RESUMO
Several novel chemical series were identified that modulate glucocerebrosidase (GCase). Compounds from these series are active on glucosylceramide, unlike other known GCase modulators. We obtained GCase crystal structures with two compounds that have distinct chemotypes. Positive allosteric modulators bind to a site on GCase and induce conformational changes, but also induce an equilibrium state between monomer and dimer.
Assuntos
Doença de Gaucher , Glucosilceramidase , Humanos , Glucosilceramidase/química , Glucosilceramidase/metabolismo , Glucosilceramidas , Hidrólise , Doença de Gaucher/tratamento farmacológicoRESUMO
AIMS: Kv1.1 (KCNA1) channels contribute to the control of neuronal excitability and have been associated with epilepsy. Kv1.1 channels can associate with the cytoplasmic Kvß1 subunit resulting in rapid inactivating A-type currents. We hypothesized that removal of channel inactivation, by modulating Kv1.1/Kvß1 interaction with a small molecule, would lead to decreased neuronal excitability and anticonvulsant activity. METHODS: We applied high-throughput screening to identify ligands able to modulate the Kv1.1-T1 domain/Kvß1 protein complex. We then selected a compound that was characterized on recombinant Kv1.1/Kvß1 channels by electrophysiology and further evaluated on sustained neuronal firing and on in vitro epileptiform activity using a high K+ -low Ca2+ model in hippocampal slices. RESULTS: We identified a novel compound able to modulate the interaction of the Kv1.1/Kvß1 complex and that produced a functional inhibition of Kv1.1/Kvß1 channel inactivation. We demonstrated that this compound reduced the sustained repetitive firing in hippocampal neurons and was able to abolish the development of in vitro epileptiform activity. CONCLUSIONS: This study describes a rational drug discovery approach for the identification of novel ligands that inhibit Kv1.1 channel inactivation and provides pharmacological evidence that such a mechanism translates into physiological effects by reducing in vitro epileptiform activity.
Assuntos
Potenciais de Ação/fisiologia , Descoberta de Drogas/métodos , Hipocampo/fisiologia , Canal de Potássio Kv1.1/fisiologia , Neurônios/fisiologia , Potenciais de Ação/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Feminino , Células HEK293 , Ensaios de Triagem em Larga Escala/métodos , Hipocampo/efeitos dos fármacos , Humanos , Canal de Potássio Kv1.1/agonistas , Canal de Potássio Kv1.1/antagonistas & inibidores , Neurônios/efeitos dos fármacos , Técnicas de Cultura de Órgãos , Bloqueadores dos Canais de Potássio/farmacologia , Estrutura Secundária de Proteína , Ratos , Xenopus laevisRESUMO
The management of back-up strategies in drug discovery and development is usually done on an ad hoc basis depending upon a series of external factors including overall portfolio status and resource and/or budget availability. These are however an essential component of risk management and merit a more structured and systematic conduct throughout the lifetime of a project. An approach based upon a thorough alignment of decision points and data availability as well as a tailor-made progression of various types of back-up program as a function of project categorization is suggested.
Assuntos
Descoberta de Drogas , Animais , Tomada de Decisões , HumanosRESUMO
A selective α2C -adrenoceptor (AR) agonist was developed for the treatment of neuropathic pain. The objective was to dissociate analgesic activity from cardiovascular and sedative side effects commonly observed with nonselective agents. A 2-amino-oxazoline derivative (compound A), identified as a dual α2C -AR agonist/α2A -AR antagonist in in vitro-binding assays, exhibited in vivo efficacy in rodent pain models. Its safety profile was compared with that of clonidine in six different in vivo models. Contrary to clonidine, compound A did not induce hypotension in pentobarbital-anesthetized rats, in conscious spontaneous hypertensive rats, or in telemetered dogs. Both agents induced similar dose-dependent decreases in heart rate in dogs and rats. In anesthetized pithed rats, clonidine showed dose-dependent hypertension and inhibited electrical nerve stimulation-induced tachycardia at doses close to its efficacious doses in the mouse formalin test, while compound A had much weaker vasoconstrictive and antichronotropic effects. Finally, in a mouse Irwin test, no sedation was observed with compound A at 30-fold its ED50 in the mouse formalin test, while sedative effects of clonidine started from three-fold its ED50 . These data confirm the advantageous safety profile of the new dual α2C -AR agonist/α2A -AR antagonist agent vs. the nonselective agonist clonidine.
Assuntos
Agonistas de Receptores Adrenérgicos alfa 2/farmacologia , Antagonistas de Receptores Adrenérgicos alfa 2/farmacologia , Analgésicos/farmacologia , Oxazóis/farmacologia , Agonistas de Receptores Adrenérgicos alfa 2/administração & dosagem , Agonistas de Receptores Adrenérgicos alfa 2/toxicidade , Antagonistas de Receptores Adrenérgicos alfa 2/administração & dosagem , Antagonistas de Receptores Adrenérgicos alfa 2/toxicidade , Analgésicos/administração & dosagem , Analgésicos/toxicidade , Animais , Clonidina/administração & dosagem , Clonidina/farmacologia , Clonidina/toxicidade , Modelos Animais de Doenças , Cães , Relação Dose-Resposta a Droga , Masculino , Camundongos , Oxazóis/administração & dosagem , Oxazóis/toxicidade , Dor/tratamento farmacológico , Ratos , Ratos Endogâmicos SHR , Ratos Sprague-DawleyRESUMO
The role of the synaptic vesicle protein 2A (SV2A) protein, target of the antiepileptic drug levetiracetam, is still mostly unknown. Considering its potential to provide in vivo functional insights into the role of SV2A in epileptic patients, the development of an SV2A positron emission tomography (PET) tracer has been undertaken. Using a 3D pharmacophore model based on close analogues of levetiracetam, we report the rationale design of three heterocyclic non-acetamide lead compounds, UCB-A, UCB-H and UCB-J, the first single-digit nanomolar SV2A ligands with suitable properties for development as PET tracers.
Assuntos
Acetamidas , Descoberta de Drogas , Compostos Heterocíclicos , Glicoproteínas de Membrana/análise , Proteínas do Tecido Nervoso/análise , Tomografia por Emissão de Pósitrons , Acetamidas/síntese química , Acetamidas/química , Animais , Compostos Heterocíclicos/síntese química , Compostos Heterocíclicos/química , Humanos , Ligantes , Masculino , Glicoproteínas de Membrana/metabolismo , Modelos Moleculares , Estrutura Molecular , Proteínas do Tecido Nervoso/metabolismo , Traçadores Radioativos , Ratos , Ratos WistarRESUMO
An X-ray crystal structure of Kelch-like ECH-associated protein (Keap1) co-crystallised with (1S,2R)-2-[(1S)-1-[(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)methyl]-1,2,3,4-tetrahydroisoquinolin-2-carbonyl]cyclohexane-1-carboxylic acid (compound (S,R,S)-1 a) was obtained. This X-ray crystal structure provides breakthrough experimental evidence for the true binding mode of the hit compound (S,R,S)-1 a, as the ligand orientation was found to differ from that of the initial docking model, which was available at the start of the project. Crystallographic elucidation of this binding mode helped to focus and drive the drug design process more effectively and efficiently.