Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 15 de 15
Filtrar
1.
Bioorg Med Chem Lett ; 28(21): 3458-3462, 2018 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-30249354

RESUMO

The discovery and lead optimisation of a novel series of SYK inhibitors is described. These were optimised for SYK potency and selectivity against Aurora B. Compounds were profiled in a human skin penetration study to identify a suitable candidate molecule for pre-clinical development. Compound 44 (GSK2646264) was selected for progression and is currently in Phase I clinical trials.


Assuntos
Anti-Inflamatórios/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Pele/efeitos dos fármacos , Quinase Syk/antagonistas & inibidores , Administração Tópica , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/síntese química , Anti-Inflamatórios/química , Domínio Catalítico , Linhagem Celular Tumoral , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Piridinas/administração & dosagem , Piridinas/síntese química , Piridinas/química , Relação Estrutura-Atividade , Quinase Syk/química
2.
Nature ; 488(7411): 404-8, 2012 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-22842901

RESUMO

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.


Assuntos
Inibidores Enzimáticos/farmacologia , Histona Desmetilases com o Domínio Jumonji/antagonistas & inibidores , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Sequência de Aminoácidos , Animais , Biocatálise/efeitos dos fármacos , Domínio Catalítico , Células Cultivadas , Inibidores Enzimáticos/metabolismo , Evolução Molecular , Histonas/química , Histonas/metabolismo , Humanos , Concentração Inibidora 50 , Histona Desmetilases com o Domínio Jumonji/química , Histona Desmetilases com o Domínio Jumonji/classificação , Histona Desmetilases com o Domínio Jumonji/metabolismo , Lisina/metabolismo , Macrófagos/enzimologia , Macrófagos/metabolismo , Metilação/efeitos dos fármacos , Camundongos , Modelos Moleculares , Especificidade por Substrato , Fator de Necrose Tumoral alfa/biossíntese
3.
Bioorg Med Chem Lett ; 26(19): 4606-4612, 2016 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-27578246

RESUMO

The optimisation of the azanaphthyridine series of Spleen Tyrosine Kinase inhibitors is described. The medicinal chemistry strategy was focused on optimising the human whole blood activity whilst achieving a sufficient margin over hERG activity. A good pharmacokinetic profile was achieved by modification of the pKa. Morpholine compound 32 is a potent SYK inhibitor showing moderate selectivity, good oral bioavailability and good efficacy in the rat Arthus model but demonstrated a genotoxic potential in the Ames assay.


Assuntos
Naftiridinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Administração Oral , Animais , Disponibilidade Biológica , Cristalografia por Raios X , Humanos , Testes de Mutagenicidade , Naftiridinas/administração & dosagem , Naftiridinas/farmacocinética , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacocinética , Ratos , Relação Estrutura-Atividade
5.
Gut ; 62(11): 1581-90, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23242119

RESUMO

OBJECTIVE: Intestinal inflammation resulting from manipulation-induced mast cell activation is a crucial mechanism in the pathophysiology of postoperative ileus (POI). Recently it has been shown that spleen tyrosine kinase (Syk) is involved in mast cell degranulation. Therefore, we have evaluated the effect of the Syk-inhibitor GSK compound 143 (GSK143) as potential treatment to shorten POI. DESIGN: In vivo: in a mouse model of POI, the effect of the Syk inhibitor (GSK143) was evaluated on gastrointestinal transit, muscular inflammation and cytokine production. In vitro: the effect of GSK143 and doxantrazole were evaluated on cultured peritoneal mast cells (PMCs) and bone marrow derived macrophages. RESULTS: In vivo: intestinal manipulation resulted in a delay in gastrointestinal transit at t=24 h (Geometric Center (GC): 4.4 ± 0.3). Doxantrazole and GSK143 significantly increased gastrointestinal transit (GC doxantrazole (10 mg/kg): 7.2 ± 0.7; GSK143 (1 mg/kg): 7.6 ± 0.6), reduced inflammation and prevented recruitment of immune cells in the intestinal muscularis. In vitro: in PMCs, substance P (0-90 µM) and trinitrophenyl (0-4 µg/ml) induced a concentration-dependent release of ß-hexosaminidase. Pretreatment with doxantrazole and GSK143 (0.03-10 µM) concentration dependently blocked substance P and trinitrophenyl induced ß-hexosaminidase release. In addition, GSK143 was able to reduce cytokine expression in endotoxin-treated bone marrow derived macrophages in a concentration-dependent manner. CONCLUSIONS: The Syk inhibitor GSK143 reduces macrophage activation and mast cell degranulation in vitro. In addition, it inhibits manipulation-induced intestinal muscular inflammation and restores intestinal transit in mice. These findings suggest that Syk inhibition may be a new tool to shorten POI.


Assuntos
Compostos de Anilina/uso terapêutico , Íleus/prevenção & controle , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Complicações Pós-Operatórias/prevenção & controle , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Tirosina Quinases/antagonistas & inibidores , Pirimidinas/uso terapêutico , Compostos de Anilina/administração & dosagem , Compostos de Anilina/farmacologia , Animais , Degranulação Celular/efeitos dos fármacos , Células Cultivadas , Citocinas/biossíntese , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/métodos , Trânsito Gastrointestinal/efeitos dos fármacos , Íleus/fisiopatologia , Ativação de Macrófagos/efeitos dos fármacos , Mastócitos/efeitos dos fármacos , Mastócitos/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/antagonistas & inibidores , Ovalbumina/farmacologia , Inibidores de Fosfodiesterase/uso terapêutico , Complicações Pós-Operatórias/fisiopatologia , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/administração & dosagem , Pirimidinas/farmacologia , Substância P/antagonistas & inibidores , Substância P/farmacologia , Quinase Syk , Tioxantenos/uso terapêutico , Xantonas/uso terapêutico
7.
J Med Chem ; 65(22): 15174-15207, 2022 11 24.
Artigo em Inglês | MEDLINE | ID: mdl-36378954

RESUMO

The bromodomain and extra terminal (BET) family of proteins are an integral part of human epigenome regulation, the dysregulation of which is implicated in multiple oncology and inflammatory diseases. Disrupting the BET family bromodomain acetyl-lysine (KAc) histone protein-protein interaction with small-molecule KAc mimetics has proven to be a disease-relevant mechanism of action, and multiple molecules are currently undergoing oncology clinical trials. This work describes an efficiency analysis of published GSK pan-BET bromodomain inhibitors, which drove a strategic choice to focus on the identification of a ligand-efficient KAc mimetic with the hypothesis that lipophilic efficiency could be drastically improved during optimization. This focus drove the discovery of the highly ligand-efficient and structurally distinct benzoazepinone KAc mimetic. Following crystallography to identify suitable growth vectors, the benzoazepinone core was optimized through an explore-exploit structure-activity relationship (SAR) approach while carefully monitoring lipophilic efficiency to deliver I-BET432 (41) as an oral candidate quality molecule.


Assuntos
Lisina , Fatores de Transcrição , Humanos , Lisina/metabolismo , Ligantes , Domínios Proteicos , Histonas/metabolismo
8.
Bioorg Med Chem Lett ; 21(20): 6188-94, 2011 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-21903390
9.
Rapid Commun Mass Spectrom ; 23(22): 3501-12, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19844970

RESUMO

The knowledge of drug metabolism in the early phases of the drug discovery process is vital for minimising compound failure at later stages. As chemically reactive metabolites may cause adverse drug reactions, it is generally accepted that avoiding formation of reactive metabolites increases the chances of success of a molecule. In order to generate this important information, a screening strategy for the rapid detection of in vitro generated reactive metabolites trapped by glutathione has been developed. The bioassay incorporated the use of native glutathione and its close analogue the glutathione ethyl ester. The generic conditions for detecting glutathione conjugates that undergo constant neutral loss of 129 Da were optimised using a glutathione-based test mix of four compounds. The final liquid chromatography/tandem mass spectrometry constant neutral loss method used low-resolution settings and a scanning window of 200 amu. Data mining was rapidly and efficiently performed using LightSight software. Unambiguous identification of the glutathione conjugates was significantly facilitated by the analytical characteristics of the conjugate pairs formed with glutathione and glutathione ethyl ester, i.e. by chromatographic retention time and mass differences. The reliability and robustness of the screening strategy was tested using a number of compounds known to form reactive metabolites. Overall, the developed screening strategy provided comprehensive and reliable identification of glutathione conjugates and is well suited for rapid routine detection of trapped reactive metabolites. This new approach allowed the identification of a previously unreported diclofenac glutathione conjugate.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Glutationa/química , Espectrometria de Massas/métodos , Processamento Eletrônico de Dados , Software
10.
J Pharm Biomed Anal ; 49(1): 100-7, 2009 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-19019615

RESUMO

Recent years have seen the introduction of new high performance liquid chromatography (HPLC) instruments and columns that are capable of achieving high resolution, high speed liquid chromatographic separations at back pressures up to 1000 bar, so-called ultra-high performance liquid chromatography (UHPLC). Ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) is gaining widespread use for this purpose, and for this approach to be successful a generically applicable, robust column is required. Here, data are presented showing the robustness of a partially porous 2.7 microm diameter particle material in this application and the accuracy and precision of an assay for a typical pharmaceutical in plasma. This stationary phase material is evaluated for performance and compared with other materials frequently used for similar analyses using a test mix currently used routinely in our laboratories to assess the performance of UHPLC-MS/MS systems. The partially porous material demonstrates similar resolving power to sub-2 microm materials under the ballistic gradient chromatography conditions employed and exhibits excellent resilience over the analysis of thousands of protein precipitated plasma extracts. It is suggested that this stationary phase material can be an invaluable tool in generic, high throughput assays for pharmaceutical bioanalysts.


Assuntos
Cromatografia Líquida de Alta Pressão/instrumentação , Descoberta de Drogas , Medicamentos Genéricos/análise , Medicamentos Genéricos/farmacocinética , Espectrometria de Massas em Tandem/métodos , Bioensaio/métodos , Calibragem , Cromatografia Líquida de Alta Pressão/métodos , Tamanho da Partícula , Preparações Farmacêuticas/sangue , Porosidade , Reprodutibilidade dos Testes
11.
Br J Pharmacol ; 176(8): 1135-1142, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30735243

RESUMO

BACKGROUND AND PURPOSE: Chronic spontaneous urticaria presents as a heterogeneous syndrome characterised by wheals, angioedema, or both for greater than 6 weeks. Spleen tyrosine kinase mediates allergen-induced mast cell degranulation via the IgE signalling pathway, a central component of wheal formation and inflammation. In this study, we investigated the effects of perfused or topically administered GSK2646264 on IgE-mediated histamine release from mast cells in an ex vivo human skin model. EXPERIMENTAL APPROACH: Using a novel SkiP device, ex vivo human skin from mastectomy surgeries was challenged with anti-IgE, complement 5a (C5a), and buffer to induce histamine release from skin mast cells. Histamine was collected via microdialysis fibres and measured fluorometrically. GSK2646264 was delivered via perfusion either using microdialysis fibres or topically in a cream. Drug concentrations in the skin were measured by LC-MS, and a pharmacokinetic/ pharmacodynamic (PK/PD) relationship developed. KEY RESULTS: Perfused GSK2646264 significantly inhibited anti-IgE (but not C5a)-induced histamine release in a concentration-dependent manner. The 0.5, 1, and 3% cream delivered GSK2646264 to the dermis above the IC90 and dose-dependently attenuated anti-IgE-induced histamine release. CONCLUSIONS AND IMPLICATIONS: GSK2646264 administered topically or direct to the dermis blocked histamine release from in situ skin mast cells. A PK/PD relationship curve suggests that dermal concentrations above 6.8 µM should lead to approximately 90% inhibition of histamine release from skin mast cells following activation of the Fc fragment of IgE receptor 1a, implicating a potential use for the compound in skin mast cell diseases such as urticaria.


Assuntos
Liberação de Histamina/efeitos dos fármacos , Inibidores de Proteínas Quinases/administração & dosagem , Piridinas/administração & dosagem , Pele/efeitos dos fármacos , Quinase Syk/antagonistas & inibidores , Administração Tópica , Relação Dose-Resposta a Droga , Feminino , Histamina/metabolismo , Liberação de Histamina/fisiologia , Humanos , Microdiálise/métodos , Técnicas de Cultura de Órgãos , Pele/metabolismo , Quinase Syk/metabolismo
12.
Science ; 356(6345): 1397-1401, 2017 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-28619718

RESUMO

The success of new therapies hinges on our ability to understand their molecular and cellular mechanisms of action. We modified BET bromodomain inhibitors, an epigenetic-based therapy, to create functionally conserved compounds that are amenable to click chemistry and can be used as molecular probes in vitro and in vivo. We used click proteomics and click sequencing to explore the gene regulatory function of BRD4 (bromodomain containing protein 4) and the transcriptional changes induced by BET inhibitors. In our studies of mouse models of acute leukemia, we used high-resolution microscopy and flow cytometry to highlight the heterogeneity of drug activity within tumor cells located in different tissue compartments. We also demonstrate the differential distribution and effects of BET inhibitors in normal and malignant cells in vivo. This study provides a potential framework for the preclinical assessment of a wide range of drugs.


Assuntos
Benzodiazepinas/uso terapêutico , Química Click , Sistemas de Liberação de Medicamentos , Epigenômica , Leucemia/tratamento farmacológico , Animais , Benzodiazepinas/farmacologia , Células Cultivadas , Modelos Animais de Doenças , Leucemia/patologia , Camundongos , Medicina de Precisão , Distribuição Tecidual , Fatores de Transcrição/antagonistas & inibidores
13.
J Chromatogr A ; 1056(1-2): 3-12, 2004 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-15595526

RESUMO

Both LC and capillary LC (CapLC) have been successfully interfaced with inductively coupled plasma mass spectrometry (ICP-MS). Gradients of acetonitrile and aqueous based solvents have been employed to separate several compounds of pharmaceutical interest. This paper will describe four application areas in the pharmaceutical industry, and examples will be shown where CapLC, LC and gel electrophoresis via laser ablation have been coupled with ICP-MS. The four areas highlighted in this paper are: (1) the use of derivatisation reactions to "make the invisible visible". Methods involving derivatisations with copper and iron will be described that can be used for the analysis of amines and carboxylic acids by ICP-MS. (2) The profiling of metal ion content (in particular bromine) in biological samples such as human plasma, this study will focus on the metabolism of bromine-labelled peptides (e.g. substance P). (3) The analysis of materials derived from single, solid-phase beads used in combinatorial chemistry, and (4) also discussed will be our findings from investigations into the use of laser ablation ICP-MS on the determination of protein phosphorylation on electrophoresis gel blots.


Assuntos
Cromatografia Líquida/métodos , Espectrometria de Massas/métodos , Humanos , Sensibilidade e Especificidade , Substância P/sangue
14.
J Pept Sci ; 12(3): 220-6, 2006 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16163665

RESUMO

The coupling of the techniques, high-performance liquid chromatography (HPLC), orthogonal acceleration time-of-flight mass spectrometry (OATOF-MS) and inductively coupled plasma mass spectrometry (ICP-MS) provides a very powerful method for identifying and quantifying the products of bradykinin metabolism. In this study, we were able to identify the major metabolites of bradykinin degradation reported in the literature. In addition, a new bradykinin metabolite corresponding to bradykinin 5,9 fragment (BK-(5,9)-fragment) was identified as a product of neutral endopeptidase (NEP) activity. This finding establishes that NEP cleaves bradykinin simultaneously at the positions 4-5 and 7-8. We also demonstrate the equivalent participation of NEP and angiotensin-converting enzyme (ACE) within the rat lung tissue membranes (RLTM) in bradykinin degradation, suggesting its suitability as a model for the assay of dual ACE/NEP inhibitors. On the contrary, in rat kidney brush border membranes (KBBM), ACE is not significantly involved in bradykinin metabolism, with NEP being the major enzyme.


Assuntos
Bradicinina/metabolismo , Membrana Celular/metabolismo , Rim/metabolismo , Pulmão/metabolismo , Animais , Bradicinina/química , Membrana Celular/enzimologia , Cromatografia Líquida de Alta Pressão , Rim/química , Rim/enzimologia , Pulmão/química , Pulmão/enzimologia , Espectrometria de Massas/métodos , Microvilosidades/metabolismo , Neprilisina/antagonistas & inibidores , Neprilisina/química , Peptidil Dipeptidase A/química , Peptidil Dipeptidase A/efeitos dos fármacos , Piridinas/farmacologia , Ratos , Sensibilidade e Especificidade , Relação Estrutura-Atividade , Tiazepinas/farmacologia , Fatores de Tempo
15.
J Pept Sci ; 8(11): 591-600, 2002 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-12487427

RESUMO

Several studies have suggested that the accumulation of bradykinin, or that of one its metabolites BK1-8, is involved in the occurrence of side effects such as AE associated with the use of various ACEi. In this work a novel approach combining HPLC-UV on-line with oaTOF-MS and ICPMS was applied to investigate in human and rat plasma the metabolism of labelled BK (79/81 Br-Phe5) BrBK in the presence of two new dual ACE/NEP inhibitors (GW660511X and omapatrilat) currently under clinical trial. In human plasma the BrBK half-life values in the absence or in the presence of GW660511X (3.8 microM) or omapatrilat (32 nM) were 38.7 +/- 2.4, 51.2 +/- 4.7 and 114.7 +/- 9.3 min, respectively and BrBK was degraded into BrBK1-8, BrBK1-7, BrBK1-5 and Br-Phe. In the presence of inhibitors, however, the levels of these resultant metabolites were different. Unlike GW660511X, omapatrilat abolished the production of BrBK1-5 and BrBK1-7, suggesting a better ACE inhibition effect over GW660511X as no NEP activity was found. In addition the production of BrBK1-8 was enhanced in the presence of these inhibitors with a greater accumulation being observed with omapatrilat. The production of Br-Phe5 was reduced with GW660511X while no significant change was observed with omapatrilat after 4 h of incubation. In rat plasma the BrBK half-life values in the absence or in the presence of GW660511X (530 nM) or omapatrilat (50 nM) were 9.31 +/- 1.7, 22.06 +/- 3.1 and 25.3 +/- 1.7 min, respectively and BrBK was degraded into BrBK1-8, BrBK1-7, BrBK1-5 and Br-Phe5 plus BrBK2-9, BrBK4-8 and BrBK2-8 metabolites not found in human plasma. GW660511X and omapatrilat reduced the production of BrBK1-5 and BrBK1-7 with more effect being observed with omapatrilat. GW660511X and omapatrilat increased the production of both BrBK1-8 and Br-Phe5 but not that of BrBK4-8 and BrBK2-8. This study shows that the potency of GW660511X in comparison with omapatrilat is more than 100-fold lower in human, but less than 10-fold lower in rat plasma, suggesting that rat may not be a suitable in vivo model for the evaluation of ACE/NEP inhibition in relation to effects in humans.


Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Bradicinina/sangue , Bradicinina/metabolismo , Piridinas/farmacologia , Tiazepinas/farmacologia , Tiazóis/farmacologia , Sequência de Aminoácidos , Inibidores da Enzima Conversora de Angiotensina/química , Animais , Bradicinina/química , Humanos , Estrutura Molecular , Neprilisina/metabolismo , Piridinas/química , Ratos , Tiazepinas/química , Fatores de Tempo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA