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1.
J Cancer Res Clin Oncol ; 145(9): 2293-2301, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31401673

RESUMO

PURPOSE: Androgen receptors (ARs) are expressed on a variety of cell types, and AR signaling plays an important role in tumor development and progression in several cancers. This in vitro study evaluated the effect of dihydrotestosterone (DHT) on the proliferation of renal cell carcinoma (RCC) cells in relation to AR status. METHODS: Steroid hormone receptor expression was evaluated using RT-PCR and Western blotting. The effect of DHT on cell proliferation and STAT5 phosphorylation was evaluated in RCC cell lines (Caki-2, A498, and SN12C) and primary RCC cells using cell viability assays and Western blotting. ARs and glucocorticoid receptors (GRs) were knocked down with small interfering RNAs before assessing changes in cell proliferation and STAT5 activation. RESULTS: DHT treatment promoted cell proliferation and increased STAT5 phosphorylation regardless of AR status. The AR antagonist bicalutamide reduced kidney cancer cell proliferation, regardless of AR status. AR and GR knockdown blocked STAT5 activation and reduced cell proliferation in all RCC cell lines. In patient-derived primary cells, DHT enhanced cell proliferation and this effect was diminished by treatment with the AR antagonists bicalutamide and enzalutamide and the GR antagonist mifepristone. CONCLUSION: DHT promotes cell proliferation through STAT5 activation in RCC cells, regardless of AR status. DHT appears to utilize the AR and GR pathways to activate STAT5, and the inhibition of AR and GR showed antitumor activity in RCC cells. These data suggest that targeting AR and GR may be a promising new approach to the treatment of RCC.


Assuntos
Carcinoma de Células Renais/patologia , Proliferação de Células/efeitos dos fármacos , Di-Hidrotestosterona/farmacologia , Neoplasias Renais/patologia , Receptores Androgênicos/fisiologia , Receptores de Glucocorticoides/fisiologia , Fator de Transcrição STAT5/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Antagonistas de Receptores de Andrógenos/farmacologia , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Fator de Transcrição STAT5/genética , Transdução de Sinais/efeitos dos fármacos , Proteínas Supressoras de Tumor/genética
2.
Medicine (Baltimore) ; 98(33): e16807, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31415393

RESUMO

BACKGROUND: Sepsis is a serious clinical condition with a poor prognosis, despite improvements in diagnosis and treatment.Therefore, novel biomarkers are necessary that can help with estimating prognosis and improving clinical outcomes of patients with sepsis. METHODS: The gene expression profiles GSE54514 and GSE63042 were downloaded from the GEO database. DEGs were screened by t test after logarithmization of raw data; then, the common DEGs between the 2 gene expression profiles were identified by up-regulation and down-regulation intersection. The DEGs were analyzed using bioinformatics, and a protein-protein interaction (PPI) survival network was constructed using STRING. Survival curves were constructed to explore the relationship between core genes and the prognosis of sepsis patients based on GSE54514 data. RESULTS: A total of 688 common DEGs were identified between survivors and non-survivors of sepsis, and 96 genes were involved in survival networks. The crucial genes Signal transducer and activator of transcription 5A (STAT5A), CCAAT/enhancer-binding protein beta (CEBPB), Myc proto-oncogene protein (MYC), and REL-associated protein (RELA) were identified and showed increased expression in sepsis survivors. These crucial genes had a positive correlation with patients' survival time according to the survival analysis. CONCLUSIONS: Our findings indicate that the genes STAT5A, CEBPB, MYC, and RELA may be important in predicting the prognosis of sepsis patients.


Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fator de Transcrição STAT5/metabolismo , Sepse/genética , Sepse/mortalidade , Fator de Transcrição RelA/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Idoso , Idoso de 80 Anos ou mais , Biologia Computacional , Bases de Dados Genéticas , Regulação para Baixo , Feminino , Marcadores Genéticos , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Mapas de Interação de Proteínas , Fatores de Tempo , Transcriptoma , Regulação para Cima
3.
BMC Bioinformatics ; 20(1): 395, 2019 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-31311516

RESUMO

BACKGROUND: Ordinary differential equation systems are frequently utilized to model biological systems and to infer knowledge about underlying properties. For instance, the development of drugs requires the knowledge to which extent malign cells differ from healthy ones to provide a specific treatment with least side effects. As these cell-type specific properties may stem from any part of biochemical cell processes, systematic quantitative approaches are necessary to identify the relevant potential drug targets. An ℓ1 regularization for the maximum likelihood parameter estimation proved to be successful, but falsely predicted cell-type dependent behaviour had to be corrected manually by using a Profile Likelihood approach. RESULTS: The choice of extended ℓ1 penalty functions significantly decreased the number of falsely detected cell-type specific parameters. Thus, the total accuracy of the prediction could be increased. This was tested on a realistic dynamical benchmark model used for the DREAM6 challenge. Among Elastic Net, Adaptive Lasso and a non-convex ℓq penalty, the latter one showed the best predictions whilst also requiring least computation time. All extended methods include a hyper-parameter in the regularization function. For an Erythropoietin (EPO) induced signalling pathway, the extended methods ℓq and Adaptive Lasso revealed an unpublished alternative parsimonious model when varying the respective hyper-parameters. CONCLUSIONS: Using ℓq or Adaptive Lasso with an a-priori choice for the hyper-parameter can lead to a more specific and accurate result than ℓ1. Scanning different hyper-parameters can yield additional pieces of information about the system.


Assuntos
Modelos Biológicos , Eritropoetina/metabolismo , Humanos , Janus Quinase 2/metabolismo , Funções Verossimilhança , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais , Biologia de Sistemas/métodos
4.
J Trace Elem Med Biol ; 54: 214-220, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31109615

RESUMO

Zinc has a strong influence on the function of the immune system and is a driving factor for immune cell development. In this regard, studies revealed cell type specific effects of zinc. During zinc deficiency for example, development and activity of myeloid cells seems to be prioritized at the cost of cells from the lymphoid lineage. In T-cells, the altered proliferation was found to be due to zinc's effect on IL-2-induced signaling processes, but in contrast to lymphoid cells, effects of zinc homeostasis on growth-factor-induced signaling in myeloid cells have not been investigated yet. The granulocyte-macrophage colony-stimulating factor (GM-CSF) is one of the major factors inducing monopoiesis. Considering the structural similarities between the GM-CSF receptor and those of the IL-receptor family as well as a similar set of signaling molecules involved, an impact of zinc on the GM-CSF signaling seems to be likely. Therefore, the effect of zinc on GM-CSF-induced signaling molecules was investigated here, using U937 cells as a model myeloid cell line. GM-CSF stimulation significantly increased STAT5 phosphorylation which was prevented completely by pre-incubation with zinc and pyrithione. U937 cells showed a strong pre-activation regarding c-Raf, which was significantly decreased by zinc and pyrithione incubation, independently from GM-CSF stimulation. As current literature was not sufficient to explain the observed effects, we hypothesized an altered receptor-complex assembly. As membrane composition and plasticity, subsumed under the term of membrane fluidity, was found to affect receptor multimerization, the impact of zinc on membrane fluidity was considered as a completely novel approach. Indeed, addition of zinc also decreased GM-CSFR expression on the cell surface and most interestingly altered membrane fluidity. In conclusion, we hypothesize that the incubation with zinc causes an alteration of membrane fluidity that hinders efficient receptor assembly as well as phosphorylation of signal molecules and therefore signal transduction.


Assuntos
Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Fluidez de Membrana/fisiologia , Células Mieloides/efeitos dos fármacos , Linhagem Celular , Humanos , Células Mieloides/metabolismo , Fosforilação/efeitos dos fármacos , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais/efeitos dos fármacos , Zinco/metabolismo
5.
BMC Bioinformatics ; 20(1): 242, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31092187

RESUMO

BACKGROUND: ErbB4/HER4 is a unique member of the ErbB family of receptor tyrosine kinases concerning its activation of anti-proliferative JAK2-STAT5 pathway when stimulated by ligand Neuregulin (NRG). Activation of this pathway leads to expression of genes like ß-casein which promote cell differentiation. Recent experimental studies on mouse HC11 mammary epithelial cells stimulated by ligand Neuregulin (NRG) showed a time-dependent switching behavior in the ß-casein expression. This behavior cannot be explained using currently available mechanistic models of the JAK-STAT pathway. We constructed an improved mechanistic model which introduces two crucial modifications to the canonical HER4-JAK2-STAT5 pathway based on literature findings. These modifications include competitive HER4 heterodimerization with other members of the ErbB family and a slower JAK2 independent activation STAT5 through HER4. We also performed global sensitivity analysis on the model to test the robustness of the predictions and parameter combinations that are sensitive to the outcome. RESULTS: Our model was able to reproduce the time-dependent switching behavior of ß-casein and also establish that the modifications mentioned above to the canonical JAK-STAT pathway are necessary to reproduce this behavior. The sensitivity studies show that the competitive HER4 heterodimerization reactions have a profound impact on the sensitivity of the pathway to NRG stimulation, while the slower JAK2-independent pathway is necessary for the late stage promotion of ß-casein mRNA transcription. The difference in the time scales of the JAK-dependent and JAK-independent pathways was found to be the main contributing factor to the time-dependent switch. The transport rates controlling activated STAT5 dimer nuclear import and ß-casein mRNA export to cytoplasm affected the time delay between NRG stimulation and peak ß-casein mRNA activity. CONCLUSION: This study highlights the effect of competitive and parallel reaction pathways on both short and long-term dynamics of receptor-mediated signaling. It provides robust and testable predictions of the dynamical behavior of the HER4 mediated JAK-STAT pathway which could be useful in designing treatments for various cancers where this pathway is activated/altered.


Assuntos
Receptores Proteína Tirosina Quinases/agonistas , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Transdução de Sinais , Animais , Caseínas/metabolismo , Diferenciação Celular , Linhagem Celular , Núcleo Celular/metabolismo , Células Epiteliais/metabolismo , Janus Quinase 2/metabolismo , Ligantes , Camundongos , Modelos Biológicos , Multimerização Proteica , Transporte Proteico , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor ErbB-4/metabolismo , Fator de Transcrição STAT5/metabolismo , Fatores de Tempo , Transcrição Genética
6.
Nat Commun ; 10(1): 2042, 2019 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-31053703

RESUMO

Metabolic pathways that regulate T-cell function show promise as therapeutic targets in diverse diseases. Here, we show that at rest cultured human effector memory and central memory CD4+ T-cells have elevated levels of glycolysis and oxidative phosphorylation (OXPHOS), in comparison to naïve T-cells. Despite having low resting metabolic rates, naive T-cells respond to TCR stimulation with robust and rapid increases in glycolysis and OXPHOS. This early metabolic switch requires Akt activity to support increased rates of glycolysis and STAT5 activity for amino acid biosynthesis and TCA cycle anaplerosis. Importantly, both STAT5 inhibition and disruption of TCA cycle anaplerosis are associated with reduced IL-2 production, demonstrating the functional importance of this early metabolic program. Our results define STAT5 as a key node in modulating the early metabolic program following activation in naive CD4+ T-cells and in turn provide greater understanding of how cellular metabolism shapes T-cell responses.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Fator de Transcrição STAT5/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas , Ciclo do Ácido Cítrico/imunologia , Glicólise/imunologia , Voluntários Saudáveis , Humanos , Memória Imunológica , Ativação Linfocitária , Fosforilação Oxidativa , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-akt/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Fator de Transcrição STAT5/imunologia
7.
Med Sci Monit ; 25: 2599-2608, 2019 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-30964854

RESUMO

BACKGROUND Mantle cell lymphoma (MCL) is a high-grade B-cell lymphoma with poor prognosis. Fludarabine is used alone or in combination for relapsed and advanced-stage MCL. The expression of the signal transducer and activator of transcription 5B (STAT5B) gene is associated with tumorigenesis in solid tumors, but its role in MCL remains unknown. The aims of this study were to investigate the role of STAT5B in GRANTA-519 human mantle cell lymphoma cells and drug resistance. MATERIAL AND METHODS GRANTA-519 human mantle cell lymphoma cells were cultured with and without 10 µM fludarabine dephosphorylated 9-ß-D-arabinofuranosyl-2-fluoroadenine, (2-F-araA) or 10 µM 4-hydroperoxycyclophosphamide (4-HC). The MTT assay assessed cell proliferation. Flow cytometry was used to investigate the cell cycle in MCL cells treated with the specific inhibitor of the Akt pathway, LY294002, and assessed cell cycle and cell apoptosis. Western blot was used to detect the expression levels of p-Akt/Akt and STAT5B/p-STAT5B. The gene expression profiles of lymph node (LN)-derived MCL cells were compared with peripheral blood (PB)-derived lymphocytes using bioinformatics and hierarchical cluster analysis. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) was performed to determine the expression of the marker of proliferation Ki-67 (MKI67) gene. RESULTS STAT5B was significantly upregulated in LN-derived MCL cells compared with PB lymphocytes. Increased expression of STAT5B was associated with increased MCL cell proliferation and reduced cell apoptosis and was associated with drug resistance and activation of Akt. CONCLUSIONS STAT5B promoted cell proliferation and drug resistance in human MCL cells by activating the Akt signaling pathway.


Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Linfoma de Célula do Manto/genética , Linfoma de Célula do Manto/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT5/genética , Transdução de Sinais , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Ativação Enzimática , Regulação Neoplásica da Expressão Gênica , Humanos , Linfonodos/patologia , Linfócitos/metabolismo , Linfoma de Célula do Manto/enzimologia , Poli(ADP-Ribose) Polimerases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fator de Transcrição STAT5/metabolismo
8.
Immunity ; 50(5): 1232-1248.e14, 2019 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-31027998

RESUMO

Regulatory T cells (Treg cells) are important for preventing autoimmunity and maintaining tissue homeostasis, but whether Treg cells can adopt tissue- or immune-context-specific suppressive mechanisms is unclear. Here, we found that the enzyme hydroxyprostaglandin dehydrogenase (HPGD), which catabolizes prostaglandin E2 (PGE2) into the metabolite 15-keto PGE2, was highly expressed in Treg cells, particularly those in visceral adipose tissue (VAT). Nuclear receptor peroxisome proliferator-activated receptor-γ (PPARγ)-induced HPGD expression in VAT Treg cells, and consequential Treg-cell-mediated generation of 15-keto PGE2 suppressed conventional T cell activation and proliferation. Conditional deletion of Hpgd in mouse Treg cells resulted in the accumulation of functionally impaired Treg cells specifically in VAT, causing local inflammation and systemic insulin resistance. Consistent with this mechanism, humans with type 2 diabetes showed decreased HPGD expression in Treg cells. These data indicate that HPGD-mediated suppression is a tissue- and context-dependent suppressive mechanism used by Treg cells to maintain adipose tissue homeostasis.


Assuntos
Dinoprostona/análogos & derivados , Dinoprostona/metabolismo , Hidroxiprostaglandina Desidrogenases/metabolismo , Gordura Intra-Abdominal/imunologia , Linfócitos T Reguladores/enzimologia , Linfócitos T Reguladores/imunologia , Células 3T3 , Animais , Linhagem Celular , Diabetes Mellitus Tipo 2/metabolismo , Células HEK293 , Homeostase/imunologia , Humanos , Hidroxiprostaglandina Desidrogenases/genética , Resistência à Insulina/genética , Gordura Intra-Abdominal/citologia , Células Jurkat , Ativação Linfocitária/imunologia , Masculino , Camundongos , Camundongos Knockout , Fator de Transcrição STAT5/metabolismo
9.
Mol Cancer ; 18(1): 84, 2019 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-30961617

RESUMO

BACKGROUND: Long noncoding RNAs (lncRNAs), defined as the transcripts longer than 200 nt without protein-coding capacity, have been found to be aberrantly expressed in diverse human diseases including cancer. A reciprocal translocation between chromosome 9 and 22 generates the chimeric Bcr-Abl oncogene, which is associated with several hematological malignancies. However, the functional relevance between aberrantly expressed lncRNAs and Bcr-Abl-mediated leukemia remains obscure. METHODS: LncRNA cDNA microarray was used to identify novel lncRNAs involved in Bcr-Abl-mediated cellular transformation. To study the functional relevance of novel imatinib-upregulated lncRNA (IUR) family in Abl-induced tumorigenesis, Abl-transformed cell survival and xenografted tumor growth in mice was evaluated. Primary bone marrow transformation and in vivo leukemia transplant using lncRNA-IUR knockdown (KD) transgenic mice were further conducted to corroborate the role of lncRNA-IUR in Abl-induced tumorigenesis. Transcriptome RNA-seq, Western blot, RNA pull down and RNA Immunoprecipitation (RIP) were employed to determine the mechanisms by which lncRNA-IUR-5 regulates Bcr-Abl-mediated tumorigenesis. RESULTS: We identified a conserved lncRNA-IUR family as a key negative regulator of Bcr-Abl-induced tumorigenesis. Increased expression of lncRNA-IUR was detected in both human and mouse Abl-transformed cells upon imatinib treatment. In contrast, reduced expression of lncRNA-IUR was observed in the peripheral blood lymphocytes derived from Bcr-Abl-positive acute lymphoblastic leukemia (ALL) patients compared to normal subjects. Knockdown of lncRNA-IUR remarkably promoted Abl-transformed leukemic cell survival and xenografted tumor growth in mice, whereas overexpression of lncRNA-IUR had opposite effects. Also, silencing murine lncRNA-IUR promoted Bcr-Abl-mediated primary bone marrow transformation and Abl-transformed leukemia cell survival in vivo. Besides, knockdown of murine lncRNA-IUR in transgenic mice provided a favorable microenvironment for development of Abl-mediated leukemia. Finally, we demonstrated that lncRNA-IUR-5 suppressed Bcr-Abl-mediated tumorigenesis by negatively regulating STAT5-mediated expression of CD71. CONCLUSIONS: The results suggest that lncRNA-IUR may act as a critical tumor suppressor in Bcr-Abl-mediated tumorigenesis by suppressing the STAT5-CD71 pathway. This study provides new insights into functional involvement of lncRNAs in leukemogenesis.


Assuntos
Antígenos CD/genética , Proteínas de Fusão bcr-abl/genética , Regulação Neoplásica da Expressão Gênica , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , RNA Longo não Codificante/genética , Receptores da Transferrina/genética , Fator de Transcrição STAT5/genética , Adolescente , Adulto , Animais , Antígenos CD/metabolismo , Antineoplásicos/farmacologia , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Linhagem Celular Tumoral , Pré-Escolar , Feminino , Proteínas de Fusão bcr-abl/metabolismo , Humanos , Mesilato de Imatinib/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Masculino , Camundongos , Camundongos Nus , Oligorribonucleotídeos/genética , Oligorribonucleotídeos/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , RNA Longo não Codificante/agonistas , RNA Longo não Codificante/antagonistas & inibidores , RNA Longo não Codificante/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Receptores da Transferrina/metabolismo , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Anticancer Res ; 39(4): 1705-1710, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30952709

RESUMO

BACKGROUND/AIM: Fluorescent gold nanoparticles demonstrate strong photoluminescence, photostability, and low cellular toxicity, making them attractive agents for biomedical applications. Mechano-growth factor (MGF) is an isoform of IGF1 and its expression has been demonstrated in malignancies including prostate cancer. MATERIALS AND METHODS: Near-infrared-emitting gold nanoparticles (AuNPs) were synthesized and conjugated to MGF. Following characterization and confirmation of conjugation, these AuNPs were used to investigate the expression of MGF in colon cancer cell lines (HT29 and SW620) and tissues comparing normal and colon cancer. The prostate cancer cell line PC3 and adenocarcinoma tissues were used as positive controls. RESULTS: Colon cancer cell lines, adenocarcinoma tissues and polyp tissues demonstrated evidence of MGF peptide expression, which was not found in normal colon tissues and human umbilical vein endothelial cells. CONCLUSION: MGF appears to be overexpressed in colon cancer tissues, offering a potential unique target for imaging and drug delivery in colon cancer.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/metabolismo , Imunofluorescência , Ouro , Nanopartículas Metálicas , Pontos Quânticos , Fator de Transcrição STAT5/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Neoplasias Colorretais/patologia , Células HT29 , Humanos , Microscopia Confocal , Microscopia de Fluorescência , Células PC-3
11.
Nature ; 569(7754): 73-78, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30996346

RESUMO

Polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) are pathologically activated neutrophils that are crucial for the regulation of immune responses in cancer. These cells contribute to the failure of cancer therapies and are associated with poor clinical outcomes. Despite recent advances in the understanding of PMN-MDSC biology, the mechanisms responsible for the pathological activation of neutrophils are not well defined, and this limits the selective targeting of these cells. Here we report that mouse and human PMN-MDSCs exclusively upregulate fatty acid transport protein 2 (FATP2). Overexpression of FATP2 in PMN-MDSCs was controlled by granulocyte-macrophage colony-stimulating factor, through the activation of the STAT5 transcription factor. Deletion of FATP2 abrogated the suppressive activity of PMN-MDSCs. The main mechanism of FATP2-mediated suppressive activity involved the uptake of arachidonic acid and the synthesis of prostaglandin E2. The selective pharmacological inhibition of FATP2 abrogated the activity of PMN-MDSCs and substantially delayed tumour progression. In combination with checkpoint inhibitors, FATP2 inhibition blocked tumour progression in mice. Thus, FATP2 mediates the acquisition of immunosuppressive activity by PMN-MDSCs and represents a target to inhibit the functions of PMN-MDSCs selectively and to improve the efficiency of cancer therapy.


Assuntos
Proteínas de Transporte de Ácido Graxo/metabolismo , Ácidos Graxos/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Neutrófilos/metabolismo , Idoso , Animais , Ácido Araquidônico/metabolismo , Dinoprostona/metabolismo , Proteínas de Transporte de Ácido Graxo/antagonistas & inibidores , Feminino , Humanos , Metabolismo dos Lipídeos , Lipídeos , Masculino , Camundongos , Pessoa de Meia-Idade , Neutrófilos/patologia , Fator de Transcrição STAT5/metabolismo
12.
Int J Mol Sci ; 20(5)2019 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-30818760

RESUMO

O-GlcNAcylation is a post-translational modification that influences tyrosine phosphorylation in healthy and malignant cells. O-GlcNAc is a product of the hexosamine biosynthetic pathway, a side pathway of glucose metabolism. It is essential for cell survival and proper gene regulation, mirroring the metabolic status of a cell. STAT3 and STAT5 proteins are essential transcription factors that can act in a mutational context-dependent manner as oncogenes or tumor suppressors. They regulate gene expression for vital processes such as cell differentiation, survival, or growth, and are also critically involved in metabolic control. The role of STAT3/5 proteins in metabolic processes is partly independent of their transcriptional regulatory role, but is still poorly understood. Interestingly, STAT3 and STAT5 are modified by O-GlcNAc in response to the metabolic status of the cell. Here, we discuss and summarize evidence of O-GlcNAcylation-regulating STAT function, focusing in particular on hyperactive STAT5A transplant studies in the hematopoietic system. We emphasize that a single O-GlcNAc modification is essential to promote development of neoplastic cell growth through enhancing STAT5A tyrosine phosphorylation. Inhibition of O-GlcNAcylation of STAT5A on threonine 92 lowers tyrosine phosphorylation of oncogenic STAT5A and ablates malignant transformation. We conclude on strategies for new therapeutic options to block O-GlcNAcylation in combination with tyrosine kinase inhibitors to target neoplastic cancer cell growth and survival.


Assuntos
Metabolismo Energético , Neoplasias/metabolismo , Neoplasias/patologia , Fator de Transcrição STAT5/metabolismo , Animais , Proliferação de Células , Sobrevivência Celular , Glicosilação , Humanos , Transdução de Sinais
13.
Cell Physiol Biochem ; 52(1): 141-155, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30790510

RESUMO

BACKGROUND/AIMS: Type 1 Diabetes (T1D) involves autoimmune attack due to reduced regulatory T cells as an effect of mutant Stat5b(C1462A) in non-obese diabetic (NOD) mice, a T1D model resulting in pancreatic ß-cell destruction. Although reactive oxygen species are considered to orchestrate the immune attack, the role of nitric oxide (·NO) still remains debatable. Since JAK-STAT pathway is known to induce Nos2, we investigated the role of STAT5B in nitric oxide generation and oxidative stress. METHODS: In this study, we have used chromatin immunoprecipitation with STAT5B antibody to explore whether STAT5B binds Nos2 promoter. Using Stat5b gene silencing and overexpression models in MIN6 mouse pancreatic ß-cell line we have assayed nitric oxide and its end products, superoxide levels, H2O2 levels, and expression of genes related to redox pathway by immunocytochemistry, biochemical assays, quantitative real time PCR and western blotting. RESULTS: Our results prove that STAT5B binds to the candidate gamma-interferon-activated (GAS) element in Nos2 promoter thereby inducing Nos2 mRNA transcription resulting in NOS2 protein expression in MIN6, a mouse pancreatic ß-cell line. Our findings are substantiated by reduced ·NO as well as nitric oxide end products (nitrate and nitrite), and increased superoxide production in Stat5b silenced MIN6 cells. Our results indicate that C1462A mutant STAT5B shows lack of ·NO generation ability. To detoxify excess superoxide as a consequence of lowered Nos2, an overexpressed SOD2 in Stat5b silenced cells results in increased H2O2 production. H2O2 metabolizing enzymes do not show upregulation upon Stat5b silencing, and thus oxidative stress is brought about by amassed H2O2. Stat5b silencing finally reduces AKT expression, a prosurvival signal. CONCLUSION: Our study enables us to conclude that ß-cell stress is aggravated by the incapability of STAT5B to induce Nos2 resulting in H2O2 accumulation and the ensuing oxidative stress enhances ß-cell damage.


Assuntos
Células Secretoras de Insulina/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico/metabolismo , Fator de Transcrição STAT5/metabolismo , Substituição de Aminoácidos , Animais , Linhagem Celular , Peróxido de Hidrogênio/farmacologia , Camundongos Endogâmicos NOD , Mutação de Sentido Incorreto , Óxido Nítrico/genética , Óxido Nítrico Sintase Tipo II/genética , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Regiões Promotoras Genéticas , Ligação Proteica , Fator de Transcrição STAT5/genética
14.
J Exp Clin Cancer Res ; 38(1): 49, 2019 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-30717771

RESUMO

BACKGROUND: The JAK2-STAT signaling pathway plays a critical role in myeloproliferative neoplasms (MPN). An activating mutation in JAK2 (V617F) is present in ~ 95% of polycythemia vera, essential thrombocythemia, and primary myelofibrosis cases. This study aims to explore the selective JAK2V617F inhibitor, evaluate the efficacy and possible mechanism of ZT55 on MPN. METHODS: HTRF assays were conducted to evaluate the selective inhibition of ZT55 for JAKs. Cell apoptosis, proliferation, and cycle arrest assays were performed to examine the effect of ZT55 on HEL cell line with JAK2V617F mutation in vitro. Western analysis was used to monitor the expression and activity of proteins on JAK2/STAT pathway. A mice xenograft model was established to evaluate the antitumor efficacy of ZT55 in vivo. Peripheral blood samples from patients with the JAK2V617F mutation were collected to estimate the effect of ZT55 on erythroid colony formation by colony-forming assay. RESULTS: We found that ZT55 showed a selective inhibition of a 0.031 µM IC50 value against JAK2. It exhibited potent effects on the cellular JAK-STAT pathway, inhibiting tyrosine phosphorylation in JAK2V617F and downstream STAT3/5 transcription factors. ZT55 inhibited the proliferation of the JAK2V617F-expressing HEL cell line, leading to cell cycle arrest at the G2/M phase and induction of caspase-dependent apoptosis. Notably, ZT55 also significantly suppressed the growth of HEL xenograft tumors in vivo. Further evaluation indicated that ZT55 blocked erythroid colony formation of peripheral blood hematopoietic progenitors from patients carrying the JAK2V617F mutation. CONCLUSION: These results suggest that ZT55 is a highly-selective JAK2 inhibitor that can induce apoptosis of human erythroleukemia cells by inhibiting the JAK2-STAT signaling.


Assuntos
Medicamentos de Ervas Chinesas/uso terapêutico , Janus Quinase 2/antagonistas & inibidores , Transtornos Mieloproliferativos/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Medicamentos de Ervas Chinesas/administração & dosagem , Medicamentos de Ervas Chinesas/farmacologia , Feminino , Humanos , Isatis/química , Janus Quinase 2/sangue , Janus Quinase 2/genética , Masculino , Camundongos , Camundongos Nus , Transtornos Mieloproliferativos/sangue , Transtornos Mieloproliferativos/genética , Células-Tronco Neoplásicas/efeitos dos fármacos , Mutação Puntual , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT5/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Leukemia ; 33(7): 1583-1597, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30679796

RESUMO

Deregulation of the Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathway is found in cancer with STAT5A/B controlling leukemic cell survival and disease progression. As mutations in STAT5B, but not STAT5A, have been frequently described in hematopoietic tumors, we used BCR/ABL as model systems to investigate the contribution of STAT5A or STAT5B for leukemogenesis. The absence of STAT5A decreased cell survival and colony formation. Even more drastic effects were observed in the absence of STAT5B. STAT5B-deficient cells formed BCR/ABL+ colonies or stable cell lines at low frequency. The rarely evolving Stat5b-/- cell lines expressed enhanced levels of BCR/ABL oncoprotein compared to wild-type cells. In line, Stat5b-/- leukemic cells induced leukemia with a significantly prolonged disease onset, whereas Stat5a-/- cells rapidly caused a fatal disease superimposable to wild-type cells. RNA-sequencing (RNA-seq) profiling revealed a marked enhancement of interferon (IFN)-α and IFN-γ signatures in Stat5b-/- cells. Inhibition of IFN responses rescued BCR/ABL+ colony formation of Stat5b-/--deficient cells. A downregulated IFN response was also observed in patients suffering from leukemia carrying STAT5B mutations. Our data define STAT5B as major STAT5 isoform driving BCR/ABL+ leukemia. STAT5B enables transformation by suppressing IFN-α/γ, thereby facilitating leukemogenesis. Our findings might help explain the high frequency of STAT5B mutations in hematopoietic tumors.


Assuntos
Transformação Celular Neoplásica/patologia , Proteínas de Fusão bcr-abl/metabolismo , Leucemia Linfocítica Granular Grande/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Mutação , Fator de Transcrição STAT5/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Antineoplásicos/farmacologia , Proliferação de Células , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/metabolismo , Proteínas de Fusão bcr-abl/genética , Humanos , Interferons/farmacologia , Leucemia Linfocítica Granular Grande/tratamento farmacológico , Leucemia Linfocítica Granular Grande/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Fator de Transcrição STAT5/genética , Taxa de Sobrevida , Proteínas Supressoras de Tumor/genética , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Nat Commun ; 10(1): 66, 2019 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-30622248

RESUMO

Protein-templated fragment ligations have been established as a powerful method for the assembly and detection of optimized protein ligands. Initially developed for reversible ligations, the method has been expanded to irreversible reactions enabling the formation of super-additive fragment combinations. Here, protein-induced Mannich ligations are discovered as a biocatalytic reaction furnishing inhibitors of the transcription factor STAT5. STAT5 protein catalyzes multicomponent reactions of a phosphate mimetic, formaldehyde, and 1H-tetrazoles yielding protein ligands with greatly increased binding affinity and ligand efficiency. Reactions are induced under physiological conditions selectively by native STAT5 but not by other proteins. Formation of ligation products and (auto-)inhibition of the reaction are quantified and the mechanism is investigated. Inhibitors assembled by STAT5 block specifically the phosphorylation of this protein in a cellular model of acute myeloid leukemia (AML), DNA-binding of STAT5 dimers, expression of downstream targets of the transcription factor, and the proliferation of cancer cells in mice.


Assuntos
Antineoplásicos/síntese química , Biocatálise , Leucemia Mieloide Aguda/tratamento farmacológico , Fator de Transcrição STAT5/química , Proteínas Supressoras de Tumor/química , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , DNA/metabolismo , Desenvolvimento de Medicamentos/métodos , Humanos , Ligantes , Camundongos , Camundongos Endogâmicos NOD , Simulação de Acoplamento Molecular , Fosforilação/efeitos dos fármacos , Fator de Transcrição STAT5/antagonistas & inibidores , Fator de Transcrição STAT5/metabolismo , Proteínas Supressoras de Tumor/antagonistas & inibidores , Proteínas Supressoras de Tumor/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Eur J Histochem ; 63(1)2019 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-30652434

RESUMO

Prolactin (PRL) production in mammals has been demonstrated in extrapituitary gland, which can activate autocrine/paracrine signaling pathways to regulate physiological activity. In the current study, we characterized the gene expression profiles of PRL, prolactin receptor (PRLR) and signal transducers and activators of transcription 5 (STAT5) in the scented glandular tissues of the muskrats, to further elucidate the relationship between PRL and the scented glandular functions of the muskrats. The weight and volume of the scented glands in the breeding season were significantly higher than those of the non-breeding season. Immunohistochemical data showed that PRL, PRLR and STAT5/phospho-STAT5 (pSTAT5) were found in the glandular and epithelial cells of the scented glands in both seasons. Furthermore, we found that PRL, PRLR and STAT5 had higher immunoreactivities in the scented glands during the breeding season when compared to those of the non-breeding season. In parallel, the gene expressions of PRL, PRLR and STAT5 were significantly higher in the scented glands during the breeding season than those of the non-breeding season. The concentrations of PRL in scented glandular tissues and sera were measured by enzyme-linked immunosorbent assay (ELISA), and their levels were both notably higher in the breeding season than those of the non-breeding season. These findings suggested that the scented glands of the muskrats were capable of extrapituitary synthesis of PRL, which might attribute PRL a specific function to an endocrine or autocrine/paracrine mediator.


Assuntos
Regulação da Expressão Gênica , Prolactina/genética , RNA Mensageiro/metabolismo , Receptores da Prolactina/genética , Fator de Transcrição STAT5/genética , Glândulas Odoríferas/metabolismo , Animais , Arvicolinae , Imuno-Histoquímica , Masculino , Receptores da Prolactina/metabolismo , Reprodução/fisiologia , Fator de Transcrição STAT5/metabolismo , Estações do Ano
18.
Int J Med Sci ; 16(1): 167-179, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30662340

RESUMO

The proliferation and adipogenesis of preadipocytes played important roles in the development of adipose tissue and contributed much to the processes of obesity. On the other hand, lipopolysaccharide (LPS), also known as endotoxin, is a key outer membrane component of gram-negative bacteria in the gut microbiota, and has a dominant role in linking inflammation to high-fat diet-induced metabolic syndrome. Studies suggested the potential roles of LPS in hepatic steatosis and in obese mice models. However, the molecular mechanisms underlying LPS-regulated obesity remained largely unknown. Here we reported that LPS stimulated expression of cyosolic phospholipase A2 (cPLA2), one of inflammation regulators of obesity, in the preadipocytes. Pretreatment the inhibitors of JAK2, STAT3, STAT5 or AMPK significantly reduced LPS-increased mRNA and protein expression of cPLA2 together with phosphorylation of JAK2, STAT3, STAT5 and AMPK, separately. Similarly, transfection of siRNA against JAK2 or AMPK abolished expression of cPLA2 and phosphorylation of JAK2 or AMPK together with downregulated expression of JAK2 and AMPK protein. LPS enhanced activation of STAT3 and STAT5 via JAK2-dependent manner in the preadipocytes. Transfection of JAK2 or AMPK siRNA further proofed the independence of JAK2 and AMPK in LPS-treated preadipocytes. In addition, LPS-increased DNA synthesis, cell numbers and cell viability of preadipocytes were attenuated by AACOCF3, AG490, BML-275, cPLA2 siRNA, JAK2 siRNA or AMPK siRNA. Attenuation JAK2/STAT or AMPK-dependent cPLA2 expression reduced LPS-mediated adipogenesis of preadipocytes. Stimulation of arachidonic acid or AMPK activator, A-769662, increased cell numbers and cell viability and promoted differentiation of preadipocytes. Collectively, these results indicated that LPS increased preadipocytes proliferation and adipogenesis via JAK/STAT and AMPK-dependent cPLA2 expression. The mechanisms of LPS-stimulated cPLA2 expression may be a link between bacteria and obesity and provides the molecular basis for preventing metabolic syndrome or hyperplasic obesity.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Adipócitos , Adipogenia/efeitos dos fármacos , Janus Quinase 2/metabolismo , Lipopolissacarídeos/farmacologia , Fosfolipases A2 Citosólicas/metabolismo , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT5/metabolismo , Células 3T3-L1 , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Endotoxinas/farmacologia , Camundongos
19.
Exp Mol Med ; 51(1): 2, 2019 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-30617282

RESUMO

Growth hormone receptor (GHR) plays a vital role in breast cancer chemoresistance and metastasis but the mechanism is not fully understood. We determined if GHR could be a potential therapeutic target for estrogen receptor negative (ER-ve) breast cancer, which are highly chemoresistant and metastatic. GHR was stably knocked down in ER-ve breast cancer cells and its effect on cell proliferation, metastatic behavior, and chemosensitivity to docetaxel (DT) was assessed. Microarray analysis was performed to identify potential GHR downstream targets involved in chemoresistance. GHR and ATP-binding cassette sub-family G member 2 (ABCG2) overexpression and knockdown studies were performed to investigate the mechanism of GHR-induced chemoresistance. Patient-derived xenografts was used to study the effect of GHR and ABCG2. Immunohistochemical data was used to determine the correlation between GHR, pAKT, pmTOR, and ABCG2 expressions. GHR silencing drastically reduced the chemoresistant and metastatic behavior of ER-ve breast cancer cells and also inhibited AKT/mTOR pathway. In contrast, activation, or overexpression of GHR increased chemoresistance and metastasis by increasing the expression and promoter activity, of ABCG2. Inhibition of JAK2/STAT5 signaling repressed GHR-induced ABCG2 promoter activity and expression. Further, ABCG2 knockdown significantly increased the chemosensitivity. Finally, patient-derived xenograft studies revealed the role of GHR in chemoresistance. Overall, these findings demonstrate that targeting GHR could be a novel therapeutic approach to overcome chemoresistance and associated metastasis in aggressive ER-ve breast cancers.


Assuntos
Inativação Gênica , Neoplasias Mamárias Experimentais/terapia , Terapêutica com RNAi/métodos , Receptores da Somatotropina/genética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Feminino , Humanos , Janus Quinase 2/metabolismo , Células MCF-7 , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos Nus , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Estrogênicos/genética , Receptores da Somatotropina/metabolismo , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo
20.
Proc Natl Acad Sci U S A ; 116(1): 211-216, 2019 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-30559202

RESUMO

Bone marrow (BM) produces all blood and immune cells deriving from hematopoietic stem cells (HSCs). The decrease of immune cell production during aging is one of the features of immunosenescence. The impact of redox dysregulation in BM aging is still poorly understood. Here we use TP53INP1-deficient (KO) mice endowed with chronic oxidative stress to assess the influence of aging-associated redox alterations in BM homeostasis. We show that TP53INP1 deletion has no impact on aging-related accumulation of HSCs. In contrast, the aging-related contraction of the lymphoid compartment is mitigated in TP53INP1 KO mice. B cells that accumulate in old KO BM are differentiating cells that can mature into functional B cells. Importantly, this phenotype results from B cell-intrinsic events associated with defective redox control. Finally, we show that oxidative stress in aged TP53INP1-deficient mice maintains STAT5 expression and activation in early B cells, driving high Pax5 expression, which provides a molecular mechanism for maintenance of B cell development upon aging.


Assuntos
Linfócitos B/fisiologia , Medula Óssea/fisiologia , Linfopoese , Proteínas Nucleares/deficiência , Receptores de Interleucina-7/metabolismo , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais , Envelhecimento/fisiologia , Animais , Linfócitos B/metabolismo , Medula Óssea/metabolismo , Linfopoese/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Oxirredução , Estresse Oxidativo
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