RESUMEN
TNF receptor associated periodic syndrome (TRAPS) is an autoinflammatory disease caused by mutations in TNF Receptor 1 (TNFR1). Current therapies for TRAPS are limited and do not target the pro-inflammatory signalling pathways that are central to the disease mechanism. Our aim was to identify drugs for repurposing as anti-inflammatories based on their ability to down-regulate molecules associated with inflammatory signalling pathways that are activated in TRAPS. This was achieved using rigorously optimized, high through-put cell culture and reverse phase protein microarray systems to screen compounds for their effects on the TRAPS-associated inflammatory signalome. 1360 approved, publically available, pharmacologically active substances were investigated for their effects on 40 signalling molecules associated with pro-inflammatory signalling pathways that are constitutively upregulated in TRAPS. The drugs were screened at four 10-fold concentrations on cell lines expressing both wild-type (WT) TNFR1 and TRAPS-associated C33Y mutant TNFR1, or WT TNFR1 alone; signalling molecule levels were then determined in cell lysates by the reverse-phase protein microarray. A novel mathematical methodology was developed to rank the compounds for their ability to reduce the expression of signalling molecules in the C33Y-TNFR1 transfectants towards the level seen in the WT-TNFR1 transfectants. Seven high-ranking drugs were selected and tested by RPPA for effects on the same 40 signalling molecules in lysates of peripheral blood mononuclear cells (PBMCs) from C33Y-TRAPS patients compared to PBMCs from normal controls. The fluoroquinolone antibiotic lomefloxacin, as well as others from this class of compounds, showed the most significant effects on multiple pro-inflammatory signalling pathways that are constitutively activated in TRAPS; lomefloxacin dose-dependently significantly reduced expression of 7/40 signalling molecules across the Jak/Stat, MAPK, NF-κB and PI3K/AKT pathways. This study demonstrates the power of signalome screening for identifying candidates for drug repurposing.
Asunto(s)
Antiinflamatorios/farmacología , Fiebre/inmunología , Fluoroquinolonas/farmacología , Enfermedades Autoinflamatorias Hereditarias/inmunología , Transducción de Señal/efectos de los fármacos , Adulto , Línea Celular Tumoral , Reposicionamiento de Medicamentos , Femenino , Ensayos Analíticos de Alto Rendimiento , Humanos , Masculino , Persona de Mediana Edad , Mutación , Receptores Tipo I de Factores de Necrosis Tumoral/genéticaRESUMEN
There are a number of small-molecule inhibitors targeting the RAS/RAF/MEK/ERK signaling pathway that have either been approved or are in clinical development for oncology across a range of disease indications. The inhibition of ERK1/2 is of significant current interest, as cell lines with acquired resistance to BRAF and MEK inhibitors have been shown to maintain sensitivity to ERK1/2 inhibition in preclinical models. This article reports on our recent work to identify novel, potent, and selective reversible ERK1/2 inhibitors from a low-molecular-weight, modestly active, and highly promiscuous chemical start point, compound 4. To guide and inform the evolution of this series, inhibitor binding mode information from X-ray crystal structures was critical in the rapid exploration of this template to compound 35, which was active when tested in in vivo antitumor efficacy experiments.
Asunto(s)
Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Animales , Disponibilidad Biológica , Línea Celular Tumoral , Perros , Descubrimiento de Drogas , Humanos , Metilación , Inhibidores de Proteínas Quinasas/farmacocinéticaRESUMEN
Computer-aided drug design encompasses a wide variety of tools and techniques, and can be implemented with a range of organisational structures and focus in different organisations. Here we outline the computational chemistry skills within Sygnature Discovery, along with the software and hardware at our disposal, and briefly discuss the methods that are not employed and why. The goal of the group is to provide support for design and analysis in order to improve the quality of compounds synthesised and reduce the timelines of drug discovery projects, and we reveal how this is achieved at Sygnature. Impact on medicinal chemistry is vital to demonstrating the value of computational chemistry, and we discuss the approaches taken to influence the list of compounds for synthesis, and how we recognise success. Finally we touch on some of the areas being developed within the team in order to provide further value to the projects and clients.
Asunto(s)
Diseño Asistido por Computadora , Descubrimiento de Drogas/métodos , Industria Farmacéutica/métodos , Biología Computacional , Diseño de Fármacos , Modelos Moleculares , Estructura Molecular , Programas Informáticos , Relación Estructura-ActividadRESUMEN
A weak screening hit with suboptimal physicochemical properties was optimized against PFKFB3 kinase using critical structure-guided insights. The resulting compounds demonstrated high selectivity over related PFKFB isoforms and modulation of the target in a cellular context. A selected example demonstrated exposure in animals following oral dosing. Examples from this series may serve as useful probes to understand the emerging biology of this metabolic target.
Asunto(s)
Diseño de Fármacos , Fosfofructoquinasa-2/antagonistas & inhibidores , Fosfofructoquinasa-2/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Administración Oral , Animales , Línea Celular , Humanos , Masculino , Ratones , Modelos Moleculares , Fosfofructoquinasa-2/química , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/farmacocinética , Ratas Wistar , Relación Estructura-ActividadRESUMEN
By careful analysis of experimental X-ray ligand crystallographic protein data across several inhibitor series we have discovered a novel, potent and selective series of iNOS inhibitors exemplified by compound 8.
Asunto(s)
Inhibidores Enzimáticos/química , Isoxazoles/química , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Piridinas/química , Animales , Sitios de Unión , Simulación por Computador , Cristalografía por Rayos X , Evaluación Preclínica de Medicamentos , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Humanos , Isoxazoles/síntesis química , Isoxazoles/farmacología , Ratones , Microsomas Hepáticos/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Estructura Terciaria de Proteína , Piridinas/farmacología , RatasRESUMEN
The design and synthesis of a new series of c-Jun N-terminal kinase inhibitors are reported. The novel series of substituted amino indazoles were designed based on a combination of hits from high-throughput screening and X-ray crystal structure information of the compounds crystallised into the JNK-1 ATP binding site.
Asunto(s)
Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Indazoles/síntesis química , Indazoles/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos/antagonistas & inhibidores , Adenosina Trifosfato/química , Adenosina Trifosfato/metabolismo , Sitios de Unión , Cristalografía por Rayos X , Diseño de Fármacos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Proteína Quinasa 8 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 8 Activada por Mitógenos/metabolismo , Modelos Moleculares , Estructura Molecular , Relación Estructura-ActividadRESUMEN
A hit-to-lead optimisation programme was carried out on the thiophenecarboxamide high throughput screening hits 1 and 2 resulting in the discovery of the potent and orally bioavailable IKK-2 inhibitor 22.