RESUMO
Allosteric integrase inhibitors (ALLINIs) of HIV-1 may hold promise as a novel mechanism for HIV therapeutics and cure. Scaffold modifications to the 4-(4,4-dimethylpiperidinyl) 2,6-dimethylpyridinyl class of ALLINIs provided a series of potent compounds with differentiated 5/6 fused ring systems. Notably, inhibitors containing the 1,2,4-triazolopyridine and imidazopyridine core exhibited single digit nM antiviral potency and low to moderate clearance after intravenous (IV) dosing in rat pharmacokinetic (PK) studies. The 1,2,4-triazolopyridines showed a higher oral exposure when compared to the imidazopyridines. Further modifications to the C5 substituent of the 1,2,4-triazolopyridines resulted in a new lead compound, which had improved rat IV/PO PK compared to the former lead compound GSK3739936, while maintaining antiviral potency. Structure-activity relationships (SAR) and rat pharmacokinetic profiles of this series are discussed.
Assuntos
Fármacos Anti-HIV , Inibidores de Integrase de HIV , Integrase de HIV , HIV-1 , Regulação Alostérica , Animais , Fármacos Anti-HIV/farmacologia , Integrase de HIV/metabolismo , Inibidores de Integrase de HIV/farmacologia , HIV-1/metabolismo , RatosRESUMO
It is estimated that more than 170 million people are infected with hepatitis C virus (HCV) worldwide. Clinical trials have demonstrated that, for the first time in human history, the potential exists to eradicate a chronic viral disease using combination therapies that contain only direct-acting antiviral agents. HCV non-structural protein 5A (NS5A) is a multifunctional protein required for several stages of the virus replication cycle. NS5A replication complex inhibitors, exemplified by daclatasvir (DCV; also known as BMS-790052 and Daklinza), belong to the most potent class of direct-acting anti-HCV agents described so far, with in vitro activity in the picomolar (pM) to low nanomolar (nM) range. The potency observed in vitro has translated into clinical efficacy, with HCV RNA declining by ~3-4 log10 in infected patients after administration of single oral doses of DCV. Understanding the exceptional potency of DCV was a key objective of this study. Here we show that although DCV and an NS5A inhibitor analogue (Syn-395) are inactive against certain NS5A resistance variants, combinations of the pair enhance DCV potency by >1,000-fold, restoring activity to the pM range. This synergistic effect was validated in vivo using an HCV-infected chimaeric mouse model. The cooperative interaction of a pair of compounds suggests that NS5A protein molecules communicate with each other: one inhibitor binds to resistant NS5A, causing a conformational change that is transmitted to adjacent NS5As, resensitizing resistant NS5A so that the second inhibitor can act to restore inhibition. This unprecedented synergistic anti-HCV activity also enhances the resistance barrier of DCV, providing additional options for HCV combination therapy and new insight into the role of NS5A in the HCV replication cycle.
Assuntos
Antivirais/farmacologia , Compostos de Bifenilo/farmacologia , Farmacorresistência Viral/efeitos dos fármacos , Hepacivirus/efeitos dos fármacos , Hepacivirus/genética , Imidazóis/farmacologia , Proteínas não Estruturais Virais/metabolismo , Regulação Alostérica/efeitos dos fármacos , Animais , Carbamatos , Linhagem Celular , Sinergismo Farmacológico , Quimioterapia Combinada , Hepacivirus/metabolismo , Hepatite C/virologia , Hepatócitos/transplante , Humanos , Camundongos , Modelos Moleculares , Conformação Proteica/efeitos dos fármacos , Multimerização Proteica/efeitos dos fármacos , Estrutura Quaternária de Proteína/efeitos dos fármacos , Pirrolidinas , Reprodutibilidade dos Testes , Valina/análogos & derivados , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/genética , Replicação Viral/efeitos dos fármacosRESUMO
An efficient large-scale synthesis of acid 1, a penultimate precursor to the HCV NS5A inhibitor BMS-986097, along with the final API step are described. Three routes were devised for the synthesis of 1 at the various stages of the program. The third generation route, the one that proved scalable and is the main subject of this paper, features a one-step Michael addition of t-butyl 2-((diphenylmethylene)amino)acetate (24) to (E)-benzyl 4-(1-hydroxycyclopropyl)but-2-enoate (28) followed by cyclization and chiral separation to form 27c, the core skeleton of cap piece 1. The epimerization and chiral resolution of 27c followed by further synthetic manipulations involving the carbamate formation, lactone reduction and cyclization, afforded cyclopropyl pyran 1. A detailed study of diphenylmethane deprotection via acid hydrolysis as well as a key lactone to tetrahydropyran conversion, in order to avoid a side reaction that afforded an alternative cyclization product, are discussed. This synthesis was applied to the preparation of more than 100 g of the final API BMS-986097 for toxicology studies.
Assuntos
Antivirais/síntese química , Glicina/análogos & derivados , Imidazóis/síntese química , Piranos/farmacologia , Pirrolidinas/síntese química , Compostos de Espiro/síntese química , Proteínas não Estruturais Virais/antagonistas & inibidores , Antivirais/química , Antivirais/farmacologia , Imidazóis/química , Imidazóis/farmacologia , Estrutura Molecular , Piranos/síntese química , Piranos/química , Pirrolidinas/química , Pirrolidinas/farmacologia , Compostos de Espiro/química , Compostos de Espiro/farmacologia , Proteínas não Estruturais Virais/metabolismoRESUMO
Daclatasvir is a first-in-class, potent, and selective inhibitor of the hepatitis C virus nonstructural protein 5A replication complex. In support of nonclinical studies during discovery and exploratory development, liquid chromatography-tandem mass spectrometry and nuclear magnetic resonance were used in connection with synthetic and radiosynthetic approaches to investigate the biotransformation of daclatasvir in vitro and in cynomolgus monkeys, dogs, mice, and rats. The results of these studies indicated that disposition of daclatasvir was accomplished mainly by the release of unchanged daclatasvir into bile and feces and, secondarily, by oxidative metabolism. Cytochrome P450s were the main enzymes involved in the metabolism of daclatasvir. Oxidative pathways included δ-oxidation of the pyrrolidine moiety, resulting in ring opening to an aminoaldehyde intermediate followed by an intramolecular reaction between the aldehyde and the proximal imidazole nitrogen atom. Despite robust formation of the resulting metabolite in multiple systems, rates of covalent binding to protein associated with metabolism of daclatasvir were modest (55.2-67.8 pmol/mg/h) in nicotinamide adenine dinucleotide phosphate (reduced form)-supplemented liver microsomes (human, monkey, rat), suggesting that intramolecular rearrangement was favored over intermolecular binding in the formation of this metabolite. This biotransformation profile supported the continued development of daclatasvir, which is now marketed for the treatment of chronic hepatitis C virus infection.
Assuntos
Biotransformação/fisiologia , Imidazóis/metabolismo , Pirrolidinas/metabolismo , Animais , Bile/metabolismo , Carbamatos , Cromatografia Líquida de Alta Pressão/métodos , Sistema Enzimático do Citocromo P-450/metabolismo , Cães , Haplorrinos , Hepatócitos/metabolismo , Humanos , Macaca fascicularis , Espectroscopia de Ressonância Magnética/métodos , Masculino , Espectrometria de Massas/métodos , Camundongos , Camundongos Endogâmicos BALB C , Microssomos Hepáticos/metabolismo , Oxirredução , Ratos , Ratos Sprague-Dawley , Valina/análogos & derivadosRESUMO
The worldwide prevalence of chronic hepatitis C virus (HCV) infection is estimated to be approaching 200 million people. Current therapy relies upon a combination of pegylated interferon-alpha and ribavirin, a poorly tolerated regimen typically associated with less than 50% sustained virological response rate in those infected with genotype 1 virus. The development of direct-acting antiviral agents to treat HCV has focused predominantly on inhibitors of the viral enzymes NS3 protease and the RNA-dependent RNA polymerase NS5B. Here we describe the profile of BMS-790052, a small molecule inhibitor of the HCV NS5A protein that exhibits picomolar half-maximum effective concentrations (EC(50)) towards replicons expressing a broad range of HCV genotypes and the JFH-1 genotype 2a infectious virus in cell culture. In a phase I clinical trial in patients chronically infected with HCV, administration of a single 100-mg dose of BMS-790052 was associated with a 3.3 log(10) reduction in mean viral load measured 24 h post-dose that was sustained for an additional 120 h in two patients infected with genotype 1b virus. Genotypic analysis of samples taken at baseline, 24 and 144 h post-dose revealed that the major HCV variants observed had substitutions at amino-acid positions identified using the in vitro replicon system. These results provide the first clinical validation of an inhibitor of HCV NS5A, a protein with no known enzymatic function, as an approach to the suppression of virus replication that offers potential as part of a therapeutic regimen based on combinations of HCV inhibitors.
Assuntos
Antivirais/farmacologia , Hepacivirus/efeitos dos fármacos , Imidazóis/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Adolescente , Adulto , Animais , Antivirais/sangue , Antivirais/química , Antivirais/uso terapêutico , Carbamatos , Linhagem Celular , Chlorocebus aethiops , Farmacorresistência Viral , Feminino , Genótipo , Células HeLa , Hepatite C/tratamento farmacológico , Hepatite C/virologia , Humanos , Imidazóis/sangue , Imidazóis/química , Concentração Inibidora 50 , Masculino , Pessoa de Meia-Idade , Pirrolidinas , Fatores de Tempo , Valina/análogos & derivados , Células Vero , Carga Viral/efeitos dos fármacos , Adulto JovemRESUMO
Daclatasvir (DCV) is a first-in-class hepatitis C virus (HCV) nonstructural 5A replication complex inhibitor (NS5A RCI) that is clinically effective in interferon-free combinations with direct-acting antivirals (DAAs) targeting alternate HCV proteins. Recently, we reported NS5A RCI combinations that enhance HCV inhibitory potential in vitro, defining a new class of HCV inhibitors termed NS5A synergists (J. Sun, D. R. O'Boyle II, R. A. Fridell, D. R. Langley, C. Wang, S. Roberts, P. Nower, B. M. Johnson F. Moulin, M. J. Nophsker, Y. Wang, M. Liu, K. Rigat, Y. Tu, P. Hewawasam, J. Kadow, N. A. Meanwell, M. Cockett, J. A. Lemm, M. Kramer, M. Belema, and M. Gao, Nature 527:245-248, 2015, doi:10.1038/nature15711). To extend the characterization of NS5A synergists, we tested new combinations of DCV and NS5A synergists against genotype (gt) 1 to 6 replicons and gt 1a, 2a, and 3a viruses. The kinetics of inhibition in HCV-infected cells treated with DCV, an NS5A synergist (NS5A-Syn), or a combination of DCV and NS5A-Syn were distinctive. Similar to activity observed clinically, DCV caused a multilog drop in HCV, followed by rebound due to the emergence of resistance. DCV-NS5A-Syn combinations were highly efficient at clearing cells of viruses, in line with the trend seen in replicon studies. The retreatment of resistant viruses that emerged using DCV monotherapy with DCV-NS5A-Syn resulted in a multilog drop and rebound in HCV similar to the initial decline and rebound observed with DCV alone on wild-type (WT) virus. A triple combination of DCV, NS5A-Syn, and a DAA targeting the NS3 or NS5B protein cleared the cells of viruses that are highly resistant to DCV. Our data support the observation that the cooperative interaction of DCV and NS5A-Syn potentiates both the genotype coverage and resistance barrier of DCV, offering an additional DAA option for combination therapy and tools for explorations of NS5A function.
Assuntos
Antivirais/farmacologia , Compostos de Bifenilo/farmacologia , Hepacivirus/efeitos dos fármacos , Imidazóis/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Carbamatos , Linhagem Celular , Sinergismo Farmacológico , Quimioterapia Combinada , Humanos , Simulação de Acoplamento Molecular , Pirrolidinas , Replicon/efeitos dos fármacos , Replicon/genética , Valina/análogos & derivadosRESUMO
The isoquinolinamide series of HCV NS5A inhibitors exemplified by compounds 2b and 2c provided the first dual genotype-1a/1b (GT-1a/1b) inhibitor class that demonstrated a significant improvement in potency toward GT-1a replicons compared to that of the initial program lead, stilbene 2a. Structure-activity relationship (SAR) studies that uncovered an alternate phenylglycine-based cap series that exhibit further improvements in virology profile, along with some insights into the pharmacophoric elements associated with the GT-1a potency, are described.
Assuntos
Antivirais/química , Glicina/análogos & derivados , Hepacivirus/enzimologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Animais , Antivirais/síntese química , Antivirais/farmacocinética , Cristalografia por Raios X , Avaliação Pré-Clínica de Medicamentos , Genótipo , Glicina/síntese química , Glicina/química , Glicina/farmacocinética , Meia-Vida , Hepacivirus/genética , Hepacivirus/fisiologia , Microssomos Hepáticos/metabolismo , Conformação Molecular , Ratos , Relação Estrutura-Atividade , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/efeitos dos fármacosRESUMO
In a recent disclosure, we described the discovery of dimeric, prolinamide-based NS5A replication complex inhibitors exhibiting excellent potency towards an HCV genotype 1b replicon. That disclosure dealt with the SAR exploration of the peripheral region of our lead chemotype, and herein is described the SAR uncovered from a complementary effort that focused on the central core region. From this effort, the contribution of the core region to the overall topology of the pharmacophore, primarily vector orientation and planarity, was determined, with a set of analogs exhibiting <10 nM EC(50) in a genotype 1b replicon assay.
Assuntos
Antivirais/química , Proteínas não Estruturais Virais/antagonistas & inibidores , Antivirais/farmacologia , Carbamatos , Hepacivirus/efeitos dos fármacos , Imidazóis/química , Imidazóis/farmacologia , Concentração Inibidora 50 , Estrutura Molecular , Prolina/análogos & derivados , Prolina/química , Prolina/farmacologia , Pirrolidinas , Relação Estrutura-Atividade , Valina/análogos & derivados , Proteínas não Estruturais Virais/química , Replicação Viral/efeitos dos fármacosRESUMO
Long-acting (LA) human immunodeficiency virus-1 (HIV-1) antiretroviral therapy characterized by a ≥1 month dosing interval offers significant advantages over daily oral therapy. However, the criteria for compounds that enter clinical development are high. Exceptional potency and low plasma clearance are required to meet dose size requirements; excellent chemical stability and/or crystalline form stability is required to meet formulation requirements, and new antivirals in HIV-1 therapy need to be largely free of side effects and drug-drug interactions. In view of these challenges, the discovery that capsid inhibitors comprising a quinazolinone core tolerate a wide range of structural modifications while maintaining picomolar potency against HIV-1 infection in vitro, are assembled efficiently in a multi-component reaction, and can be isolated in a stereochemically pure form is reported herein. The detailed characterization of a prototypical compound, GSK878, is presented, including an X-ray co-crystal structure and subcutaneous and intramuscular pharmacokinetic data in rats and dogs.
Assuntos
Fármacos Anti-HIV , Infecções por HIV , HIV-1 , Humanos , Ratos , Animais , Cães , Capsídeo , Proteínas do Capsídeo , Quinazolinonas/farmacologia , Quinazolinonas/uso terapêutico , Fármacos Anti-HIV/farmacocinética , Infecções por HIV/tratamento farmacológicoRESUMO
In a previous disclosure,(1) we reported the dimerization of an iminothiazolidinone to form 1, a contributor to the observed inhibition of HCV genotype 1b replicon activity. The dimer was isolated via bioassay-guided fractionation experiments and shown to be a potent inhibitor of genotype 1b HCV replication for which resistance mapped to the NS5A protein. The elements responsible for governing HCV inhibitory activity were successfully captured in the structurally simplified stilbene prolinamide 2. We describe herein the early SAR and profiling associated with stilbene prolinamides that culminated in the identification of analogs with PK properties sufficient to warrant continued commitment to this chemotype. These studies represent the key initial steps toward the discovery of daclatasvir (BMS-790052), a compound that has demonstrated clinical proof-of-concept for inhibiting the NS5A replication complex in the treatment of HCV infection.
Assuntos
Antivirais/farmacologia , Imidazóis/farmacologia , Prolina/análogos & derivados , Estilbenos/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Antivirais/síntese química , Antivirais/química , Carbamatos , Relação Dose-Resposta a Droga , Imidazóis/síntese química , Imidazóis/química , Estrutura Molecular , Prolina/síntese química , Prolina/química , Prolina/farmacologia , Pirrolidinas , Estilbenos/síntese química , Estilbenos/química , Relação Estrutura-Atividade , Valina/análogos & derivadosRESUMO
Allosteric HIV-1 integrase inhibitors (ALLINIs) have been of interest recently because of their novel mechanism of action. Strategic modifications to the C5 moiety of a class of 4-(4,4-dimethylpiperidinyl)-2,6-dimethylpyridinyl ALLINIs led to the identification of a tetrahydroisoquinoline heterocycle as a suitable spacer element to project the distal hydrophobic aryl ring. Subsequent optimization of the aryl substitutions identified 12 as an ALLINI with single-digit nanomolar inhibitory potency and low clearance across preclinical species. In preclinical toxicology studies with 12 in rats, lipid hepatocellular vacuolation was observed. Removal of the C6 methyl group resulted in GSK3839919 (22), which exhibited a reduced incidence and severity of lipid vacuolation in both in vitro assays and in vivo studies while maintaining the potency and pharmacokinetic (PK) properties of the prototype. The virology, PK, and toxicology profiles of 22 are discussed.
RESUMO
Hepatitis C virus (HCV) non-structural protein 5A (NS5A) is a multi-functional protein that is expressed in basally phosphorylated (p56) and in hyperphosphorylated (p58) forms. NS5A phosphorylation has been implicated in regulating multiple aspects of HCV replication. We recently reported the identification of a class of compounds that potently inhibit HCV RNA replication by targeting NS5A. Although the precise mechanism of inhibition of these compounds is not well understood, one activity that has been described is their ability to block expression of the hyperphosphorylated form of NS5A. Here, we report that an NS5A inhibitor impaired hyperphosphorylation without affecting basal phosphorylation at the C-terminal region of NS5A. This inhibitor activity did not require NS5A domains II and III and was distinct from that of a cellular kinase inhibitor that also blocked NS5A hyperphosphorylation, results that are consistent with an inhibitor-binding site within the N-terminal region of NS5A. In addition, we observed that an NS5A inhibitor promoted the accumulation of an HCV polyprotein intermediate, suggesting that inhibitor binding to NS5A may occur prior to the completion of polyprotein processing. Finally, we observed that NS5A p56 and p58 separated into different membrane fractions during discontinuous sucrose gradient centrifugation, consistent with these NS5A phosphoforms performing distinct replication functions. The p58 localization pattern was disrupted by an NS5A inhibitor. Collectively, our results suggest that NS5A inhibitors probably impact several aspects of HCV expression and regulation. These findings may help to explain the exceptional potency of this class of HCV replication complex inhibitors.
Assuntos
Antivirais/farmacologia , Inibidores Enzimáticos/farmacologia , Hepacivirus/efeitos dos fármacos , Hepacivirus/enzimologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Proteínas não Estruturais Virais/metabolismo , Fosforilação , RNA Polimerase Dependente de RNA/antagonistas & inibidores , RNA Polimerase Dependente de RNA/metabolismoRESUMO
This article summarizes key aspects of progress made during 2004 toward the design, discovery and development of antiviral agents for clinical use. Important developments in the identification, characterization and clinical utility of inhibitors of human immunodeficiency virus; the hepatitis viruses, hepatitis B, hepatitis C; the herpes family of viruses, herpes simplex viruses 1 and 2, varicella zoster virus, Epstein-Barr virus and human cytomegalovirus; the respiratory viruses, influenza, respiratory syncytial virus, human metapneumovirus, picornaviruses, measles and the severe acute respiratory syndrome coronavirus; human papilloma virus; rotavirus; Ebola virus and West Nile virus, are reviewed.
Assuntos
Antivirais/química , Antivirais/farmacologia , Desenho de Fármacos , HIV-1/efeitos dos fármacos , HIV-2/efeitos dos fármacos , Vírus de Hepatite/efeitos dos fármacos , Herpesviridae/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Fármacos Anti-HIV/química , Fármacos Anti-HIV/farmacologia , Ensaios Clínicos como Assunto , Avaliação Pré-Clínica de Medicamentos , Inibidores de Integrase de HIV/química , Inibidores de Integrase de HIV/farmacologia , Humanos , Dados de Sequência Molecular , Estrutura Molecular , Inibidores da Síntese de Ácido Nucleico , Nucleosídeos/química , Nucleosídeos/farmacologia , Orthomyxoviridae/efeitos dos fármacos , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , Inibidores da Transcriptase Reversa/química , Inibidores da Transcriptase Reversa/farmacologia , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/efeitos dos fármacosRESUMO
The discovery and development of the first-in-class hepatitis C virus (HCV) NS5A replication complex inhibitor daclatasvir (6) provides a compelling example of the power of phenotypic screening to identify leads engaging novel targets in mechanistically unique ways. HCV NS5A replication complex inhibitors are pan-genotypic in spectrum, and this mechanistic class provides the most potent HCV inhibitors in vitro that have been described to date. Clinical trials with 6 demonstrated a potent effect on reducing plasma viral load and, in combination with mechanistically orthogonal HCV inhibitors, established the ability to cure even the most difficult infections without the need for immune stimulation. In this Drug Annotation, we describe the discovery of the original high-throughput screening lead 7 and the chemical conundrum and challenges resolved in optimizing to 6 as a clinical candidate and finally we summarize the results of select clinical studies.
Assuntos
Antivirais/química , Hepacivirus/efeitos dos fármacos , Imidazóis/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Carbamatos , Ensaios Clínicos como Assunto , Farmacorresistência Viral , Genótipo , Hepacivirus/genética , Humanos , Imidazóis/química , Imidazóis/uso terapêutico , Pirrolidinas , Relação Estrutura-Atividade , Valina/análogos & derivadosRESUMO
Lead inhibitors that target the function of the hepatitis C virus (HCV) nonstructural 5A (NS5A) protein have been identified by phenotypic screening campaigns using HCV subgenomic replicons. The demonstration of antiviral activity in HCV-infected subjects by the HCV NS5A replication complex inhibitor (RCI) daclatasvir (1) spawned considerable interest in this mechanistic approach. In this Perspective, we summarize the medicinal chemistry studies that led to the discovery of 1 and other chemotypes for which resistance maps to the NS5A protein and provide synopses of the profiles of many of the compounds currently in clinical trials. We also summarize what is currently known about the NS5A protein and the studies using NS5A RCIs and labeled analogues that are helping to illuminate aspects of both protein function and inhibitor interaction. We conclude with a synopsis of the results of notable clinical trials with HCV NS5A RCIs.
Assuntos
Descoberta de Drogas , Hepacivirus/metabolismo , Proteínas não Estruturais Virais/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Hepacivirus/fisiologia , Proteínas não Estruturais Virais/metabolismoRESUMO
A series of symmetrical E-stilbene prolinamides that originated from the library-synthesized lead 3 was studied with respect to HCV genotype 1a (G-1a) and genotype 1b (G-1b) replicon inhibition and selectivity against BVDV and cytotoxicity. SAR emerging from an examination of the prolinamide cap region revealed 11 to be a selective HCV NS5A inhibitor exhibiting submicromolar potency against both G-1a and G-1b replicons. Additional structural refinements resulted in the identification of 30 as a potent, dual G-1a/1b HCV NS5A inhibitor.
Assuntos
Antivirais/farmacologia , Genótipo , Hepacivirus/efeitos dos fármacos , Inibidores de Proteases/farmacologia , RNA Polimerase Dependente de RNA/antagonistas & inibidores , Replicon/efeitos dos fármacos , Proteínas não Estruturais Virais/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Antivirais/química , Hepacivirus/genética , Hepacivirus/fisiologia , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Inibidores de Proteases/química , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
A medicinal chemistry campaign that was conducted to address a potential genotoxic liability associated with an aniline-derived scaffold in a series of HCV NS5A inhibitors with dual GT-1a/-1b inhibitory activity is described. Anilides 3b and 3c were used as vehicles to explore structural modifications that retained antiviral potency while removing the potential for metabolism-based unmasking of the embedded aniline. This effort resulted in the discovery of a highly potent biarylimidazole chemotype that established a potency benchmark in replicon assays, particularly toward HCV GT-1a, a strain with significant clinical importance. Securing potent GT-1a activity in a chemotype class lacking overt structural liabilities was a critical milestone in the effort to realize the full clinical potential of targeting the HCV NS5A protein.
Assuntos
Antivirais/farmacologia , Genótipo , Hepacivirus/efeitos dos fármacos , Imidazóis/farmacologia , RNA Polimerase Dependente de RNA/antagonistas & inibidores , Replicon/efeitos dos fármacos , Proteínas não Estruturais Virais/antagonistas & inibidores , Antivirais/química , Antivirais/farmacocinética , Cristalografia por Raios X , Descoberta de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Inibidores Enzimáticos/farmacologia , Hepacivirus/enzimologia , Hepacivirus/genética , Hepacivirus/fisiologia , Imidazóis/química , Imidazóis/farmacocinética , Espectroscopia de Ressonância Magnética , Espectrometria de Massas por Ionização por Electrospray , Relação Estrutura-AtividadeRESUMO
The biphenyl derivatives 2 and 3 are prototypes of a novel class of NS5A replication complex inhibitors that demonstrate high inhibitory potency toward a panel of clinically relevant HCV strains encompassing genotypes 1-6. However, these compounds exhibit poor systemic exposure in rat pharmacokinetic studies after oral dosing. The structure-activity relationship investigations that improved the exposure properties of the parent bis-phenylimidazole chemotype, culminating in the identification of the highly potent NS5A replication complex inhibitor daclatasvir (33) are described. An element critical to success was the realization that the arylglycine cap of 2 could be replaced with an alkylglycine derivative and still maintain the high inhibitory potency of the series if accompanied with a stereoinversion, a finding that enabled a rapid optimization of exposure properties. Compound 33 had EC50 values of 50 and 9 pM toward genotype-1a and -1b replicons, respectively, and oral bioavailabilities of 38-108% in preclinical species. Compound 33 provided clinical proof-of-concept for the NS5A replication complex inhibitor class, and regulatory approval to market it with the NS3/4A protease inhibitor asunaprevir for the treatment of HCV genotype-1b infection has recently been sought in Japan.
Assuntos
Antivirais/farmacologia , Inibidores Enzimáticos/farmacologia , Hepacivirus/efeitos dos fármacos , Imidazóis/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Animais , Antivirais/química , Antivirais/farmacocinética , Área Sob a Curva , Carbamatos , Cães , Descoberta de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Hepacivirus/enzimologia , Hepacivirus/fisiologia , Imidazóis/química , Imidazóis/farmacocinética , Espectroscopia de Ressonância Magnética , Pirrolidinas , Ratos , Espectrometria de Massas por Ionização por Electrospray , Relação Estrutura-Atividade , Valina/análogos & derivadosRESUMO
The hepatitis C virus NS5A protein is an established and clinically validated target for antiviral intervention by small molecules. Characterizations are presented of compounds identified as potent inhibitors of HCV replication to provide insight into structural elements that interact with the NS5A protein. UV-activated cross linking and affinity isolation was performed with one series to probe the physical interaction between the inhibitors and the NS5A protein expressed in HCV replicon cells. Resistance mapping with the second series was used to determine the functional impact of specific inhibitor subdomains on the interaction with NS5A. The data provide evidence for a direct high-affinity interaction between these inhibitors and the NS5A protein, with the interaction dependent on inhibitor stereochemistry. The functional data supports a model of inhibition that implicates inhibitor binding by covalently combining distinct pharmacophores across an NS5A dimer interface to achieve maximal inhibition of HCV replication.
Assuntos
Antivirais/farmacologia , Inibidores Enzimáticos/farmacologia , Hepacivirus/efeitos dos fármacos , Hepacivirus/enzimologia , RNA Polimerase Dependente de RNA/antagonistas & inibidores , Proteínas não Estruturais Virais/antagonistas & inibidores , Farmacorresistência Viral , Humanos , Ligação ProteicaRESUMO
We have recently identified BMS-345541 (1) as a highly selective and potent inhibitor of IKK-2 (IC50 = 0.30 microM), which however was considerably less potent against IKK-1 (IC50 = 4.0 microM). In order to further explore the SAR around the imidazoquinoxaline tricyclic structure of 1, we prepared a series of tetracyclic analogues (7, 13, and 18). The synthesis and biological activities of these potent IKK inhibitors are described.