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1.
Drug Des Devel Ther ; 18: 2653-2679, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38974119

RESUMO

Purpose: Over the last few years, covalent fragment-based drug discovery has gained significant importance. Thus, striving for more warhead diversity, we conceived a library consisting of 20 covalently reacting compounds. Our covalent fragment library (CovLib) contains four different warhead classes, including five α-cyanoacacrylamides/acrylates (CA), three epoxides (EO), four vinyl sulfones (VS), and eight electron-deficient heteroarenes with a leaving group (SNAr/SN). Methods: After predicting the theoretical solubility of the fragments by LogP and LogS during the selection process, we determined their experimental solubility using a turbidimetric solubility assay. The reactivities of the different compounds were measured in a high-throughput 5,5'-dithiobis-(2-nitrobenzoic acid) DTNB assay, followed by a (glutathione) GSH stability assay. We employed the CovLib in a (differential scanning fluorimetry) DSF-based screening against different targets: c-Jun N-terminal kinase 3 (JNK3), ubiquitin-specific protease 7 (USP7), and the tumor suppressor p53. Finally, the covalent binding was confirmed by intact protein mass spectrometry (MS). Results: In general, the purchased fragments turned out to be sufficiently soluble. Additionally, they covered a broad spectrum of reactivity. All investigated α-cyanoacrylamides/acrylates and all structurally confirmed epoxides turned out to be less reactive compounds, possibly due to steric hindrance and reversibility (for α-cyanoacrylamides/acrylates). The SNAr and vinyl sulfone fragments are either highly reactive or stable. DSF measurements with the different targets JNK3, USP7, and p53 identified reactive fragment hits causing a shift in the melting temperatures of the proteins. MS confirmed the covalent binding mode of all these fragments to USP7 and p53, while additionally identifying the SNAr-type electrophile SN002 as a mildly reactive covalent hit for p53. Conclusion: The screening and target evaluation of the CovLib revealed first interesting hits. The highly cysteine-reactive fragments VS004, SN001, SN006, and SN007 covalently modify several target proteins and showed distinct shifts in the melting temperatures up to +5.1 °C and -9.1 °C.


Assuntos
Proteína Quinase 10 Ativada por Mitógeno , Proteína Supressora de Tumor p53 , Peptidase 7 Específica de Ubiquitina , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/química , Peptidase 7 Específica de Ubiquitina/antagonistas & inibidores , Peptidase 7 Específica de Ubiquitina/metabolismo , Peptidase 7 Específica de Ubiquitina/química , Humanos , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 10 Ativada por Mitógeno/química , Sulfonas/química , Sulfonas/farmacologia , Estrutura Molecular , Solubilidade , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Relação Estrutura-Atividade , Acrilamidas/química , Acrilamidas/farmacologia , Acrilatos/química , Acrilatos/farmacologia , Ligação Proteica
2.
Eur J Med Chem ; 245(Pt 1): 114894, 2023 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-36343411

RESUMO

Despite innumerable efforts to develop effective therapeutics, it is difficult to achieve breakthrough treatments for Alzheimer's disease (AD), and the main reason is probably the absence of a clear target. Here, we reveal c-Jun N-terminal kinase 3 (JNK3), a protein kinase explicitly expressed in the brain and involved in neuronal apoptosis, with a view toward providing effective treatment for AD. For many years, we have worked on JNK3 inhibitors and have discovered 2-aryl-1-pyrimidinyl-1H-imidazole-5-yl acetonitrile-based JNK3 inhibitors with superb potency (IC50 < 1.0 nM) and excellent selectivity over other protein kinases including isoforms JNK1 (>300 fold) and JNK2 (∼10 fold). Based on in vitro biological activity and DMPK properties, the lead compounds were selected for further in vivo studies. We confirmed that repeat administration of JNK3 inhibitors improved cognitive memory in APP/PS1 and the 3xTg mouse model. Overall, our results show that JNK3 could be a potential target protein for AD.


Assuntos
Doença de Alzheimer , Imidazóis , Proteína Quinase 10 Ativada por Mitógeno , Inibidores de Proteínas Quinases , Animais , Camundongos , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/enzimologia , Apoptose/efeitos dos fármacos , Imidazóis/química , Imidazóis/farmacologia , Imidazóis/uso terapêutico , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Isoformas de Proteínas/antagonistas & inibidores , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Modelos Animais de Doenças
3.
Int J Mol Sci ; 22(20)2021 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-34681742

RESUMO

As members of the MAPK family, c-Jun-N-terminal kinases (JNKs) regulate the biological processes of apoptosis. In particular, the isoform JNK3 is expressed explicitly in the brain at high levels and is involved in the pathogenesis of neurodegenerative diseases such as Alzheimer's disease (AD) and Parkinson's disease (PD). In this study, we prepared a series of five 6-dihydroxy-1H-benzo[d]imidazoles as JNK3 inhibitors and found them have potential as neuroprotective agents. Following a previous lead scaffold, benzimidazole moiety was modified with various aryl groups and hydroxylation, and the resulting compounds exhibited JNK3 inhibitory activity with improved potency and selectivity. Out of 37 analogues synthesized, (S)-cyclopropyl(3-((4-(2-(2,3-dihydrobenzo[b][1,4]dioxin -6-yl)-5,6-dihydroxy-1H-benzo[d]imidazol-1-yl)pyrimidin-2-yl)amino) piperidin-1-yl)methanone (35b) demonstrated the highest JNK3 inhibition (IC50 = 9.7 nM), as well as neuroprotective effects against Aß-induced neuronal cell death. As a protein kinase inhibitor, it also showed excellent selectivity over other protein kinases including isoforms JNK1 (>1000 fold) and JNK2 (-10 fold).


Assuntos
Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/farmacologia , Peptídeos beta-Amiloides/toxicidade , Animais , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Ligação de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Proteína Quinase 10 Ativada por Mitógeno/química , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Modelos Moleculares , Neurônios/patologia , Síndromes Neurotóxicas/prevenção & controle , Fragmentos de Peptídeos/toxicidade , Ratos
4.
Int J Mol Sci ; 22(7)2021 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-33918172

RESUMO

Current treatments for neurodegenerative diseases (ND) are symptomatic and do not affect disease progression. Slowing this progression remains a crucial unmet need for patients and their families. c-Jun N-terminal kinase 3 (JNK3) are related to several ND hallmarks including apoptosis, oxidative stress, excitotoxicity, mitochondrial dysfunction, and neuroinflammation. JNK inhibitors can play an important role in addressing neuroprotection. This research aims to evaluate the neuroprotective, anti-inflammatory, and antioxidant effects of a synthetic compound (FMU200) with known JNK3 inhibitory activity in SH-SY5Y and RAW264.7 cell lines. SH-SY5Y cells were pretreated with FMU200 and cell damage was induced by 6-hydroxydopamine (6-OHDA) or hydrogen peroxide (H2O2). Cell viability and neuroprotective effect were assessed with an MTT assay. Flow cytometric analysis was performed to evaluate cell apoptosis. The H2O2-induced reactive oxygen species (ROS) generation and mitochondrial membrane potential (ΔΨm) were evaluated by DCFDA and JC-1 assays, respectively. The anti-inflammatory effect was determined in LPS-induced RAW264.7 cells by ELISA assay. In undifferentiated SH-SY5Y cells, FMU200 decreased neurotoxicity induced by 6-OHDA in approximately 20%. In RA-differentiated cells, FMU200 diminished cell death in approximately 40% and 90% after 24 and 48 h treatment, respectively. FMU200 reduced both early and late apoptotic cells, decreased ROS levels, restored mitochondrial membrane potential, and downregulated JNK phosphorylation after H2O2 exposure. In LPS-stimulated RAW264.7 cells, FMU200 reduced TNF-α levels after a 3 h treatment. FMU200 protects neuroblastoma SH-SY5Y cells against 6-OHDA- and H2O2-induced apoptosis, which may result from suppressing the JNK pathways. Our findings show that FMU200 can be a useful candidate for the treatment of neurodegenerative disorders.


Assuntos
Potencial da Membrana Mitocondrial/efeitos dos fármacos , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Doenças Neurodegenerativas/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Humanos , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Células RAW 264.7
5.
Cell Mol Life Sci ; 78(1): 287-298, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32189007

RESUMO

Unveiling the key pathways underlying postnatal beta-cell proliferation can be instrumental to decipher the mechanisms of beta-cell mass plasticity to increased physiological demand of insulin during weight gain and pregnancy. Using transcriptome and global Serine Threonine Kinase activity (STK) analyses of islets from newborn (10 days old) and adult rats, we found that highly proliferative neonatal rat islet cells display a substantially elevated activity of the mitogen activated protein 3 kinase 12, also called dual leucine zipper-bearing kinase (Dlk). As a key upstream component of the c-Jun amino terminal kinase (Jnk) pathway, Dlk overexpression was associated with increased Jnk3 activity and was mainly localized in the beta-cell cytoplasm. We provide the evidence that Dlk associates with and activates Jnk3, and that this cascade stimulates the expression of Ccnd1 and Ccnd2, two essential cyclins controlling postnatal beta-cell replication. Silencing of Dlk or of Jnk3 in neonatal islet cells dramatically hampered primary beta-cell replication and the expression of the two cyclins. Moreover, the expression of Dlk, Jnk3, Ccnd1 and Ccnd2 was induced in high replicative islet beta cells from ob/ob mice during weight gain, and from pregnant female rats. In human islets from non-diabetic obese individuals, DLK expression was also cytoplasmic and the rise of the mRNA level was associated with an increase of JNK3, CCND1 and CCND2 mRNA levels, when compared to islets from lean and obese patients with diabetes. In conclusion, we find that activation of Jnk3 signalling by Dlk could be a key mechanism for adapting islet beta-cell mass during postnatal development and weight gain.


Assuntos
Células Secretoras de Insulina/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Transdução de Sinais , Animais , Proliferação de Células/efeitos dos fármacos , Ciclina D1/genética , Ciclina D1/metabolismo , Ciclina D2/genética , Ciclina D2/metabolismo , Feminino , Glucose/farmacologia , Humanos , Insulina/metabolismo , Células Secretoras de Insulina/citologia , MAP Quinase Quinase Quinases/antagonistas & inibidores , MAP Quinase Quinase Quinases/genética , Camundongos , Camundongos Endogâmicos C57BL , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 10 Ativada por Mitógeno/genética , Obesidade/metabolismo , Obesidade/patologia , Pâncreas/crescimento & desenvolvimento , Pâncreas/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos
6.
Eur J Med Chem ; 201: 112445, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32603981

RESUMO

The c-Jun N-terminal kinase 3 (JNK3) plays key roles in a wide range of diseases, including neurodegeneration diseases, inflammation diseases, cancers, cardiovascular diseases, and metabolic disorders. Previously, we have identified a lead compound, (Z)-3-(2-(naphthalen-1-yl)-2-oxoethylidene)-3,4-dihydroquinoxalin-2(1H)-one (J46), which contains a 3,4-dihydroquinoxalin-2(1H)-one core structure as a key fragment to inhibit JNK3. However, compound J46 displayed high DDR1 and EGFR (T790M, L858R) inhibition and poor physicochemical properties, especially clogD and water-solubility, in its biological studies. Herein, we optimized compound J46 by structure-based drug design and exploiting the selectivity and physicochemical properties of various warhead groups to obtain compound J46-37, which not only exhibited a potent inhibition against JNK3 but also showed more than 50-fold potency better than DDR1 and EGFR (T790M, L858R). Furthermore, the selectivity and structure-activity relationship of novel synthesized 3,4-dihydroquinoxalin-2(1H)-one derivatives were analyzed by molecular docking and molecular dynamics simulation. Overall, compound J46-37, as a highly selective inhibitor of JNK3 with well physicochemical properties, is worth developing as therapies for the treatment of diseases related to JNK3.


Assuntos
Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/química , Quinoxalinas/química , Ensaios Enzimáticos , Humanos , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Estrutura Molecular , Ligação Proteica , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/metabolismo , Quinoxalinas/síntese química , Quinoxalinas/metabolismo , Relação Estrutura-Atividade
7.
Cells ; 9(6)2020 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-32580272

RESUMO

Sirtuin 6 (SIRT6), as a NAD + -dependent deacetylase, plays an indispensable role in the regulation of health and physiology. Loss of SIRT6 causes spontaneous colitis in mice and makes intestinal epithelial cells prone to stress. However, whether SIRT6 overexpression increases resistance to colitis remains unknown. Here, in vivo results demonstrated that SIRT6 overexpression attenuates DSS-induced colitis in terms of clinical manifestations, histopathological damage, loss of tight junction function and imbalanced intestinal microenvironment. Additionally, we also found that the activation of NF-κB and c-Jun induced by DSS is diminished by SIRT6 overexpression. Furthermore, SIRT6 may regulate TAK1 to inhibit NF-κB and c-Jun signaling. Thus, our findings highlight the protective effect of SIRT6 on colon, further supporting the perspective that SIRT6 may be a therapeutic target for intestine injury under stress.


Assuntos
Colite/metabolismo , Sirtuínas/biossíntese , Animais , Colite/induzido quimicamente , Colite/genética , Colite/prevenção & controle , Modelos Animais de Doenças , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Transdução de Sinais , Sirtuínas/genética , Sirtuínas/metabolismo
8.
Int J Mol Sci ; 21(5)2020 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-32131443

RESUMO

We designed and synthesized 1-pyrimidinyl-2-aryl-4, 6-dihydropyrrolo [3,4-d] imidazole-5(1H)-carboxamide derivatives as selective inhibitors of c-Jun-N-terminal Kinase 3 (JNK3), a target for the treatment of neurodegenerative diseases. Based on the compounds found in previous studies, a novel scaffold was designed to improve pharmacokinetic characters and activity, and compound 18a, (R)-1-(2-((1-(cyclopropanecarbonyl)pyrrolidin-3-yl)amino)pyrimidin-4-yl)-2-(3,4-dichlorophenyl)-4,6-dihydro pyrrolo [3,4-d]imidazole-5(1H)-carboxamide, showed the highest IC50 value of 2.69 nM. Kinase profiling results also showed high selectivity for JNK3 among 38 kinases, having mild activity against JNK2, RIPK3, and GSK3ß, which also known to involve in neuronal apoptosis.


Assuntos
Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/síntese química , Sítios de Ligação , Imidazóis/química , Proteína Quinase 10 Ativada por Mitógeno/química , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Ligação Proteica , Inibidores de Proteínas Quinases/farmacologia
9.
J Comput Aided Mol Des ; 34(6): 671-682, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32040807

RESUMO

The c-Jun N-terminal kinase 3 (JNK3) signaling cascade is activated during cerebral ischemia leading to neuronal damage. The present study was carried out to identify and evaluate novel JNK3 inhibitors using in-silico and in-vitro approach. A total of 380 JNK3 inhibitors belonging to different organic groups was collected from the previously reported literature. These molecules were used to generate a pharmacophore model. This model was used to screen a chemical database (SPECS) to identify newer molecules with similar chemical features. The top 1000 hits molecules were then docked against the JNK3 enzyme coordinate following GLIDE rigid receptor docking (RRD) protocol. Best posed molecules of RRD were used during induced-fit docking (IFD), allowing receptor flexibility. Other computational predictions such as binding free energy, electronic configuration and ADME/tox were also calculated. Inferences from the best pharmacophore model suggested that, in order to have specific JNK3 inhibitory activity, the molecules must possess one H-bond donor, two hydrophobic and two ring features. Docking studies suggested that the main interaction between lead molecules and JNK3 enzyme consisted of hydrogen bond interaction with methionine 149 of the hinge region. It was also observed that the molecule with better MM-GBSA dG binding free energy, had greater correlation with JNK3 inhibition. Lead molecule (AJ-292-42151532) with the highest binding free energy (dG = 106.8 Kcal/mol) showed better efficacy than the SP600125 (reference JNK3 inhibitor) during cell-free JNK3 kinase assay (IC50 = 58.17 nM) and cell-based neuroprotective assay (EC50 = 7.5 µM).


Assuntos
Proteína Quinase 10 Ativada por Mitógeno/química , Fármacos Neuroprotetores/química , Compostos Orgânicos/química , Inibidores de Proteínas Quinases/química , Isquemia Encefálica/tratamento farmacológico , Isquemia Encefálica/enzimologia , Isquemia Encefálica/patologia , Avaliação Pré-Clínica de Medicamentos , Humanos , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Neurônios/patologia , Ligação Proteica/efeitos dos fármacos , Relação Estrutura-Atividade , Interface Usuário-Computador
10.
J Enzyme Inhib Med Chem ; 35(1): 372-376, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31856610

RESUMO

3-alkyl-5-aryl-1-pyrimidyl-1H-pyrazole derivatives were designed and synthesised as selective inhibitors of JNK3, a target for the treatment of neurodegenerative diseases. Following previous studies, we have designed JNK3 inhibitors to reduce the molecular weight and successfully identified a lead compound that exhibits equipotent activity towards JNK3. Kinase profiling results also showed high selectivity for JNK3 among 38 kinases. Among the derivatives, the IC50 value of 8a, (R)-2-(1-(2-((1-(cyclopropanecarbonyl)pyrrolidin-3-yl)amino)pyrimidin-4-yl)-5-(3,4-dichlorophenyl)-1H-pyrazol-3-yl)acetonitrile exhibited 227 nM, showing the highest inhibitory activity against JNK3.


Assuntos
Descoberta de Drogas , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Pirazóis/farmacologia , Relação Dose-Resposta a Droga , Humanos , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Simulação de Acoplamento Molecular , Estrutura Molecular , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Pirazóis/síntese química , Pirazóis/química , Relação Estrutura-Atividade
11.
J Med Chem ; 62(14): 6645-6664, 2019 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-31268308

RESUMO

Alzheimer's disease (AD) is one of the most challenging diseases around the world with no effective clinical treatment. Previous studies have suggested c-Jun N-terminal kinase 3 (JNK3) as an attractive therapeutic target for AD. Herein, we report 3-substituted indolin-2-one derivatives as the first isoform-selective JNK3 inhibitors by multistage screening. In this study, comparative structure-based virtual screening was performed, and J30-8 was identified with a half-maximal inhibitory concentration of 40 nM, which exhibited over 2500-fold isoform selectivity and marked kinome-wide selectivity. Further study indicated that 1 µM J30-8 exhibited neuroprotective activity in vitro so as to alleviate the spatial memory impairment in vivo through reducing plaque burden and inhibiting the phosphorylation of JNKs, Aß precursor protein, and Tau protein. All of these indicated J30-8 as proved isoform-selective JNK3 inhibitors that might serve as a useful tool for further JNK3 studies with AD as well as for the development of JNK3 inhibitors for the potential treatment of neurodegenerative diseases.


Assuntos
Indóis/química , Indóis/farmacologia , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Animais , Desenho de Fármacos , Descoberta de Drogas , Feminino , Humanos , Indóis/farmacocinética , Camundongos , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Modelos Moleculares , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/metabolismo , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/farmacocinética , Fármacos Neuroprotetores/farmacologia , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacocinética
12.
J Mol Graph Model ; 91: 30-51, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31158642

RESUMO

The kinase c-Jun N-terminal Kinase 3 (JNK3) plays an important role in neurodegenerative diseases. JNK3 inhibitors have shown promising results in treating Alzheimer's and Parkinson's diseases. This prompted us to model this interesting target via three established structure-based computational workflows; namely, docking-based Comparative Intermolecular Contacts Analysis (db-CICA), pharmacophore modeling via molecular-dynamics based Ligand-Receptor Contact Analysis (md-LRCA), and QSAR-guided selection of crystallographic pharmacophores. Moreover, we compared the performances of resulting pharmacophores against binding models generated via a newly introduced technique, namely, QSAR-guided selection of docking-based pharmacophores. The resulting pharmacophores were validated by receiver operating characteristic (ROC) curve analysis and used as virtual search queries to screen the National Cancer Institute (NCI) database for promising anti-JNK3 hits of novel chemotypes. Eleven nanomolar and low micromolar hits were identified, three of which were captured by QSAR-guided docking-based pharmacophores.


Assuntos
Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Simulação de Acoplamento Molecular , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Relação Quantitativa Estrutura-Atividade , Sítios de Ligação , Humanos , Ligantes , Proteína Quinase 10 Ativada por Mitógeno/química , Simulação de Dinâmica Molecular , Curva ROC
13.
J Mol Graph Model ; 90: 235-242, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31103916

RESUMO

In this work, the application of a new strategy called NWFE ensemble (nonparametric weighted feature extraction ensemble) method is proposed. Subspace-supervised projections based on NWFE are incorporated into the construction of ensembles of classifiers to facilitate the correct classification of wrongly classified instances without being detrimental to the overall performance of the ensemble. The performance of NWFE is investigated with a c-Jun N-terminal kinase-3 inhibitor benchmark dataset using different chemical compound representation models. Compared with the standard method, the results obtained show that the applied method improves the prediction performance using two classifiers based on decision trees and support vector machines.


Assuntos
Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Algoritmos , Árvores de Decisões , Humanos , Máquina de Vetores de Suporte
14.
Inflammopharmacology ; 27(6): 1217-1227, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31037574

RESUMO

BACKGROUND: Mitogen-activated protein kinase (MAPK) signaling plays an important role in inflammatory diseases such as rheumatoid arthritis (RA).The aim of our study was to elucidate the therapeutic potential of the highly selective p38 MAPK inhibitor Skepinone-L and the dual inhibitor LN 950 (p38 MAPK and JNK 3) in the K/BxN serum transfer model of RA. Additionally, we aimed to monitor MAPK treatment non-invasively in vivo using the hypoxia tracer [18F]fluoromisonidazole ([18F]FMISO) and positron emission tomography (PET). METHODS: To induce experimental arthritis, we injected glucose-6-phosphate isomerase autoantibody-containing serum in BALB/c mice. MAPK inhibitor or Sham treatment was administered per os once daily. On days 3 and 6 after arthritis induction, we conducted PET imaging with [18F]FMISO. At the end of the experiment, ankles were harvested for histopathological analysis. RESULTS: Skepinone-L and LN 950 were applicable to suppress the severity of experimental arthritis confirmed by reduced ankle swelling and histopathological analysis. Skepinone-L (3.18 ± 0.19 mm) and LN 950 (3.40 ± 0.13 mm) treatment yielded a significantly reduced ankle thickness compared to Sham-treated mice (3.62 ± 0.11 mm) on day 5 after autoantibody transfer, a time-point characterized by severe arthritis. Hypoxia imaging with [18F]FMISO revealed non-conclusive results and might not be an appropriate tool to monitor MAPK therapy in experimental RA. CONCLUSION: Both the selective p38 MAPK inhibitor Skepinone-L and the dual (p38 MAPK and JNK 3) inhibitor LN 950 exhibited significant therapeutic effects during experimental arthritis. Thus, our study contributes to the ongoing discussion on the use of p38 MAPK as a potential target in RA.


Assuntos
Artrite Experimental/tratamento farmacológico , Dibenzocicloeptenos/uso terapêutico , Imidazóis/uso terapêutico , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/uso terapêutico , Piridinas/uso terapêutico , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Animais , Artrite Experimental/diagnóstico por imagem , Dibenzocicloeptenos/farmacologia , Modelos Animais de Doenças , Glucose-6-Fosfato Isomerase/imunologia , Imidazóis/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Misonidazol/análogos & derivados , Misonidazol/farmacocinética , Tomografia por Emissão de Pósitrons , Piridinas/farmacologia
15.
Anal Biochem ; 532: 26-28, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28552758

RESUMO

In order to evaluate the isoform selectivity of novel inhibitors within the c-Jun N-terminal kinase (JNK) family, a fluorescence polarization-based competition binding assay, previously developed for JNK3, was extended to the other isoforms JNK1 and JNK2. The assay is based on the displacement of a versatile fluorescent pyridinylimidazole-based probe and was validated by testing the precursor of the probe as well as standard JNK inhibitors.


Assuntos
Polarização de Fluorescência , Corantes Fluorescentes/metabolismo , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Proteína Quinase 9 Ativada por Mitógeno/metabolismo , Inibidores de Proteínas Quinases/metabolismo , Ligação Competitiva , Humanos , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 8 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 9 Ativada por Mitógeno/antagonistas & inibidores , Ligação Proteica
16.
Acta Biomater ; 53: 59-69, 2017 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-28216301

RESUMO

In order to identify the mechanisms by which skeletal maturity alters the mechanosensitivity of mesenchymal stromal cells (MSCs) and, the implications for osteogenesis and angiogenesis during bone formation, we compared the response of MSCs derived from children and skeletally-mature healthy adults cultured on soft and stiff collagen-coated polyacrylamide substrates. MSCs from children were more mechanosensitive, showing enhanced angiogenesis and osteogenesis on stiff substrates as indicated by increased endothelial tubule formation, PGF production, nuclear-translocation of YAP, ALP activity and mineralisation. To examine these mechanisms in more detail, a customised PCR array identified an age-dependent, stiffness-induced upregulation of NOX1, VEGFR1, VEGFR2, WIF1 and, of particular interest, JNK3 in cells from children compared to adults. When JNK3 activity was inhibited, a reduction in stiffness-induced driven osteogenesis was observed - suggesting that JNK3 might serve as a novel target for recapitulating the enhanced regenerative potential of children in adults suffering from bone degeneration. STATEMENT OF SIGNIFICANCE: We investigated the age-associated changes in the capacity of MSCs for bone regeneration involving the mechanosensitive signalling pathways, which reduce the ability of adult cells to respond to biophysical cues in comparison to cells from children, who are still undergoing bone development. Our results offer new insights into the mechanobiology of MSCs and sheds new light on age-altered mechanosensitivity and, on why children have such an immense capacity to regenerate their skeletal system. We have identified the mechanisms by which skeletal maturity alters the mechanosensitivity of mesenchymal stromal cells and an age-dependent, stiffness-induced upregulation of a number of prominent genes including, most notably, JNK3 in children cells, thus suggesting its potential to promote enhanced bone repair.


Assuntos
Envelhecimento/fisiologia , Mecanotransdução Celular , Células-Tronco Mesenquimais/fisiologia , Neovascularização Fisiológica , Osteogênese , Transporte Ativo do Núcleo Celular , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Envelhecimento/genética , Fenômenos Biomecânicos , Regeneração Óssea , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Células Cultivadas , Criança , Humanos , Sistema de Sinalização das MAP Quinases , Células-Tronco Mesenquimais/citologia , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 10 Ativada por Mitógeno/genética , Fosfoproteínas/metabolismo , Fator de Crescimento Placentário/genética , Fator de Crescimento Placentário/metabolismo , Fatores de Transcrição , Regulação para Cima , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Proteínas de Sinalização YAP , Adulto Jovem
17.
Ross Fiziol Zh Im I M Sechenova ; 103(3): 268-83, 2017 Mar.
Artigo em Russo | MEDLINE | ID: mdl-30199207

RESUMO

The article reviews the literature regarding the role of c-Jun-N-terminal kinases (JNK) and its inhibitors in brain damage in the settings of ischemia and reperfusion injury. The implication of JNK in signaling mechanisms involved in ischemia-reperfusion-induced cerebral injury are discussed. Described effects associated with JNK inhibition using synthetic and natural substances in experimental models of ischemic and reperfusion injury of the brain. Results of experimental studies demonstrated that JNK represent promising therapeutic targets for brain protection against ischemic stroke. However, multiple physiologic functions of various JNK family members do not allow for the systemic use of non-specific JNK inhibitors for therapeutic purposes. The authors conclude that the continuous search for selective inhibitors of JNK3 remains an important task.


Assuntos
Isquemia Encefálica/tratamento farmacológico , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Fármacos Neuroprotetores/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Acetonitrilas/farmacologia , Animais , Antracenos/farmacologia , Benzotiazóis/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Encéfalo/patologia , Isquemia Encefálica/enzimologia , Isquemia Encefálica/genética , Isquemia Encefálica/patologia , Regulação da Expressão Gênica , Ginsenosídeos/isolamento & purificação , Ginsenosídeos/farmacologia , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 10 Ativada por Mitógeno/genética , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Oximas/farmacologia , Extratos Vegetais/química , Quinoxalinas/farmacologia , Traumatismo por Reperfusão/enzimologia , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/patologia , Transdução de Sinais
18.
J Med Chem ; 60(2): 594-607, 2017 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-27977190

RESUMO

The concept of covalent inhibition of c-Jun N-terminal kinase 3 (JNK3) was successfully transferred to our well validated pyridinylimidazole scaffold varying several structural features in order to deduce crucial structure-activity relationships. Joint targeting of the hydrophobic region I and methylation of imidazole-N1 position increased the activity and reduced the number of off-targets. The most promising covalent inhibitor, the tetrasubstituted imidazole 3-acrylamido-N-(4-((4-(4-(4-fluorophenyl)-1-methyl-2-(methylthio)-1H-imidazol-5-yl)pyridin-2-yl)amino)phenyl)benzamide (7) inhibits the JNK3 in the subnanomolar range (IC50 = 0.3 nM), shows high metabolic stability in human liver microsomes, and displays excellent selectivity in a screening against a panel of 410 kinases. Covalent bond formation to Cys-154 was confirmed by incubation of the inhibitors with wild-type JNK3 and JNK3-C154A mutant followed by mass spectrometry.


Assuntos
Acrilamidas/farmacologia , Benzamidas/farmacologia , Imidazóis/farmacologia , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Acrilamidas/síntese química , Acrilamidas/química , Benzamidas/síntese química , Benzamidas/química , Humanos , Imidazóis/síntese química , Imidazóis/química , Masculino , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Piridinas/síntese química , Piridinas/química , Relação Estrutura-Atividade
19.
J Neurotrauma ; 34(3): 695-706, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27526795

RESUMO

The mechanism by which spinal cord injury (SCI) induces neuronal death has not been thoroughly understood. Investigation on the molecular signal pathways involved in SCI-mediated neuronal apoptosis is important for development of new therapeutics for SCI. In the current study, we explore the role of heme oxygenase-1 (HO-1) in the modulation of mixed lineage kinase 3/mitogen-activated protein kinase kinase/cJUN N-terminal kinase 3 (MLK3/MKK7/JNK3) signaling, which is a pro-apoptotic pathway, after SCI. We found that MLK3/MKK7/JNK3 signaling was activated by SCI in a time-dependent manner, demonstrated by increase in activating phosphorylation of MLK3, MKK7, and JNK3. SCI also induced HO-1 expression. Administration of HO-1-expressing adeno-associated virus before SCI introduced expression of exogenous HO-1 in injured spinal cords. Exogenous HO-1 reduced phosphorylation of MLK3, MKK7, and JNK3. Consistent with its inhibitory effect on MLK3/MKK7/JNK3 signaling, exogenous HO-1 decreased SCI-induced neuronal apoptosis and improved neurological score. Further, we found that exogenous HO-1 inhibited expression of cell division cycle 42 (Cdc42), which is crucial for MLK3 activation. In vitro experiments indicated that Cdc42 was essential for neuronal apoptosis, while transduction of neurons with HO-1-expressing adeno-associated virus significantly reduced neuronal apoptosis to enhance neuronal survival. Therefore, our study disclosed a novel mechanism by which HO-1 exerted its neuroprotective efficacy. Our discovery might be valuable for developing a new therapeutic approach for SCI.


Assuntos
Heme Oxigenase (Desciclizante)/biossíntese , MAP Quinase Quinase 7/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Traumatismos da Medula Espinal/metabolismo , Proteína cdc42 de Ligação ao GTP/metabolismo , Animais , Apoptose/fisiologia , Células Cultivadas , Regulação para Baixo/fisiologia , MAP Quinase Quinase 7/antagonistas & inibidores , MAP Quinase Quinase Quinases/antagonistas & inibidores , Masculino , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Neurônios/metabolismo , Neurônios/patologia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , Traumatismos da Medula Espinal/patologia , Proteína cdc42 de Ligação ao GTP/antagonistas & inibidores , MAP Quinase Quinase Quinase 11 Ativada por Mitógeno
20.
Brain Res ; 1653: 67-74, 2016 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-27769787

RESUMO

Cilostazol(CTL) is a phosphodiesterase inhibitor, which has been widely used as anti-platelet agent. It also has preventive effects on various central nervous system (CNS) diseases, including ischemic stroke, Parkinson's disease and Alzheimer disease. However, the molecular mechanism underlying the protective effects of CTL is still unclear, and whether CTL can prevent I/R induced cognitive deficit has not been reported. Transient global brain ischemia was induced by 4-vessel occlusion in adult male Sprague-Dawley rats. The open field tasks and Morris water maze were used to assess the effect of CTL on anxiety-like behavioral and cognitive impairment after I/R. Western blotting were performed to examine the expression of related proteins, and HE-staining was used to detect the percentage of neuronal death in the hippocampal CA1 region. Here we found that CTL significantly improved cognitive deficits and the behavior of rats in Morris water maze and open field tasks (P<0.05). HE staining results showed that CTL could significantly protect CA1 neurons against cerebral I/R (P<0.05). Additionally, Akt1 phosphorylation levels were evidently up-regulated (P<0.05), while the activation of JNK3, which is an important contributor to I/R-induced neuron apoptosis, was reduced by CTL after I/R (P<0.05), and caspase-3 levels were also decreased by CTL treatment. Furthermore, all of CTL's protective effects were reversed by LY294002, which is a PI3K/Akt1 inhibitor. Taken together, our results suggest that CTL could protect hippocampal neurons and ameliorate the impairment of learning/memory abilities and locomotor/ exploratory activities in ischemic stroke via a PI3K-Akt1/JNK3/caspase-3 dependent mechanism.


Assuntos
Isquemia Encefálica/tratamento farmacológico , Transtornos Cognitivos/tratamento farmacológico , Hipocampo/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Tetrazóis/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Isquemia Encefálica/complicações , Isquemia Encefálica/enzimologia , Isquemia Encefálica/patologia , Caspase 3/metabolismo , Cilostazol , Transtornos Cognitivos/enzimologia , Transtornos Cognitivos/etiologia , Transtornos Cognitivos/patologia , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/farmacologia , Hipocampo/enzimologia , Hipocampo/patologia , Masculino , Proteína Quinase 10 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Neurônios/patologia , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Sprague-Dawley , Traumatismo por Reperfusão/complicações , Traumatismo por Reperfusão/enzimologia , Traumatismo por Reperfusão/patologia
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