RESUMO
ER aminopeptidase 1 (ERAP1) is an intracellular enzyme that generates antigenic peptides and is an emerging target for cancer immunotherapy and the control of autoimmunity. ERAP1 inhibitors described previously target the active site and are limited in selectivity, minimizing their clinical potential. To address this, we targeted the regulatory site of ERAP1 using a high-throughput screen and discovered a small molecule hit that is highly selective for ERAP1. (4aR,5S,6R,8S,8aR)-5-(2-(Furan-3-yl)ethyl)-8-hydroxy-5,6,8a-trimethyl-3,4,4a,5,6,7,8,8a-octahydronaphthalene-1-carboxylic acid is a natural product found in Dodonaea viscosa that constitutes a submicromolar, highly selective, and cell-active modulator of ERAP1. Although the compound activates hydrolysis of small model substrates, it is a competitive inhibitor for physiologically relevant longer peptides. Crystallographic analysis confirmed that the compound targets the regulatory site of the enzyme that normally binds the C-terminus of the peptide substrate. Our findings constitute a novel starting point for the development of selective ERAP1 modulators that have potential for further clinical development.
Assuntos
Aminopeptidases/antagonistas & inibidores , Apresentação de Antígeno/efeitos dos fármacos , Diterpenos Clerodânicos/farmacologia , Epitopos/metabolismo , Peptídeos/metabolismo , Inibidores de Proteases/farmacologia , Sítio Alostérico , Aminopeptidases/química , Aminopeptidases/metabolismo , Animais , Domínio Catalítico , Cristalografia por Raios X , Diterpenos Clerodânicos/química , Diterpenos Clerodânicos/metabolismo , Ativadores de Enzimas/química , Ativadores de Enzimas/metabolismo , Ativadores de Enzimas/farmacologia , Epitopos/química , Células HeLa , Humanos , Camundongos , Antígenos de Histocompatibilidade Menor/química , Antígenos de Histocompatibilidade Menor/metabolismo , Peptídeos/química , Inibidores de Proteases/química , Inibidores de Proteases/metabolismo , Ligação Proteica , Proteólise/efeitos dos fármacosRESUMO
A deconstruction of previously reported phosphoinositide 3-kinase δ (PI3Kδ) inhibitors and subsequent regrowth led to the identification of a privileged fragment for PI3Kδ, which was exploited to deliver a potent, efficient, and selective lead series with a novel binding mode observed in the PI3Kδ crystal structure.
Assuntos
Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Relação Estrutura-Atividade , Administração por Inalação , Animais , Classe Ia de Fosfatidilinositol 3-Quinase/química , Cristalografia por Raios X , Cães , Avaliação Pré-Clínica de Medicamentos , Canal de Potássio ERG1/metabolismo , Inibidores Enzimáticos/administração & dosagem , Ligação de Hidrogênio , Isoquinolinas/química , Células Madin Darby de Rim Canino , RatosRESUMO
A four-step process of high-quality modeling of existing data, deconstruction, identification of replacement cores, and an innovative synthetic regrowth strategy led to the rapid discovery of a novel oral series of PI3Kδ inhibitors with promising selectivity and excellent in vivo characteristics.
Assuntos
Inibidores de Fosfoinositídeo-3 Quinase , Inibidores de Proteínas Quinases/farmacologia , Doenças Respiratórias/tratamento farmacológico , Administração por Inalação , Animais , Disponibilidade Biológica , Classe Ia de Fosfatidilinositol 3-Quinase/metabolismo , Relação Dose-Resposta a Droga , Masculino , Modelos Moleculares , Estrutura Molecular , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/química , Ratos , Ratos Sprague-Dawley , Doenças Respiratórias/metabolismo , Relação Estrutura-AtividadeRESUMO
ASK1, a member of the MAPK Kinase Kinase family of proteins has been shown to play a key role in cancer, neurodegeneration and cardiovascular diseases and is emerging as a possible drug target. Here we describe a 'replacement-soaking' method that has enabled the high-throughput X-ray structure determination of ASK1/ligand complexes. Comparison of the X-ray structures of five ASK1/ligand complexes from 3 different chemotypes illustrates that the ASK1 ATP binding site is able to accommodate a range of chemical diversity and different binding modes. The replacement-soaking system is also able to tolerate some protein flexibility. This crystal system provides a robust platform for ASK1/ligand structure determination and future structure based drug design.
Assuntos
MAP Quinase Quinase Quinase 5/antagonistas & inibidores , MAP Quinase Quinase Quinase 5/química , Estaurosporina/química , Sítios de Ligação , Doenças Cardiovasculares/tratamento farmacológico , Cristalografia por Raios X , Desenho de Fármacos , Humanos , Ligantes , MAP Quinase Quinase Quinase 5/genética , MAP Quinase Quinase Quinase 5/metabolismo , Neoplasias/tratamento farmacológico , Doenças Neurodegenerativas/tratamento farmacológico , Células Sf9 , Transdução de SinaisRESUMO
The jumonji (JMJ) family of histone demethylases are Fe2+- and α-ketoglutarate-dependent oxygenases that are essential components of regulatory transcriptional chromatin complexes. These enzymes demethylate lysine residues in histones in a methylation-state and sequence-specific context. Considerable effort has been devoted to gaining a mechanistic understanding of the roles of histone lysine demethylases in eukaryotic transcription, genome integrity and epigenetic inheritance, as well as in development, physiology and disease. However, because of the absence of any selective inhibitors, the relevance of the demethylase activity of JMJ enzymes in regulating cellular responses remains poorly understood. Here we present a structure-guided small-molecule and chemoproteomics approach to elucidating the functional role of the H3K27me3-specific demethylase subfamily (KDM6 subfamily members JMJD3 and UTX). The liganded structures of human and mouse JMJD3 provide novel insight into the specificity determinants for cofactor, substrate and inhibitor recognition by the KDM6 subfamily of demethylases. We exploited these structural features to generate the first small-molecule catalytic site inhibitor that is selective for the H3K27me3-specific JMJ subfamily. We demonstrate that this inhibitor binds in a novel manner and reduces lipopolysaccharide-induced proinflammatory cytokine production by human primary macrophages, a process that depends on both JMJD3 and UTX. Our results resolve the ambiguity associated with the catalytic function of H3K27-specific JMJs in regulating disease-relevant inflammatory responses and provide encouragement for designing small-molecule inhibitors to allow selective pharmacological intervention across the JMJ family.