI4KIIIα as a drug target included its evaluation for pharmacologic intervention in vivo through conditional transgenic murine lines that mimic target-specific inhibition in adult mice. Homozygotes that induce a knockout of the kinase domain or knock in a single amino acid substitution, kinase-defective PI4KIIIα, displayed a lethal phenotype with a fairly widespread mucosal epithelial degeneration of the gastrointestinal tract. This essential host physiologic role raises doubt about the pursuit of PI4KIIIα inhibitors for treatment of chronic HCV infection.
Asunto(s)
1-Fosfatidilinositol 4-Quinasa/metabolismo , Hepacivirus/fisiología , Interacciones Huésped-Patógeno , Replicación Viral , 1-Fosfatidilinositol 4-Quinasa/antagonistas & inhibidores , Animales , Antivirales/farmacología , Línea Celular , Análisis Mutacional de ADN , Farmacorresistencia Viral , Inhibidores Enzimáticos/farmacología , Femenino , Genes Esenciales , Hepatocitos/enzimología , Hepatocitos/virología , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Mutantes/genética , Proteínas no Estructurales Virales/genéticaRESUMEN
The emergence of resistance to existing classes of antiretroviral drugs necessitates finding new HIV-1 targets for drug discovery. The viral capsid (CA) protein represents one such potential new target. CA is sufficient to form mature HIV-1 capsids in vitro, and extensive structure-function and mutational analyses of CA have shown that the proper assembly, morphology, and stability of the mature capsid core are essential for the infectivity of HIV-1 virions. Here we describe the development of an in vitro capsid assembly assay based on the association of CA-NC subunits on immobilized oligonucleotides. This assay was used to screen a compound library, yielding several different families of compounds that inhibited capsid assembly. Optimization of two chemical series, termed the benzodiazepines (BD) and the benzimidazoles (BM), resulted in compounds with potent antiviral activity against wild-type and drug-resistant HIV-1. Nuclear magnetic resonance (NMR) spectroscopic and X-ray crystallographic analyses showed that both series of inhibitors bound to the N-terminal domain of CA. These inhibitors induce the formation of a pocket that overlaps with the binding site for the previously reported CAP inhibitors but is expanded significantly by these new, more potent CA inhibitors. Virus release and electron microscopic (EM) studies showed that the BD compounds prevented virion release, whereas the BM compounds inhibited the formation of the mature capsid. Passage of virus in the presence of the inhibitors selected for resistance mutations that mapped to highly conserved residues surrounding the inhibitor binding pocket, but also to the C-terminal domain of CA. The resistance mutations selected by the two series differed, consistent with differences in their interactions within the pocket, and most also impaired virus replicative capacity. Resistance mutations had two modes of action, either directly impacting inhibitor binding affinity or apparently increasing the overall stability of the viral capsid without affecting inhibitor binding. These studies demonstrate that CA is a viable antiviral target and demonstrate that inhibitors that bind within the same site on CA can have distinct binding modes and mechanisms of action.
Asunto(s)
Fármacos Anti-VIH/farmacología , Cápside/efectos de los fármacos , Productos del Gen gag/antagonistas & inhibidores , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , Bencimidazoles/farmacología , Benzodiazepinas/farmacología , Cápside/metabolismo , Línea Celular , Productos del Gen gag/química , Productos del Gen gag/genética , Productos del Gen gag/metabolismo , Infecciones por VIH/tratamiento farmacológico , VIH-1/química , VIH-1/genética , VIH-1/fisiología , Humanos , Estructura Terciaria de Proteína , Ensamble de Virus/efectos de los fármacosRESUMEN
Human papillomaviruses are responsible for multiple human diseases, including cervical cancer caused by multiple high-risk types and genital warts caused by the low-risk types 6 and 11. Based on the research indicating that low-risk HPV could be successfully targeted by inhibitors of viral DNA replication, we carried out several high-throughput screens for inhibitors of DNA replication activities. Two series were identified in screens for inhibitors of the interaction between the viral proteins E1 and E2. The two series were demonstrated to bind to overlapping sites on the transactivation domain of E2, at the E1-binding interface, by a series of biochemical and biophysical experiments. A member of the first series was also cocrystallized with the E2 transactivation domain. For both series, structure-activity investigations are described, which resulted in several hundred fold improvements in activity. The best compounds in each series had low nanomolar activity against the HPV11 E1-E2 interaction, and EC(50) values in cellular DNA replication assays of approximately 1 µM. Binding modes for the two series are compared, and some general conclusions about the discovery of protein-protein interaction inhibitors are drawn from the work described.
Asunto(s)
Carbamatos , Indanos , Papillomaviridae , Piperidinas , Proteínas Virales/antagonistas & inhibidores , Proteínas Virales/química , Antivirales/química , Antivirales/metabolismo , Antivirales/farmacología , Sitios de Unión , Carbamatos/química , Carbamatos/metabolismo , Carbamatos/farmacología , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Evaluación Preclínica de Medicamentos/métodos , Humanos , Indanos/química , Indanos/metabolismo , Indanos/farmacología , Simulación de Dinámica Molecular , Papillomaviridae/genética , Papillomaviridae/metabolismo , Piperidinas/química , Piperidinas/metabolismo , Piperidinas/farmacología , Unión Proteica/efectos de los fármacos , Conformación Proteica , Proteínas Virales/metabolismoRESUMEN
The discovery of a 1,5-dihydrobenzo[b][1,4]diazepine-2,4-dione series of inhibitors of HIV-1 capsid assembly is described. Synthesis of analogs of the 1,5-dihydrobenzo[b][1,4]diazepine-2,4-dione hit established structure-activity relationships. Replacement of the enamine functionality of the hit series with either an imidazole or a pyrazole ring led to compounds that inhibited both capsid assembly and reverse transcriptase. Optimization of the bicyclic benzodiazepine scaffold to include a 3-phenyl substituent led to lead compound 48, a pure capsid assembly inhibitor with improved antiviral activity.
Asunto(s)
Fármacos Anti-VIH/química , Benzodiazepinonas/química , Proteínas de la Cápside/antagonistas & inhibidores , VIH-1/efectos de los fármacos , Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/farmacología , Benzodiazepinonas/síntesis química , Benzodiazepinonas/farmacología , Proteínas de la Cápside/metabolismo , Evaluación Preclínica de Medicamentos , Transcriptasa Inversa del VIH/antagonistas & inhibidores , Transcriptasa Inversa del VIH/metabolismo , Humanos , Imidazoles/química , Pirazoles/química , Inhibidores de la Transcriptasa Inversa/síntesis química , Inhibidores de la Transcriptasa Inversa/química , Inhibidores de la Transcriptasa Inversa/farmacología , Relación Estructura-ActividadRESUMEN
High-throughput screening hit 1 was identified as a potent, broad-spectrum, non-nucleoside reverse transcriptase inhibitor (NNRTI) of HIV-1 replication. Analysis of the bound conformation of analogs of this inhibitor via molecular modeling and NMR contributed to the design of novel tertiary amide, carbamate, and thiocarbamate based NNRTIs.
Asunto(s)
Fármacos Anti-VIH/farmacología , Transcriptasa Inversa del VIH/antagonistas & inhibidores , Inhibidores de la Transcriptasa Inversa/farmacología , Replicación Viral/efectos de los fármacos , Amidas/química , Fármacos Anti-VIH/síntesis química , Carbamatos/química , Diseño de Fármacos , Farmacorresistencia Viral , Humanos , Concentración 50 Inhibidora , Espectroscopía de Resonancia Magnética , Modelos Químicos , Inhibidores de la Transcriptasa Inversa/síntesis química , Relación Estructura-Actividad , Tiocarbamatos/químicaRESUMEN
Human papillomaviruses (HPVs) are the causative agents of benign and malignant lesions of the epithelium. Despite their high prevalence, there is currently no antiviral drug for the treatment of HPV-induced lesions. The ATPase and helicase activities of the highly conserved E1 protein of HPV are essential for viral DNA replication and pathogenesis and hence are considered valid antiviral targets. We recently described novel biphenylsulfonacetic acid inhibitors of the ATPase activity of E1 from HPV type 6 (HPV6). Based on kinetics and mutagenesis studies, we now report that these compounds act by an allosteric mechanism. They are hyperbolic competitive inhibitors of the ATPase activity of HPV6 E1 and also inhibit its helicase activity. Compounds in this series can also inhibit the ATPase activity of the closely related enzyme from HPV11; however, the most potent inhibitors of HPV6 E1 are significantly less active against the type 11 protein. We identified a single critical residue in HPV6 E1, Tyr-486, substituted by a cysteine in HPV11, which is primarily responsible for this difference in inhibitor potency. Interestingly, HPV18 E1, which also has a tyrosine at this position, could be inhibited by biphenylsulfonacetic acid derivatives, thereby raising the possibility that this class of inhibitors could be optimized as antiviral agents against multiple HPV types. These studies implicate Tyr-486 as a key residue for inhibitor binding and define an allosteric pocket on HPV E1 that can be exploited for future drug discovery efforts.
Asunto(s)
Acetatos/farmacología , Adenosina Trifosfato/metabolismo , Compuestos de Bifenilo/farmacología , Inhibidores Enzimáticos/farmacología , Proteínas Oncogénicas Virales/antagonistas & inhibidores , Sulfonas/farmacología , Tirosina/metabolismo , Regulación Alostérica , Compuestos de Bifenilo/química , Humanos , Hidrólisis , Proteínas Oncogénicas Virales/metabolismo , Papillomaviridae/enzimología , Relación Estructura-ActividadRESUMEN
The aminothiazolylphenyl-containing compounds BILS 179 BS and BILS 45 BS are novel inhibitors of the herpes simplex virus helicase-primase with antiviral activity in vitro and in animal models of HSV disease. To verify the mechanism of antiviral action, resistant viruses were selected by serial passage or by single-step plaque selection of HSV-1 KOS in the presence of inhibitors. Three resistant isolates K138r3, K22r5, and K22r1 were found to be 38-, 316-, and 2500-fold resistant to BILS 22 BS, a potent analog of BILS 45 BS. All three viruses had growth properties in vitro similar to wild-type HSV-1 KOS but they were sensitive to acyclovir. Cutaneous and intra-cerebral inoculation of mice with K22r1 or K22r5 resulted in pathogenicity equivalent to that of HSV-1 KOS. Both isolates were fully competent for reactivation from latency following corneal inoculation. Helicase-primase purified from cells infected with resistant viruses showed decreased inhibition in an in vitro DNA-dependent ATPase assay that correlated well with antiviral resistance. Marker transfer experiments and DNA sequence analysis identified single base pair mutations clustered in the N-terminus of the UL5 gene that resulted in single amino acid changes in the UL5 protein. Taken together, the results indicate that helicase-primase inhibitors prevent HSV growth by inhibiting HSV helicase-primase through specific interaction with the UL5 protein.
Asunto(s)
Antivirales/farmacología , ADN Helicasas/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Herpesvirus Humano 1/efectos de los fármacos , Herpesvirus Humano 1/aislamiento & purificación , Tiazoles/química , Animales , ADN Primasa , Modelos Animales de Enfermedad , Farmacorresistencia Viral , Herpesvirus Humano 1/enzimología , Herpesvirus Humano 1/genética , Ratones , Ratones Endogámicos BALB C , Mutagénesis , Tiazoles/farmacología , Proteínas ViralesRESUMEN
The virally encoded NS3 protease is essential to the life cycle of the hepatitis C virus (HCV), an important human pathogen causing chronic hepatitis, cirrhosis of the liver, and hepatocellular carcinoma. The design and synthesis of 15-membered ring beta-strand mimics which are capable of inhibiting the interactions between the HCV NS3 protease enzyme and its polyprotein substrate will be described. The binding interactions between a macrocyclic ligand and the enzyme were explored by NMR and molecular dynamics, and a model of the ligand/enzyme complex was developed.
Asunto(s)
Inhibidores de Proteasas/síntesis química , Inhibidores de Proteasas/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Sitios de Unión , Diseño de Fármacos , Espectroscopía de Resonancia Magnética , Espectrometría de Masas , Modelos Moleculares , Imitación Molecular , Inhibidores de Proteasas/químicaRESUMEN
The synthesis of BILN 2061, an NS3 protease inhibitor with proven antiviral effect in humans, was accomplished in a convergent manner from four building blocks. The procedure described here was suitable for the preparation of multigram quantities of BILN 2061 for preclinical pharmacological evaluation.
Asunto(s)
Antivirales/síntesis química , Antivirales/farmacología , Carbamatos/síntesis química , Carbamatos/farmacología , Compuestos Macrocíclicos/síntesis química , Compuestos Macrocíclicos/farmacología , Inhibidores de Proteasas/síntesis química , Inhibidores de Proteasas/farmacología , Quinolinas/síntesis química , Quinolinas/farmacología , Tiazoles/síntesis química , Tiazoles/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Antivirales/química , Carbamatos/química , Hepacivirus/efectos de los fármacos , Hepacivirus/enzimología , Humanos , Compuestos Macrocíclicos/química , Estructura Molecular , Inhibidores de Proteasas/química , Quinolinas/química , Tiazoles/química , Proteínas no Estructurales Virales/metabolismoRESUMEN
From the discovery of competitive hexapeptide inhibitors, potent and selective HCV NS3 protease macrocyclic inhibitors have been identified. Structure-activity relationship studies were performed focusing on optimizing the N-terminal carbamate and the aromatic substituent on the (4R)-hydroxyproline moiety. Inhibitors meeting the potency criteria in the cell-based assay and with improved oral bioavailability in rats were identified. BILN 2061 was selected as the best compound, the first NS3 protease inhibitor reported with antiviral activity in man.
Asunto(s)
Antivirales/síntesis química , Carbamatos/síntesis química , Hepacivirus/enzimología , Compuestos Heterocíclicos/síntesis química , Inhibidores de Proteasas/síntesis química , Proteínas no Estructurales Virales/antagonistas & inhibidores , Administración Oral , Animales , Antivirales/química , Antivirales/farmacología , Disponibilidad Biológica , Carbamatos/química , Carbamatos/farmacología , Compuestos Heterocíclicos/química , Compuestos Heterocíclicos/farmacología , Inyecciones Intravenosas , Prolina/química , Inhibidores de Proteasas/química , Inhibidores de Proteasas/farmacología , Ratas , Relación Estructura-ActividadRESUMEN
The Boehringer Ingelheim compound collection was screened for inhibitors of the ATPase activity of human papillomavirus E1 helicase to develop antiviral agents that inhibit human papillomavirus (HPV) DNA replication. This screen led to the discovery of (biphenyl-4-sulfonyl)acetic acid 1, which inhibits the ATPase activity of HPV type 6 E1 helicase with a low micromolar IC(50) value. A hit-to-lead exercise rapidly converted 1 into a low nanomolar lead series.
Asunto(s)
Acetatos/síntesis química , Adenosina Trifosfatasas/antagonistas & inhibidores , Compuestos de Bifenilo/síntesis química , Proteínas Oncogénicas Virales/antagonistas & inhibidores , Papillomaviridae/enzimología , Sulfonas/síntesis química , Acetatos/química , Adenosina Trifosfatasas/química , Antivirales/síntesis química , Antivirales/química , Compuestos de Bifenilo/química , Humanos , Proteínas Oncogénicas Virales/química , Relación Estructura-Actividad , Sulfonas/químicaRESUMEN
Hepatitis C virus (HCV) infection is a serious cause of chronic liver disease worldwide with more than 170 million infected individuals at risk of developing significant morbidity and mortality. Current interferon-based therapies are suboptimal especially in patients infected with HCV genotype 1, and they are poorly tolerated, highlighting the unmet medical need for new therapeutics. The HCV-encoded NS3 protease is essential for viral replication and has long been considered an attractive target for therapeutic intervention in HCV-infected patients. Here we identify a class of specific and potent NS3 protease inhibitors and report the evaluation of BILN 2061, a small molecule inhibitor biologically available through oral ingestion and the first of its class in human trials. Administration of BILN 2061 to patients infected with HCV genotype 1 for 2 days resulted in an impressive reduction of HCV RNA plasma levels, and established proof-of-concept in humans for an HCV NS3 protease inhibitor. Our results further illustrate the potential of the viral-enzyme-targeted drug discovery approach for the development of new HCV therapeutics.
Asunto(s)
Antivirales/uso terapéutico , Carbamatos/farmacología , Hepacivirus/efectos de los fármacos , Hepacivirus/fisiología , Hepatitis C/tratamiento farmacológico , Compuestos Macrocíclicos , Quinolinas , Inhibidores de Serina Proteinasa/uso terapéutico , Tiazoles/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Administración Oral , Antivirales/administración & dosificación , Antivirales/farmacocinética , Antivirales/farmacología , Carbamatos/administración & dosificación , Carbamatos/química , Carbamatos/farmacocinética , Método Doble Ciego , Hepacivirus/enzimología , Hepacivirus/genética , Hepatitis C/virología , Humanos , Masculino , Poliproteínas/metabolismo , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Inhibidores de Serina Proteinasa/administración & dosificación , Inhibidores de Serina Proteinasa/farmacocinética , Inhibidores de Serina Proteinasa/farmacología , Tiazoles/administración & dosificación , Tiazoles/química , Tiazoles/farmacocinética , Carga Viral , Proteínas no Estructurales Virales/metabolismo , Proteínas Virales/metabolismoRESUMEN
This study investigated the oral bioavailability and efficacy of BILS 45 BS, a selective herpes simplex virus (HSV) helicase-primase inhibitor, against acyclovir (ACV)-resistant (ACV(r)) infections mediated by the HSV type 1 (HSV-1) dlsptk and PAA(r)5 mutant strains. In vitro, the compound was more potent than ACV against wild-type clinical and laboratory HSV-1 strains and ACV(r) HSV isolates, as determined by a standard plaque reduction assay, with a mean 50% effective concentration of about 0.15 microM. The oral bioavailability of BILS 45 BS in hairless mice was 49%, with a peak concentration in plasma of 31.5 microM after administration of a single dose of 25 mg/kg. Following cutaneous infection of nude mice, both the HSV-1 dlsptk and PAA(r)5 mutant strains induced significant, reproducible, and persistent cutaneous lesions that lasted for more than 2 weeks. Oral treatment with ACV (100 or 125 mg/kg/day, three times a day by gavage) did not affect either mutant-induced infection. In contrast, BILS 45 BS at an oral dose of 100 mg/kg/day almost completely abolished cutaneous lesions mediated by both ACV(r) HSV-1 mutants. The 50% effective doses of BILS 45 BS were 56.7 and 61 mg/kg/day against dlsptk- and PAA(r)5-induced infections, respectively. Taken together, our results demonstrate very effective oral therapy of experimental ACV(r) HSV-1 infections in nude mice and support the potential use of HSV helicase-primase inhibitors for the treatment of nucleoside-resistant HSV disease in humans.