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This study aimed to use a computational approach that combined the classification-based QSAR model, molecular docking, ADME studies, and molecular dynamics (MD) to identify potential inhibitors of Fyn kinase. First, a robust classification model was developed from a dataset of 1,078 compounds with known Fyn kinase inhibitory activity, using the XGBoost algorithm. After that, molecular docking was performed between potential compounds identified from the QSAR model and Fyn kinase to assess their binding strengths and key interactions, followed by MD simulations. ADME studies were additionally conducted to preliminarily evaluate the pharmacokinetics and drug-like characteristics of these compounds. The results showed that our obtained model exhibited good predictive performance with an accuracy of 0.95 on the test set, affirming its reliability in identifying potent Fyn kinase inhibitors. Through the application of this model in conjunction with molecular docking and ADME studies, nine compounds were identified as potential Fyn kinase inhibitors, including 208 (ZINC70708110), 728 (ZINC8792432), 734 (ZINC8792187), 736 (ZINC8792350), 738 (ZINC8792286), 739 (ZINC8792309), 817 (ZINC33901069), 852 (ZINC20759145), and 1227 (ZINC100006936). MD simulations further demonstrated that the four most promising compounds, 728, 734, 736, and 852 exhibited stable binding with Fyn kinase during the simulation process. Additionally, a web-based platform ( https://fynkinase.streamlit.app/ ) has been developed to streamline the screening process. This platform enables users to predict the activity of their substances of interest on Fyn kinase from their SMILES, using our classification-based QSAR model and molecular docking.
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In this study, we explored the potential of novel inhibitors for FYN kinase, a critical target in cancer and neurodegenerative disorders, by integrating advanced cheminformatics, machine learning, and molecular simulation techniques. Our approach involved analyzing key interactions for FYN inhibition using established multi-kinase inhibitors such as Staurosporine, Dasatinib, and Saracatinib. We utilized ECFP4 circular fingerprints and the t-SNE machine learning algorithm to compare molecular similarities between FDA-approved drugs and known clinical trial inhibitors. This led to the identification of potential inhibitors, including Afatinib, Copanlisib, and Vandetanib. Using the DrugSpaceX platform, we generated a vast library of 72,196 analogues from these leads, which after careful refinement, resulted in 6008 promising candidates. Subsequent clustering identified 48 analogues with significant similarity to known inhibitors. Notably, two candidates derived from Vandetanib, DE27123047 and DE27123035, exhibited strong docking affinities and stable binding in molecular dynamics simulations. These candidates showed high potential as effective FYN kinase inhibitors, as evidenced by MMGBSA calculations and MCE-18 scores exceeding 50. Additionally, our exploration into their molecular architecture revealed potential modification sites on the quinazolin-4-amine scaffold, suggesting opportunities for strategic alterations to enhance activity and optimize ADME properties. Our research is a pioneering effort in drug discovery, unveiling novel candidates for FYN inhibition and demonstrating the efficacy of a multi-layered computational strategy. The molecular insights gained provide a pathway for strategic refinements and future experimental validations, setting a new direction in targeted drug development against diseases involving FYN kinase.
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Currently, no effective therapeutics exist for the treatment of incurable neurodegenerative diseases such as Alzheimer's disease (AD). The cellular prion protein (PrPC) acts as a high-affinity receptor for amyloid beta oligomers (AßO), a main neurotoxic species mediating AD pathology. The interaction of AßO with PrPC subsequently activates Fyn tyrosine kinase and neuroinflammation. Herein, we used our previously developed peptide aptamer 8 (PA8) binding to PrPC as a therapeutic to target the AßO-PrP-Fyn axis and prevent its associated pathologies. Our in vitro results indicated that PA8 prevents the binding of AßO with PrPC and reduces AßO-induced neurotoxicity in mouse neuroblastoma N2a cells and primary hippocampal neurons. Next, we performed in vivo experiments using the transgenic 5XFAD mouse model of AD. The 5XFAD mice were treated with PA8 and its scaffold protein thioredoxin A (Trx) at a 14.4 µg/day dosage for 12 weeks by intraventricular infusion through Alzet® osmotic pumps. We observed that treatment with PA8 improves learning and memory functions of 5XFAD mice as compared to Trx-treated 5XFAD mice. We found that PA8 treatment significantly reduces AßO levels and Aß plaques in the brain tissue of 5XFAD mice. Interestingly, PA8 significantly reduces AßO-PrP interaction and its downstream signaling such as phosphorylation of Fyn kinase, reactive gliosis as well as apoptotic neurodegeneration in the 5XFAD mice compared to Trx-treated 5XFAD mice. Collectively, our results demonstrate that treatment with PA8 targeting the AßO-PrP-Fyn axis is a promising and novel approach to prevent and treat AD.
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Doença de Alzheimer , Aptâmeros de Peptídeos , Proteínas PrPC , Camundongos , Animais , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Camundongos Transgênicos , Peptídeos beta-Amiloides/metabolismo , Proteínas PrPC/metabolismo , Modelos Animais de DoençasRESUMO
In developing and developed countries, an increasing elderly population is observed. This affects the growing percentage of people struggling with neurodegenerative diseases, including Alzheimer's disease. Nevertheless, the pathomechanism of this disease is still unknown. This contributes to problems with early diagnosis of the disease as well as with treatment. One of the most popular hypotheses of Alzheimer's disease is related to the pathological deposition of amyloid-ß (Aß) in the brain of ill people. In this paper, we discuss issues related to Aß and its relationship in the development of Alzheimer's disease. The structure of Aß and its interaction with the cell membrane are discussed. Not only do the extracellular plaques affect nerve cells, but other forms of this peptide as well.
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Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Proteínas de Membrana/metabolismo , Doença de Alzheimer/metabolismo , Animais , HumanosRESUMO
Animal models have been instrumental in reproducing key aspects of human tauopathy. In pursuing these efforts, the mouse continues to have a prominent role. In this chapter, we focus on models that overexpress wild-type or mutant forms of tau, the latter being based on mutations found in familial cases of frontotemporal dementia. We review some of these models in more detail and discuss what they have revealed about the underlying pathomechanisms, as well as highlighting new developments that exploit gene editing tools such as TALEN and CRISPR. Interestingly, when investigating the role of tau in impairing cellular functions, common themes emerge. Because tau is a scaffolding protein that aggregates in the somatodendritic domain under pathological conditions, it traps proteins such as parkin and JIP1, preventing them from executing their normal function in mitophagy and axonal transport, respectively. Another aspect is the emerging role of tau in the translational machinery and the finding that the somatodendritic accumulation of tau in Alzheimer's disease may in part be due to the induction of the de novo synthesis of tau by amyloid-ß via the Fyn/ERK/S6 pathway. We further discuss treatment strategies such as tau-based vaccinations and therapeutic ultrasound and conclude by discussing whether there is a future for animal models of tauopathies.
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Modelos Animais de Doenças , Tauopatias/metabolismo , Tauopatias/terapia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Humanos , Tauopatias/genética , Tauopatias/patologia , Ultrassonografia de Intervenção , Vacinas/uso terapêutico , Proteínas tau/genética , Proteínas tau/metabolismoRESUMO
Fyn tyrosine kinase inhibitors are considered potential therapeutic agents for a variety of human cancers. Furthermore, the involvement of Fyn kinase in signalling pathways that lead to severe pathologies, such as Alzheimer's and Parkinson's diseases, has also been demonstrated. In this study, starting from 3-(benzo[d][1,3]dioxol-5-ylamino)-6-methyl-1,2,4-triazin-5(2H)-one (VS6), a hit compound that showed a micromolar inhibition of Fyn (IC50 = 4.8 µM), we computationally investigated the binding interactions of the 3-amino-1,2,4-triazin-5(2H)-one scaffold and started a preliminary hit to lead optimisation. This analysis led us to confirm the hypothesised binding mode of VS6 and to identify a new derivative that is about 6-fold more active than VS6 (compound 3, IC50 = 0.76 µM).
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Antineoplásicos/farmacologia , Citosina/análogos & derivados , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-fyn/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Sítios de Ligação/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citosina/síntese química , Citosina/química , Citosina/farmacologia , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Modelos Moleculares , Estrutura Molecular , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Relação Estrutura-AtividadeRESUMO
BACKGROUND: Mitochondrial translation machinery solely exists for the synthesis of 13 mitochondrially-encoded subunits of the oxidative phosphorylation (OXPHOS) complexes in mammals. Therefore, it plays a critical role in mitochondrial energy production. However, regulation of the mitochondrial translation machinery is still poorly understood. In comprehensive proteomics studies with normal and diseased tissues and cell lines, we and others have found the majority of mitochondrial ribosomal proteins (MRPs) to be phosphorylated. Neither the kinases for these phosphorylation events nor their specific roles in mitochondrial translation are known. METHODS: Mitochondrial kinases are responsible for phosphorylation of MRPs enriched from bovine mitoplasts by strong cation-exchange chromatography and identified by mass spectrometry-based proteomics analyses of kinase rich fractions. Phosphorylation of recombinant MRPs and 55S ribosomes was assessed by in vitro phosphorylation assays using the kinase-rich fractions. The effect of identified kinase on OXPHOS and mitochondrial translation was assessed by various cell biological and immunoblotting approaches. RESULTS: Here, we provide the first evidence for the association of Fyn kinase, a Src family kinase, with mitochondrial translation components and its involvement in phosphorylation of 55S ribosomal proteins in vitro. Modulation of Fyn expression in human cell lines has provided a link between mitochondrial translation and energy metabolism, which was evident by the changes in 13 mitochondrially encoded subunits of OXPHOS complexes. CONCLUSIONS AND GENERAL SIGNIFICANCE: Our findings suggest that Fyn kinase is part of a complex mechanism that regulates protein synthesis and OXPHOS possibly by tyrosine phosphorylation of translation components in mammalian mitochondria.
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Mamíferos/metabolismo , Mamíferos/fisiologia , Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , Proteínas Mitocondriais/metabolismo , Biossíntese de Proteínas/fisiologia , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Animais , Bovinos , Linhagem Celular , Linhagem Celular Tumoral , Células HEK293 , Células Hep G2 , Humanos , Immunoblotting/métodos , Ribossomos Mitocondriais/metabolismo , Ribossomos Mitocondriais/fisiologia , Fosforilação Oxidativa , Fosforilação/fisiologia , Proteômica/métodos , Proteínas Ribossômicas/metabolismoRESUMO
Fyn kinase is widely expressed in neuronal and glial cells of the brain, where it exerts multiple functional roles that affect fundamental physiological processes. The aim of our study was to investigate the, so far unknown, functional role of Fyn in the retina. We report that Fyn is expressed, in vivo, in a subpopulation of Müller glia. We used a mouse model of Fyn genetic ablation and Müller-enriched primary cultures to demonstrate that Fyn deficiency induces morphological alterations in the mature retina, a reduction in the thickness of the outer and inner nuclear layers and alterations in postnatal Müller cell physiology. These include shortening of Müller cell processes, a decrease in cell proliferation, inactivation of the Akt signal transduction pathway, a reduced number of focal adhesions points and decreased adhesion of these cells to the ECM. As abnormalities in Müller cell physiology have been previously associated to a compromised retinal function we evaluated behavioral responses to visual stimulation. Our results associate Fyn deficiency with impaired visual optokinetic responses under scotopic and photopic light conditions. Our study reveals novel roles for Fyn kinase in retinal morphology and Müller cell physiology and suggests that Fyn is required for optimal visual processing.
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Células Ependimogliais/metabolismo , Proteínas Proto-Oncogênicas c-fyn/genética , Animais , Adesão Celular , Proliferação de Células , Células Cultivadas , Células Ependimogliais/citologia , Células Ependimogliais/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Transdução de Sinais , Visão OcularRESUMO
Diabetic neuropathy is a common complication of diabetes. This study evaluated the role of Fyn kinase and N-methyl-d-aspartate receptors (NMDARs) in the spinal cord in diabetic neuropathy using an animal model of high-fat diet-induced prediabetes. We found that prediabetic wild-type mice exhibited tactile allodynia and thermal hypoalgesia after a 16-wk high-fat diet, relative to normal diet-fed wild-type mice. Furthermore, prediabetic wild-type mice exhibited increased tactile allodynia and thermal hypoalgesia at 24 wk relative to 16 wk. Such phenomena were correlated with increased expression and activation of NR2B subunit of NMDARs, as well as Fyn-NR2B interaction in the spinal cord. Fyn(-/-) mice developed prediabetes after 16-wk high-fat diet treatment and exhibited thermal hypoalgesia, without showing tactile allodynia or altered expression and activation of NR2B subunit, relative to normal diet-fed Fyn(-/-) mice. Finally, intrathecal administrations of Ro 25-6981 (selective NR2B subunit-containing NMDAR antagonist) dose-dependently alleviated tactile allodynia, but not thermal hypoalgesia, at 16 and 24 wk in prediabetic wild-type mice. Our results suggested that Fyn-mediated NR2B signaling plays a critical role in regulation of prediabetic neuropathy and that the increased expression/function of NR2B subunit-containing NMDARs may contribute to the progression of neuropathy in type 2 diabetes.
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Neuropatias Diabéticas/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Estado Pré-Diabético/fisiopatologia , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Pressão Sanguínea/fisiologia , Peso Corporal/fisiologia , Neuropatias Diabéticas/induzido quimicamente , Neuropatias Diabéticas/patologia , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Ingestão de Alimentos/efeitos dos fármacos , Ingestão de Alimentos/genética , Antagonistas de Aminoácidos Excitatórios/farmacologia , Hiperalgesia/etiologia , Hiperalgesia/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Limiar da Dor/efeitos dos fármacos , Limiar da Dor/fisiologia , Fenóis/farmacologia , Piperidinas/farmacologia , Estado Pré-Diabético/induzido quimicamente , Proteínas Proto-Oncogênicas c-fyn/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Medula Espinal/efeitos dos fármacos , Medula Espinal/metabolismo , Medula Espinal/patologiaRESUMO
Oligodendrocytes, the myelin forming cells of the CNS, are characterized by their numerous membranous extensions, which enwrap neuronal axons and form myelin sheaths. During differentiation oligodendrocytes pass different morphological stages, downregulate the expression of the proteoglycan NG2, and acquire major myelin specific proteins, such as myelin basic proteins (MBP) and proteolipid protein. MBP mRNA is transported in RNA granules along the microtubules (MTs) to the periphery and translated locally. MTs participate in the elaboration and stabilization of the myelin forming extensions and are essential for cellular sorting processes. Their dynamic properties are regulated by microtubule associated proteins (MAPs). The MAP tau is present in oligodendrocytes and involved in the regulation and stabilization of the MT network. To further elucidate the functional significance of tau in oligodendrocytes, we have downregulated tau by siRNA technology and studied the effects on cell differentiation and neuron-glia contact formation. The data show that tau knockdown impairs process outgrowth and leads to a decrease in MBP expression. Furthermore, MBP mRNA transport to distant cellular extensions is impaired and cells remain in the NG2 stage. In myelinating cocultures with dorsal root ganglion neurons, oligodendrocyte precursor cells after tau miR RNA lentiviral knockdown develop into NG2 positive cells with very long and thin processes, contacting axons loosely, but fail to form internodes. This demonstrates that tau is important for MBP mRNA transport and involved in process formation. The disturbance of the balance of tau leads to abnormalities in oligodendrocyte differentiation, neuron-glia contact formation and the early myelination process.
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Proteína Básica da Mielina/metabolismo , Oligodendroglia/metabolismo , Transporte de RNA/fisiologia , RNA Mensageiro/metabolismo , Proteínas tau/metabolismo , Animais , Encéfalo/metabolismo , Linhagem Celular , Células Cultivadas , Técnicas de Cocultura , Regulação para Baixo , Gânglios Espinais/metabolismo , Humanos , Microtúbulos/metabolismo , Bainha de Mielina/metabolismo , Neurônios/metabolismo , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Ratos Wistar , Tubulina (Proteína)/metabolismo , Proteínas tau/genéticaRESUMO
Mast cells, constituents of virtually all organs and tissues, are critical cells in IgE-mediated allergic responses. The aim of this study was to investigate the effect and mechanism of an indoxyl chromogenic compound, 5-bromo-4-chloro-3-indolyl 1,3-diacetate, CAC-0982, on IgE-mediated mast cell activation and allergic responses in mice. CAC-0982 reversibly suppressed antigen-stimulated degranulation in murine mast cells (IC50, ~3.8µM) and human mast cells (IC50, ~3.0µM). CAC-0982 also inhibited the expression and secretion of IL-4 and TNF-α in mast cells. Furthermore, CAC-0982 suppressed the mast cell-mediated allergic responses in mice in a dose-dependent manner (ED50 27.9mg/kg). As for the mechanism, CAC-0982 largely suppressed the phosphorylation of Syk and its downstream signaling molecules, including LAT, Akt, Erk1/2, p38, and JNK. Notably, the tyrosine kinase assay of antigen-stimulated mast cells showed that CAC-0982 inhibited Fyn kinase, one of the upstream tyrosine kinases for Syk activation in mast cells. Taken together, these results suggest that CAC-0982 may be used as a new treatment for regulating IgE-mediated allergic diseases through the inhibition of the Fyn/Syk pathway in mast cells.
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Imunoglobulina E/imunologia , Indóis/farmacologia , Mastócitos/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Animais , Humanos , Hipersensibilidade/tratamento farmacológico , Hipersensibilidade/imunologia , Interleucina-4/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Camundongos , Fosforilação , Proteínas Tirosina Quinases/metabolismo , Transdução de Sinais , Quinase Syk , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Voltage-gated Na(+) channels (VGSCs) are heteromeric proteins composed of pore-forming α subunits and smaller ß subunits. The ß subunits are multifunctional channel modulators and are members of the immunoglobulin superfamily of cell adhesion molecules (CAMs). ß1, encoded by SCN1B, is best characterized in the central nervous system (CNS), where it plays a critical role in regulating electrical excitability, neurite outgrowth and migration during development. ß1 is also expressed in breast cancer (BCa) cell lines, where it regulates adhesion and migration in vitro. In the present study, we found that SCN1B mRNA/ß1 protein were up-regulated in BCa specimens, compared with normal breast tissue. ß1 upregulation substantially increased tumour growth and metastasis in a xenograft model of BCa. ß1 over-expression also increased vascularization and reduced apoptosis in the primary tumours, and ß1 over-expressing tumour cells had an elongate morphology. In vitro, ß1 potentiated outgrowth of processes from BCa cells co-cultured with fibroblasts, via trans-homophilic adhesion. ß1-mediated process outgrowth in BCa cells required the presence and activity of fyn kinase, and Na(+) current, thus replicating the mechanism by which ß1 regulates neurite outgrowth in CNS neurons. We conclude that when present in breast tumours, ß1 enhances pathological growth and cellular dissemination. This study is the first demonstration of a functional role for ß1 in tumour growth and metastasis in vivo. We propose that ß1 warrants further study as a potential biomarker and targeting ß1-mediated adhesion interactions may have value as a novel anti-cancer therapy.
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Neoplasias da Mama/patologia , Neoplasias Hepáticas/secundário , Neoplasias Pulmonares/secundário , Neuritos/patologia , Subunidade beta-1 do Canal de Sódio Disparado por Voltagem/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Apoptose , Western Blotting , Mama/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proliferação de Células , Feminino , Humanos , Técnicas Imunoenzimáticas , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Neuritos/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Proteínas Proto-Oncogênicas c-fyn/genética , Proteínas Proto-Oncogênicas c-fyn/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas , Subunidade beta-1 do Canal de Sódio Disparado por Voltagem/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Regulatory T (Treg) cells are indispensable in maintaining the immune homeostasis and preventing autoimmune diseases. Regulatory T (Treg) cells include thymus derived Treg cells (tTregs) and peripherally induced Treg cells (iTreg), which are differentiated from antigen stimulated CD4+ naïve T cells in presence of TGFß. tTregs are quite stable, and more immune suppressive, while iTreg cells are less stable, and are prone to differentiate into inflammatory T cells. Therefore, identification of small molecules that could promote the differentiation of iTreg cells is an attractive strategy for autoimmune diseases. Inhibition of AKT/mTOR pathway promotes their differentiation. Whether inhibition of Lck/Fyn kinase activity (upstream of AKT/mTOR pathway) can be used to promote the differentiation of iTreg cells has not been determined. Here, we showed that Srci1, a small molecular inhibitor of Lck/Fyn, promoted the differentiation of FOXP3+ iTreg cells. Srci1 treatment resulted in inhibition of phosphorylation of key components of AKT/mTOR pathway, including mTOR, p70 S6K, 4EBP1, and promoted the expression of Foxp3 and its target genes, thereby promoted differentiation of in vitro iTreg cells. Srci1 treated iTreg cells showed more similar gene expression profile to that of tTreg cells. Our results thus suggest that inhibition of Lck/Fyn kinase activity can promote the differentiation of iTreg cells, and may have implication in autoimmune diseases.
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Diferenciação Celular , Proteína Tirosina Quinase p56(lck) Linfócito-Específica , Proteínas Proto-Oncogênicas c-akt , Transdução de Sinais , Linfócitos T Reguladores , Serina-Treonina Quinases TOR , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Diferenciação Celular/efeitos dos fármacos , Animais , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Camundongos , Fatores de Transcrição Forkhead/metabolismo , Células Cultivadas , Camundongos Endogâmicos C57BL , HumanosRESUMO
Alzheimer's disease, characterized by the accumulation of abnormal protein aggregates and neuronal damage in the brain, leads to a gradual decline in cognitive function and memory. As a complex neurodegenerative disorder, it involves disruptions in various biochemical pathways and neurotransmitter systems, contributing to the progressive loss of neurons and synaptic connections. The complexity of Alzheimer's signaling pathways complicates treatment, presenting a formidable challenge in the quest for effective therapeutic interventions. A member of the Src family of kinases (SFKs), Fyn, is a type of non-receptor tyrosine kinase that has been linked to multiple essential CNS processes, such as myelination and synaptic transmission. Fyn is an appealing target for AD treatments because it is uniquely linked to the two major pathologies in AD by its interaction with tau, in addition to being activated by amyloid-beta (Aß) through PrPC. Fyn mediates neurotoxicity and synaptic impairments caused by Aß and is involved in regulating the process of Aß synthesis.Additionally, the tau protein's tyrosine phosphorylation is induced by Fyn. Fyn is also a challenging target because of its widespread body expression and strong homology with other kinases of the Src family, which could cause unintentional off-target effects. This review emphasizes signaling pathways mediated by Fyn that govern neuronal development and plasticity while also summarizing the most noteworthy recent research relevant to Fyn kinase's function in the brain. Additionally, the therapeutic inhibition of Fyn kinase has been discussed, with a focus on the Fyn kinase inhibitors that are in clinical trials, which presents a fascinating opportunity for targeting Fyn kinase in the creation of possible therapeutic approaches for the management of Alzheimer's disease.
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INTRODUCTION: Fyn kinase is an Src family kinase (SFK) widely expressed in many tissues, including the CNS. Recently, Fyn kinase activation has been associated with pathological mechanisms underlying neurodegenerative diseases and, as such, the role of Fyn dysfunction is under investigation. In particular, Fyn is implicated as a major upstream regulator of neuroinflammation in Parkinson's Disease (PD). Chronic neuroinflammation has been observed not just in the substantia nigra (SN), but also in several key regions of the brain, with disruption associated with symptoms presentation in PD. This study aimed to characterise the anatomical distribution of Fyn in key brain regions affected in PD, namely the prefrontal cortex, hippocampus, striatum and SN. METHODS: Fresh and fixed post-mortem PD brain samples (n = 10) were collected and compared with neurologically healthy age-matched controls (n = 7) to assess markers of Fyn activity and neuroinflammation. RESULTS: Increased Fyn phosphorylation was observed in SN and striatum of post-mortem samples from PD patients compared with controls. No such increase was observed in the prefrontal cortex or hippocampus. In contrast with previous findings, no increase in microglial activation or astrocyte reactivity was observed in PD brains across regions. CONCLUSION: Taken together, these results indicate that Fyn dysfunction may be involved in the pathological processes observed in PD; however, this appears to be independent of inflammatory mechanisms. Further investigations are required to elucidate if increased Fyn activity is a potential cause or consequence of pathological processing in PD.
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Doença de Parkinson , Humanos , Doença de Parkinson/complicações , Doenças Neuroinflamatórias , Encéfalo/patologia , Substância Negra/patologia , FosforilaçãoRESUMO
Fyn kinase SH3 domain interaction with PXXP motif in the Tau protein is implicated in AD pathology and is central to NMDAR function. Among seven PXXP motifs localized in proline-rich domain of Tau protein, tandem 5th and 6th PXXP motifs are critical to Fyn-SH3 domain interaction. Here, we report the crystal structure of Fyn-SH3 -Tau (207-221) peptide consisting of 5th and 6th PXXP motif complex to 1.01 Å resolution. Among five AD-specific phosphorylation sites encompassing the 5th and 6th PXXP motifs, only S214 residue showed interaction with SH3 domain. Biophysical studies showed that Tau (207-221) with S214-phosphorylation (pS214) inhibits its interaction with Fyn-SH3 domain. The individual administration of Tau (207-221) with/without pS214 peptides to a single neuron increased the decay time of evoked NMDA current response. Recordings of spontaneous NMDA EPSCs at +40 mV indicate an increase in frequency and amplitude of events for the Tau (207-221) peptide. Conversely, the Tau (207-221) with pS214 peptide exhibited a noteworthy amplitude increase alongside a prolonged decay time. These outcomes underscore the distinctive modalities of action associated with each peptide in the study. Overall, this study provides insights into how Tau (207-221) with/without pS214 affects the molecular framework of NMDAR signaling, indicating its involvement in Tau-related pathogenesis.
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Domínios Proteicos Ricos em Prolina , Proteínas Proto-Oncogênicas c-fyn , Receptores de N-Metil-D-Aspartato , Domínios de Homologia de src , Proteínas tau , N-Metilaspartato/química , Peptídeos/química , Fosforilação , Ligação Proteica , Proteínas Proto-Oncogênicas c-fyn/química , Proteínas Proto-Oncogênicas c-fyn/genética , Proteínas tau/química , Proteínas tau/genética , Humanos , Receptores de N-Metil-D-Aspartato/química , Estabilidade ProteicaRESUMO
Alzheimer's disease is characterized by progressive impairment of neuronal functions culminating in neuronal loss and dementia. A universal feature of dementia is protein aggregation, a process by which a monomer forms intermediate oligomeric assembly states and filaments that develop into end-stage hallmark lesions. In Alzheimer's disease, this is exemplified by extracellular amyloid-ß (Aß) plaques which have been placed upstream of tau, found in intracellular neurofibrillary tangles and dystrophic neurites. This implies causality that can be modeled as a linear activation cascade. When Aß load is reduced, for example, in response to an anti-Aß immunotherapy, cognitive functions improve in plaque-forming mice. They also deteriorate less in clinical trial cohorts although real-world clinical benefits remain to be demonstrated. Given the existence of aged humans with unimpaired cognition despite a high plaque load, the central role of Aß has been challenged. A counter argument has been that clinical symptoms would eventually develop if these aged individuals were to live long enough. Alternatively, intrinsic mechanisms that protect the brain in the presence of pathology may exist. In fact, Aß toxicity can be abolished by either reducing or manipulating tau (through which Aß signals), at least in preclinical models. In addition to manipulating steps in this linear pathocascade model, mechanisms of restoring brain reserve can also counteract Aß toxicity. Low-intensity ultrasound is a neuromodulatory modality that can improve cognitive functions in Aß-depositing mice without the need for removing Aß. Together, this highlights a dissociation of Aß and cognition, with important implications for therapeutic interventions.
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Doença de Alzheimer , Proteínas tau , Animais , Humanos , Proteínas tau/metabolismo , Doença de Alzheimer/terapia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Cognição/fisiologia , Peptídeos beta-Amiloides/metabolismo , Placa Amiloide/patologia , Terapia por Ultrassom/métodosRESUMO
STUDY QUESTION: Does the position of the germinal vesicle (GV) in human oocytes correlate with molecular and morphological parameters as well as with maturation-competence? SUMMARY ANSWER: The position of GV in human oocytes correlates with density of microtubule (MT) filaments, concentration of Fyn, nucleolus localization and the ability of the oocytes to complete maturation following GV breakdown (GVBD). WHAT IS KNOWN ALREADY: Our knowledge is confined to oocytes of young mice where maturation-competence is correlated with a central GV and regulated by MTs and the presence of a chromatin ring. Fyn kinase is localized at the spindle and cortex of mouse oocytes and plays a role in both maturation and MT stabilization. STUDY DESIGN, SIZE, DURATION: Spatial localization of the GV and nucleolus (central or peripheral), the presence of a chromatin ring, the localization of Fyn, MT density and oocyte maturation were assessed in 153 human oocytes, 335 oocytes from young mice (2-month-old) and 146 oocytes from old mice (12-month-old). PARTICIPANTS/MATERIALS, SETTING, METHODS: GV human oocytes were donated by consenting female patients (n = 57), 21-45-year-old undergoing IVF/ICSI. As a control, GV mouse oocytes were collected from female mice after injection of pregnant mares' serum gonadotrophin. Human and mouse GV oocytes allocated for immunocytochemistry were fixed on day of retrieval, stained with specific antibodies and imaged using a confocal laser-scanning microscope. Human and mouse oocytes allocated for maturation were incubated for 48 and 24 h, respectively. GVBD and extrusion of the first polar body (PBI) were assessed using differential interference contrast optics. MAIN RESULTS AND THE ROLE OF CHANCE: GV location was peripheral and independent of age in 69.9% of the human oocytes, but GV location did vary with age in mice oocytes; it was central in 89.9% of the oocytes retrieved from young-mice and peripheral in 52.1% of the oocytes retrieved from old mice (P < 0.05). A central GV, whether in human or mouse oocytes, was highly correlated with a central nucleolus, absence of Fyn at the GV and a dense MT network (P < 0.05), whereas a peripheral GV correlated with peripheral nucleolus, presence of Fyn at the GV and a flimsy MT network. After 48 h in culture, no degeneration was observed in human central-GV oocytes, however, 12/95 (12.6%) of the peripheral-GV oocytes degenerated (P < 0.05). No correlation was observed between GV position and presence of a chromatin ring. The percentage of human oocytes that extruded the PBI after completing GVBD was significantly higher (73.7%) in central than in peripheral-GV oocytes (45.8%; P < 0.05). In mice oocytes, central location of the GV correlated with maturation competence in young (P < 0.05) but not old mice. LIMITATIONS, REASONS FOR CAUTION: The fact that the human GV oocytes used in this study were exposed to gonadotrophic stimulation but failed to mature in vivo might be a sign of their low quality and this should be considered when drawing conclusions from the data. Furthermore, our observation that only peripheral-GV human oocytes were degraded may indicate that they are of a lower quality than central-GV human oocytes. WIDER IMPLICATIONS OF THE FINDINGS: We suggest that the central location of GV within the oocytes, which is associated with an absence of Fyn at the GV and the presence of thick filamentous MTs in the ooplasm, may serve as a predictor of successful maturation and provide new insights for the use of IVM.
Assuntos
Envelhecimento , Oócitos/citologia , Oogênese , Adulto , Animais , Biomarcadores/metabolismo , Nucléolo Celular/metabolismo , Cromatina/metabolismo , Feminino , Fármacos para a Fertilidade Feminina/farmacologia , Humanos , Técnicas de Maturação in Vitro de Oócitos , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Confocal , Microtúbulos/metabolismo , Pessoa de Meia-Idade , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Indução da Ovulação , Corpos Polares/citologia , Corpos Polares/metabolismo , Transporte Proteico , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Especificidade da Espécie , Adulto JovemRESUMO
Parkinson's disease (PD) is a progressive neurodegenerative disorder characterized by the selective degeneration of dopaminergic neurons in the substantia nigra pars compacta resulting in an irreversible and a debilitating motor dysfunction. Though both genetic and idiopathic factors are implicated in the disease etiology, idiopathic PD comprise the majority of clinical cases and is caused by exposure to environmental toxicants and oxidative stress. Fyn kinase activation has been identified as an early molecular signaling event that primes neuroinflammatory and neurodegenerative events associated with dopaminergic cell death. However, the upstream regulator of Fyn activation remains unidentified. We investigated whether the lipid and tyrosine phosphatase PTEN (Phosphatase and Tensin homolog deleted on chromosome 10) could be the upstream regulator of Fyn activation in PD models as PTEN has been previously reported to contribute to Parkinsonian pathology. Our findings, using bioluminescence resonance energy transfer (BRET) and immunoblotting, indicate for the first time that PTEN is a critical early stress sensor in response to oxidative stress and neurotoxicants in in vitro models of PD. Pharmacological attenuation of PTEN activity rescues dopaminergic neurons from neurotoxicant-induced cytotoxicity by modulating Fyn kinase activation. Our findings also identify PTEN's novel roles in contributing to mitochondrial dysfunction which contribute to neurodegenerative processes. Interestingly, we found that PTEN positively regulates interleukin-1ß (IL-1ß) and the transcription of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). Taken together, we have identified PTEN as a disease course altering pharmacological target that may be further validated for the development of novel therapeutic strategies targeting PD.
Assuntos
Neurônios Dopaminérgicos , PTEN Fosfo-Hidrolase , Doença de Parkinson , Humanos , Neurônios Dopaminérgicos/metabolismo , Estresse Oxidativo , Doença de Parkinson/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Transdução de Sinais/fisiologia , Animais , RatosRESUMO
Aims: Huntington's disease (HD) is an autosomal-dominant neurodegenerative disorder with no effective therapies. Mutant huntingtin protein (mHTT), the main HD proteinaceous hallmark, has been linked to reactive oxygen species (ROS) formation and mitochondrial dysfunction, among other pathological mechanisms. Importantly, Src-related kinases, c-Src and Fyn, are activated by ROS and regulate mitochondrial activity. However, c-Src/Fyn involvement in HD is largely unexplored. Thus, in this study, we aimed at exploring changes in Src/Fyn proteins in HD models and their role in defining altered mitochondrial function and dynamics and redox regulation. Results: We show, for the first time, that c-Src/Fyn phosphorylation/activation and proteins levels are decreased in several human and mouse HD models mainly due to autophagy degradation, concomitantly with mHtt-expressing cells showing enhanced TFEB-mediated autophagy induction and autophagy flux. c-Src/Fyn co-localization with mitochondria is also reduced. Importantly, the expression of constitutive active c-Src/Fyn to restore active Src kinase family (SKF) levels improves mitochondrial morphology and function, namely through improved mitochondrial transmembrane potential, mitochondrial basal respiration, and ATP production, but it did not affect mitophagy. In addition, constitutive active c-Src/Fyn expression diminishes the levels of reactive species in cells expressing mHTT. Innovation: This work supports a relevant role for c-Src/Fyn proteins in controlling mitochondrial function and redox regulation in HD, revealing a potential HD therapeutic target. Conclusion: c-Src/Fyn restoration in HD improves mitochondrial morphology and function, precluding the rise in oxidant species and cell death. Antioxid. Redox Signal. 38, 95-114.