RESUMO
Breast cancer is a heterogeneous disease that represents the most common cancer around the world; it comprises 12% of new cases according to the World Health Organization. Despite new approaches in early diagnosis and current treatment, breast cancer is still the leading cause of death for cancer mortality. New targeted therapies against key signalling transduction molecules are required. Phosphoinositide 3-kinase (PI3K) regulates multiple biological functions such as proliferation, survival, migration, and growth. It is well established that PI3K isoform-selective inhibitors show fewer toxic side effects compared to broad spectrum inhibition of PI3K (pan-PI3K inhibitors). Therefore, we tested the PI3K p110δ-selective inhibitor, IC87114, and Vps34-selective inhibitor, Vps34-IN1, on the breast cancer cell lines MCF-7 and MDA-MB-231, representing hormone-responsive and triple-negative breast cancer cells, respectively. Our data show that both inhibitors decreased migration of MCF-7 and MDA-MB-231 cells, and Vps34 also significantly impacted MCF-7 cell proliferation. Three-dimensional (3D) in vitro culture models show that IC87114 and Vps34-IN1 treatment reduced the growth of MCF-7 and MDA-MB-231 cells in 3D tumour spheroid cultures. This study identifies IC87114 and Vps34-IN1 as potential therapeutic approaches in breast cancer.
Assuntos
Neoplasias da Mama , Classe III de Fosfatidilinositol 3-Quinases , Inibidores de Fosfoinositídeo-3 Quinase , Neoplasias de Mama Triplo Negativas , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Classe III de Fosfatidilinositol 3-Quinases/antagonistas & inibidores , Classe Ia de Fosfatidilinositol 3-Quinase , Feminino , Humanos , Células MCF-7 , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismoRESUMO
Steroid hormones and growth factors control neuritogenesis through their cognate receptors under physiological and pathological conditions. We have already shown that nerve growth factor and androgens induce neurite outgrowth of PC12 cells through a reciprocal crosstalk between the NGF receptor, TrkA and the androgen receptor. Here, we report that androgens or NGF induce neuritogenesis in PC12 cells through inactivation of RhoA. Ectopic expression of the dominant negative RhoA N19 promotes, indeed, the neurite-elongation of unchallenged and androgen- or NGF-challenged PC12 cells and the increase in the expression levels of ßIII tubulin, a specific neuronal marker. Pharmacological inhibition of the Ser/Thr kinase ROCK, an RhoA effector, induces neuritogenesis in unchallenged PC12 cells, and potentiates the effect of androgens and NGF, confirming the role of RhoA/ROCK axis in the neuritogenesis induced by androgen and NGF, through the phosphorylation of Akt. These findings suggest that therapies based on new selective androgen receptor modulators and/or RhoA/ROCK inhibitors might exert beneficial effects in the treatment of neuro-disorders, neurological diseases and ageing-related processes.
Assuntos
Androgênios , Neuritos , Animais , Ratos , Androgênios/farmacologia , Androgênios/metabolismo , Fator de Crescimento Neural/farmacologia , Fator de Crescimento Neural/metabolismo , Neuritos/metabolismo , Crescimento Neuronal , Células PC12 , Receptores Androgênicos/metabolismoRESUMO
BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is the most common subtype of renal cancer. Large-scale metabolomic data have associated metabolic alterations with the pathogenesis and progression of renal carcinoma and have correlated mitochondrial activity with poor survival in a subset of patients. The aim of this study was to determine whether targeting mitochondria-lysosome interaction could be a novel therapeutic approach using patient-derived organoids as avatar for drug response. METHODS: RNAseq data analysis and immunohistochemistry were used to show overexpression of Purinergic receptor 4 (P2XR4) in clear cell carcinomas. Seahorse experiments, immunofluorescence and fluorescence cell sorting were used to demonstrate that P2XR4 regulates mitochondrial activity and the balance of radical oxygen species. Pharmacological inhibitors and genetic silencing promoted lysosomal damage, calcium overload in mitochondria and cell death via both necrosis and apoptosis. Finally, we established patient-derived organoids and murine xenograft models to investigate the antitumor effect of P2XR4 inhibition using imaging drug screening, viability assay and immunohistochemistry. RESULTS: Our data suggest that oxo-phosphorylation is the main source of tumor-derived ATP in a subset of ccRCC cells expressing P2XR4, which exerts a critical impact on tumor energy metabolism and mitochondrial activity. Prolonged mitochondrial failure induced by pharmacological inhibition or P2XR4 silencing was associated with increased oxygen radical species, changes in mitochondrial permeability (i.e., opening of the transition pore complex, dissipation of membrane potential, and calcium overload). Interestingly, higher mitochondrial activity in patient derived organoids was associated with greater sensitivity to P2XR4 inhibition and tumor reduction in a xenograft model. CONCLUSION: Overall, our results suggest that the perturbed balance between lysosomal integrity and mitochondrial activity induced by P2XR4 inhibition may represent a new therapeutic strategy for a subset of patients with renal carcinoma and that individualized organoids may be help to predict drug efficacy.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Humanos , Animais , Camundongos , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Receptores Purinérgicos P2X4/metabolismo , Cálcio/metabolismo , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Mitocôndrias/metabolismo , Linhagem Celular TumoralRESUMO
Physiological apoptosis is induced by a switch from survival to death signalling. Dysregulation of this process is frequently associated with cancer. A powerful model for this apoptotic switch is mammary gland involution, during which redundant milk-producing epithelial cells undergo apoptosis. Signal transducer and activator of transcription 3 (Stat3) is an essential mediator of this switch but the mechanism has not yet been defined. Stat3-dependent cell death during involution can be blocked by activation of Akt/protein kinase B (PKB), a downstream effector of the phosphoinositide-3-OH kinase (PI(3)K) pathway. Here we show that expression of the PI(3)K regulatory subunits p55alpha and p50alpha is induced by Stat3 during involution. In the absence of Stat3 in vivo, upregulation of p55alpha and p50alpha is abrogated, levels of activated Akt are sustained and apoptosis is prevented. Chromatin immunoprecipitation assays show that Stat3 binds directly to the p55alpha and p50alpha promoters in vivo. Overexpression of either p55alpha or p50alpha reduces levels of activated Akt. We propose a novel mechanism in which Stat3 regulates apoptosis by inducing expression of distinct PI(3)K regulatory subunits to downregulate PI(3)K-Akt-mediated survival signalling.
Assuntos
Apoptose/fisiologia , Proteínas de Ligação a DNA/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Transativadores/metabolismo , Animais , Linhagem Celular , Células Epiteliais/metabolismo , Camundongos , Camundongos Knockout , Camundongos Mutantes , Dados de Sequência Molecular , Fosfatidilinositol 3-Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Subunidades Proteicas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Fator de Transcrição STAT3 , Transdução de Sinais/fisiologiaRESUMO
The p110 isoforms of phosphoinositide 3-kinase (PI3K) are acutely regulated by extracellular stimuli. The class IA PI3K catalytic subunits (p110alpha, p110beta, and p110delta) occur in complex with a Src homology 2 (SH2) domain-containing p85 regulatory subunit, which has been shown to link p110alpha and p110delta to Tyr kinase signaling pathways. The p84/p101 regulatory subunits of the p110gamma class IB PI3K lack SH2 domains and instead couple p110gamma to G protein-coupled receptors (GPCRs). Here, we show, using small-molecule inhibitors with selectivity for p110beta and cells derived from a p110beta-deficient mouse line, that p110beta is not a major effector of Tyr kinase signaling but couples to GPCRs. In macrophages, both p110beta and p110gamma contributed to Akt activation induced by the GPCR agonist complement 5a, but not by the Tyr kinase ligand colony-stimulating factor-1. In fibroblasts, which express p110beta but not p110gamma, p110beta mediated Akt activation by the GPCR ligands stromal cell-derived factor, sphingosine-1-phosphate, and lysophosphatidic acid but not by the Tyr kinase ligands PDGF, insulin, and insulin-like growth factor 1. Introduction of p110gamma in these cells reduced the contribution of p110beta to GPCR signaling. Taken together, these data show that p110beta and p110gamma can couple redundantly to the same GPCR agonists. p110beta, which shows a much broader tissue distribution than the leukocyte-restricted p110gamma, could thus provide a conduit for GPCR-linked PI3K signaling in the many cell types where p110gamma expression is low or absent.
Assuntos
Fosfatidilinositol 3-Quinases/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animais , Classe I de Fosfatidilinositol 3-Quinases , Complemento C5a/farmacologia , Fibroblastos/enzimologia , Teste de Complementação Genética , Isoenzimas/genética , Isoenzimas/metabolismo , Ligantes , Fator Estimulador de Colônias de Macrófagos/farmacologia , Macrófagos/enzimologia , Camundongos , Camundongos Mutantes , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Acoplados a Proteínas G/agonistas , Transdução de SinaisRESUMO
The tumor microenvironment modulates cancer growth. Extracellular vesicles (EVs) have been identified as key mediators of intercellular communication, but their role in tumor growth is largely unexplored. Here, we demonstrate that EVs from sarcoma patients promote neoangiogenesis via a purinergic X receptor 4 (P2XR4) -dependent mechanism in vitro and in vivo. Using a proteomic approach, we analyzed the protein content of plasma EVs and identified critical activated pathways in human umbilical vein endothelial cells (HUVECs) and human progenitor hematopoietic cells (CD34+). We then showed that vessel formation was due to rapid mitochondrial activation, intracellular Ca2+ mobilization, increased extracellular ATP, and trafficking of the lysosomal P2XR4 to the cell membrane, which is required for cell motility and formation of stable branching vascular networks. Cell membrane translocation of P2XR4 was induced by proteins and chemokines contained in EVs (e.g. Del-1 and SDF-1). Del-1 was found expressed in many EVs from sarcoma tumors and several tumor types. P2XR4 blockade reduced EVs-induced vessels in angioreactors, as well as intratumor vascularization in mouse xenografts. Together, these findings identify P2XR4 as a key mediator of EVs-induced tumor angiogenesis via a signaling mediated by mitochondria-lysosome-sensing response in endothelial cells, and indicate a novel target for therapeutic interventions.
Assuntos
Micropartículas Derivadas de Células/metabolismo , Lisossomos/metabolismo , Neovascularização Patológica/metabolismo , Receptores Purinérgicos P2X4/metabolismo , Sarcoma/irrigação sanguínea , Sarcoma/patologia , Animais , Cálcio/metabolismo , Movimento Celular , Citosol/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Camundongos , Mitocôndrias/metabolismo , Retina/patologia , Sarcoma/sangue , Transdução de Sinais , ViscosidadeRESUMO
Inflammatory substances released by mast cells induce and maintain the allergic response. Mast cell differentiation and activation are regulated, respectively, by stem cell factor (SCF; also known as Kit ligand) and by allergen in complex with allergen-specific immunoglobulin E (IgE). Activated SCF receptors and high-affinity receptors for IgE (FcvarepsilonRI) engage phosphoinositide 3-kinases (PI(3)Ks) to generate intracellular lipid second messenger signals. Here, we report that genetic or pharmacological inactivation of the p110delta isoform of PI(3)K in mast cells leads to defective SCF-mediated in vitro proliferation, adhesion and migration, and to impaired allergen-IgE-induced degranulation and cytokine release. Inactivation of p110delta protects mice against anaphylactic allergic responses. These results identify p110delta as a new target for therapeutic intervention in allergy and mast-cell-related pathologies.
Assuntos
Hipersensibilidade/enzimologia , Mastócitos/enzimologia , Fosfatidilinositol 3-Quinases/metabolismo , Anafilaxia/enzimologia , Anafilaxia/imunologia , Animais , Adesão Celular/efeitos dos fármacos , Contagem de Células , Degranulação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Classe I de Fosfatidilinositol 3-Quinases , Citocinas/metabolismo , Derme/citologia , Genes Essenciais/genética , Humanos , Hipersensibilidade/imunologia , Interleucina-3/farmacologia , Isoenzimas/antagonistas & inibidores , Isoenzimas/genética , Isoenzimas/metabolismo , Mastócitos/citologia , Mastócitos/imunologia , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/genética , Inibidores de Fosfoinositídeo-3 Quinase , Receptores de IgE/imunologia , Sistemas do Segundo Mensageiro/fisiologia , Fator de Células-Tronco/farmacologiaRESUMO
Phosphoinositide 3-kinases (PI3Ks) generate lipids that control a wide variety of intracellular signalling pathways. Part of this diversity in PI3K actions stems from the broad range of protein effectors of the PI3K lipids. A further layer of complexity is added by the existence of multiple isoforms of PI3K. Gene-targeting studies in the mouse have recently uncovered key roles for specific PI3K isoforms in immunity, metabolism and cardiac function. Remarkably, some of these actions do not require PI3K catalytic activity. In addition, loss-of-expression of certain PI3K genes leads to increased PI3K signalling following insulin stimulation. PI3K gene targeting has, in many cases, led to altered expression of the non-targeted PI3K subunits, making it difficult to exclude that some of the reported phenotypes result from 'knock-on' effects of PI3K gene deletion. Targeting strategies that take into account the complex interplay between members of the PI3K family will be crucial to gain a full understanding of the physiological roles of the isoforms of PI3K.
Assuntos
Marcação de Genes , Isoenzimas/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/fisiologia , Animais , Insulina/metabolismo , Isoenzimas/genética , Leucócitos/enzimologia , Camundongos , Camundongos Knockout , Fenótipo , Fosfatidilinositol 3-Quinases/genética , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismoRESUMO
In NIH3T3 cells, 0.001 nM of the synthetic androgen R1881 induces and stimulates association of androgen receptor (AR) with Src and phosphatidylinositol 3-kinase (Pl3-kinase), respectively, thereby triggering S-phase entry. 10 nM R1881 stimulates Rac activity and membrane ruffling in the absence of the receptor-Src-PI3-kinase complex assembly. The antiandrogen Casodex and specific inhibitors of Src and PI3-kinase prevent both hormonal effects, DNA synthesis and cytoskeletal changes. Neither low nor high R1881 concentration allows receptor nuclear translocation and receptor-dependent transcriptional activity in fibroblasts, although they harbor the classical murine AR. The very low amount of AR in NIH3T3 cells (7% of that present in LNCaP cells) activates the signaling pathways, but apparently is not sufficient to stimulate gene transcription. This view is supported by the appearance of receptor nuclear translocation as well as receptor-mediated transcriptional activity after overexpression of AR in fibroblasts. In addition, AR-negative Cos cells transiently transfected with a very low amount of hAR cDNA respond to low and high R1881 concentrations with signaling activation. Interestingly, they do not show significant transcriptional activation under the same experimental conditions. Fibroblasts are the first example of cells that respond to steroid hormones with activation of signaling pathways in the absence of endogenous receptor transcriptional activity. The data reported also show that hormone concentration can be crucial in determining the type of cell responsiveness.
Assuntos
Androgênios/metabolismo , Citoesqueleto/metabolismo , DNA/biossíntese , Fibroblastos/metabolismo , Receptores Androgênicos/metabolismo , Células 3T3 , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Transporte Ativo do Núcleo Celular/genética , Androgênios/farmacologia , Animais , Anticorpos/farmacologia , Células COS , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/ultraestrutura , DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Feto , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Humanos , Masculino , Camundongos , Receptores Androgênicos/efeitos dos fármacos , Receptores Androgênicos/genética , Fase S/efeitos dos fármacos , Fase S/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Células Estromais/citologia , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Transcrição Gênica/efeitos dos fármacos , Transcrição Gênica/genética , Células Tumorais Cultivadas , Proteínas rac de Ligação ao GTP/efeitos dos fármacos , Proteínas rac de Ligação ao GTP/genética , Proteínas rac de Ligação ao GTP/metabolismoRESUMO
Breast cancer (BC) is the most common cancer among women, and current available therapies often have high success rates. Nevertheless, BC might acquire drug resistance and sometimes relapse. Current knowledge about the most aggressive forms of BC points to the role of specific cells with stem properties located within BC, the so-called "BC stem cells" (BCSCs). The role of BCSCs in cancer formation, growth, invasiveness, therapy resistance and tumor recurrence is becoming increasingly clear. The growth and metastatic properties of BCSCs are regulated by different pathways, which are only partially known. Sex steroid receptors (SSRs), which are involved in BC etiology and progression, promote BCSC proliferation, dedifferentiation and migration. However, in the literature, there is incomplete information about their roles. Particularly, there are contrasting conclusions about the expression and role of the classical BC hormonal biomarkers, such as estrogen receptor alpha (ERα), together with scant, albeit promising information concerning ER beta (ERß) and androgen receptor (AR) properties that control different transduction pathways in BCSCs. In this review, we will discuss the role that SRs expressed in BCSCs play to BC progression and recurrence and how these findings have opened new therapeutic possibilities.
RESUMO
Prostate cancer (PC) remains a widespread malignancy in men. Since the androgen/androgen receptor (AR) axis is associated with the pathogenesis of prostate cancer, suppression of AR-dependent signaling by androgen deprivation therapy (ADT) still represents the primary intervention for this disease. Despite the initial response, prostate cancer frequently develops resistance to ADT and progresses. As such, the disease becomes metastatic and few therapeutic options are available at this stage. Although the majority of studies are focused on the role of AR signaling, compelling evidence has shown that estrogens and their receptors control prostate cancer initiation and progression through a still debated mechanism. Epithelial versus mesenchymal transition (EMT) is involved in metastatic spread as well as drug-resistance of human cancers, and many studies on the role of this process in prostate cancer progression have been reported. We discuss here the findings on the role of estrogen/estrogen receptor (ER) axis in epithelial versus mesenchymal transition of prostate cancer cells. The pending questions concerning this issue are presented, together with the impact of the available data in clinical management of prostate cancer patients.
RESUMO
Breast cancer is one of the most common malignancies in Western society. Localized breast cancer, before it spreads, can be cured by surgery. However, the high mortality rate associated with breast cancer is due to a propensity of the tumor to metastasize when the primary tumor is small or undetectable. Although steroid receptor status has been recognized as the most precise predictor of response to hormone therapy, a significant number of tumors expressing these receptors metastasize and patients do not respond to the antihormone therapy. The mechanism leading to breast cancer progression and resistance to the hormone therapy is not completely understood at the present time. Compelling evidence shows that hormone-bound steroid receptors in breast cancer cells activate complex signaling networks, which include MAPK- and G protein-dependent pathways. These responses, which occur within seconds or minutes after steroid administration, are not due to changes in gene expression. Depending on cell systems, steroid activation of these networks leads to different and profound effects on extra nuclear and nuclear events. In such a way steroids foster cell cycle, reduce apoptosis and stimulate cell migration of target cells. All these processes are deregulated in breast cancer. In this review we will discuss new aspects of signaling pathways activated by steroids and their integration with other pathways in breast cancer. Recent findings on the discovery of compounds specifically interfering in such a complex network will be presented.
Assuntos
Neoplasias da Mama/embriologia , Neoplasias da Mama/metabolismo , AMP Cíclico/metabolismo , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Sistema de Sinalização das MAP Quinases , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Modelos Biológicos , Receptores de Esteroides/metabolismo , Transdução de SinaisRESUMO
Breast cancer (BC) is a hormone-related tumor. Despite the progress in BC therapy, this disease still remains the most common cancer amongst women around the world. This is likely due to the amazing BC heterogeneity. Accumulating evidence suggests a role for androgen signaling in BC. Nevertheless, a precise understanding of the mechanism of androgen action in this disease remains a challenging puzzle. Androgen receptor (AR) is often expressed in BC and several studies suggest that its role depends on the tumor microenvironment as well as the relative levels of circulating estrogens and androgens. However, the AR function in BC is still conflicting. Although AR expression is often associated with a favorable prognosis in EREstradiol Receptorα-positive (ERα +) BC, many findings suggest that, in some instances, high levels of AR can contribute to the therapy-resistance. Again, in ERα negative BC (ERα -), AR is mainly expressed in tumors with apocrine differentiation and a lower Nottingham grade. Moreover, AR stimulates cellular proliferation in triple negative breast cancer (ERα -, PgR -, and HER-2-Neu -). This finding is substantiated by the observation that high levels of circulating androgens are associated with an increased risk of developing BC in post-menopausal woman. Treatment of ERα- BC with AR antagonists, such as bicalutamide or enzalutamide, reduces, indeed, the tumor growth. In this review, we will analyze the putative role of AR in BC. Emerging therapies based on the use of new agonists or antagonists or inhibitors will be here discussed.
RESUMO
Expression of a dominant negative atypical protein kinase C (aPKC), PKCzeta, prevents nuclear translocation of extracellular regulated kinase 2 (ERK-2), p27 nuclear reduction, and DNA synthesis induced by estradiol in human mammary cancer-derived MCF-7 cells. aPKC action upstream of these events has been analyzed. In hormone-stimulated NIH 3T3 and Cos cells ectopically expressing human estrogen receptor alpha (hERalpha), aPKC is activated by phosphatidylinositol 3-kinase (PI 3-kinase) and, in turn, controls the Ras/MEK-1/ERK cascade. In MCF-7 and Cos cells stimulated by hormone, PI 3-kinase activates PKCzeta by Thr410 phosphorylation. Serine phosphorylation of PKCzeta is simultaneously induced. PKCzeta activation leads to recruitment of Ras to a multimolecular complex that also includes hERalpha, Src, PI 3-kinase, and aPKC. We propose that PKCzeta pushes Ras and the signaling complex close together in such a way that it facilitates the Src-dependent Ras activation. This activation is crucial for the interplay between estradiol-triggered signaling and cell cycle machinery.
Assuntos
Estradiol/metabolismo , Fase G1/fisiologia , Isoenzimas/metabolismo , Proteína Quinase C/metabolismo , Fase S/fisiologia , Animais , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p27 , Ativação Enzimática , Receptor alfa de Estrogênio , Humanos , Isoenzimas/genética , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Células NIH 3T3 , Fosfatidilinositol 3-Quinases/metabolismo , Proteína Quinase C/genética , Subunidades Proteicas/metabolismo , Transporte Proteico/fisiologia , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais/fisiologia , Proteínas Supressoras de Tumor/metabolismo , Proteínas ras/metabolismo , Quinases da Família src/metabolismoRESUMO
Under conditions of short-term hormone deprivation, epidermal growth factor (EGF) induces DNA synthesis, cytoskeletal changes, and Src activation in MCF-7 and LNCaP cells. These effects are drastically inhibited by pure estradiol or androgen antagonists, implicating a role of the steroid receptors in these findings. Interestingly, EGF triggers rapid association of Src with androgen receptor (AR) and estradiol receptor alpha (ERalpha) in MCF-7 cells or ERbeta in LNCaP cells. Here, we show that, through EGF receptor (EGFR) and erb-B2, EGF induces tyrosine phosphorylation of ER preassociated with AR, thereby triggering the assembly of ER/AR with Src and EGFR. Remarkably, experiments in Cos cells show that this complex stimulates EGF-triggered EGFR tyrosine phosphorylation. In turn, estradiol and androgen antagonists, through the Src-associated receptors, prevent Src activation by EGF and heavily reduce EGFR tyrosine phosphorylation and the subsequent multiple effects, including DNA synthesis and cytoskeletal changes in MCF-7 cells. In addition, knockdown of ERalpha or AR gene by small interfering RNA (siRNA) almost abolishes EGFR tyrosine phosphorylation and DNA synthesis in EGF-treated MCF-7 cells. The present findings reveal that steroid receptors have a key role in EGF signaling. EGFR tyrosine phosphorylation, depending on Src, is a part of this mechanism. Understanding of EGF-triggered growth and invasiveness of mammary and prostate cancer cells expressing steroid receptors is enhanced by this report, which reveals novel aspects of steroid receptor action.
Assuntos
Antagonistas de Androgênios/farmacologia , Neoplasias da Mama/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/metabolismo , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Quinases da Família src/metabolismo , Antagonistas de Receptores de Andrógenos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , DNA de Neoplasias/biossíntese , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/genética , Feminino , Humanos , Masculino , Fosforilação , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , RNA Interferente Pequeno/genética , Receptores Androgênicos/genética , Transdução de SinaisRESUMO
Endocrine disrupting chemicals (EDCs) are man-made substances widespread in the environment that include, among many others, bisphenol A (BPA), organochlorinated pesticides and hormone derivatives detectable in meat from animals raised in concentrated animal feeding operations. Increasing evidence indicates that EDCs have a negative impact on human health as well as on male and female fertility. They may also be associated with some endocrine diseases and increased incidence of breast and prostate cancer. This review aims to summarize available data on the (potential) impact of some common EDCs, focusing particularly on BPA, prostate cancer and their mechanisms of action. These compounds interfere with normal hormone signal pathway transduction, resulting in prolonged exposure of receptors to stimuli or interference with cellular hormone signaling in target cells. Understanding the effects of BPA and other EDCs as well as their molecular mechanism(s) may be useful in sensitizing the scientific community and the manufacturing industry to the importance of finding alternatives to their indiscriminate use.
Assuntos
Compostos Benzidrílicos/efeitos adversos , Disruptores Endócrinos/efeitos adversos , Fenóis/efeitos adversos , Neoplasias da Próstata/induzido quimicamente , Saúde , Humanos , MasculinoRESUMO
Bisphenol A (BPA) belongs to the class of chemicals known as endocrine disruptors and has been also involved in the pathogenesis and progression of endocrine related cancer such as breast and prostate cancers. Here, we have investigated the effect of BPA in human prostate cancer LNCaP cells and in human non-transformed epithelial prostate EPN cells. Our data showed that BPA induces the down regulation of cyclin D1 expression and the upregulation of the cell cycle inhibitors p21 and p27, leading to cell cycle arrest. Interestingly, we found that the BPA anti-proliferative response depends on a strong and rapid activation of epidermal growth factor receptor (EGFR), which stimulates ERK-dependent pathway. This, in turn, induces expression of p53 and its phosphorylation on residue Ser15, which is responsible for cell cycle arrest. EGFR activation occurs upon a cross talk with androgen (AR) and estradiol receptor-ß (ERß) which are known to bind BPA. Altogether, these findings show a novel signaling pathway in which EGFR activation plays a key role on BPA-induced cell cycle inhibition through a pathway involving AR and ERß/EGFR complexes, ERK and p53. Our results provide new insights for understanding the molecular mechanisms in human prostate cancer. On the other, they could allow the development of new compounds that may be used to overcome human prostate cancer resistance to endocrine therapy in promising target therapeutic approaches.
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Inflammatory cells play key roles in restenosis upon vascular surgical procedures such as bypass grafts, angioplasty and stent deployment but the molecular mechanisms by which these cells affect restenosis remain unclear. The p110δ isoform of phosphoinositide 3-kinase (PI3K) is mainly expressed in white blood cells. Here, we have investigated whether p110δ PI3K is involved in the pathogenesis of restenosis in a mouse model of carotid injury, which mimics the damage following arterial grafts. We used mice in which p110δ kinase activity has been disabled by a knockin (KI) point mutation in its ATP-binding site (p110δD910A/D910A PI3K mice). Wild-type (WT) and p110δD910A/D910A mice were subjected to longitudinal carotid injury. At 14 and 30 days after carotid injury, mice with inactive p110δ showed strongly decreased infiltration of inflammatory cells (including T lymphocytes and macrophages) and vascular smooth muscle cells (VSMCs), compared with WT mice. Likewise, PI-3065, a p110δ-selective PI3K inhibitor, almost completely prevented restenosis after artery injury. Our data showed that p110δ PI3K plays a main role in promoting neointimal thickening and inflammatory processes during vascular stenosis, with its inhibition providing significant reduction in restenosis following carotid injury. p110δ-selective inhibitors, recently approved for the treatment of human B-cell malignancies, therefore, present a new therapeutic opportunity to prevent the restenosis upon artery injury.
Assuntos
Lesões das Artérias Carótidas/enzimologia , Estenose das Carótidas/enzimologia , Classe I de Fosfatidilinositol 3-Quinases/imunologia , Inflamação/enzimologia , Animais , Artérias Carótidas/enzimologia , Artérias Carótidas/imunologia , Artérias Carótidas/metabolismo , Artérias Carótidas/patologia , Lesões das Artérias Carótidas/genética , Lesões das Artérias Carótidas/imunologia , Lesões das Artérias Carótidas/patologia , Estenose das Carótidas/genética , Estenose das Carótidas/imunologia , Estenose das Carótidas/patologia , Classe I de Fosfatidilinositol 3-Quinases/genética , Modelos Animais de Doenças , Técnicas de Introdução de Genes , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Masculino , Camundongos Endogâmicos C57BL , Neointima/enzimologia , Neointima/genética , Neointima/imunologia , Neointima/patologia , Mutação PuntualRESUMO
The effects of dynamic exercise on restenosis after vascular injury are still unknown. The consequences of balloon dilation-induced injury on neointimal hyperplasia, vascular negative remodeling, and reendothelialization were assessed in sedentary and trained rats. Ex vivo eNOS vascular expression and activity were investigated in carotid arteries isolated from sedentary and exercised rats. The in vivo effects of eNOS inhibition by L-NMMA on vessel wall after balloon dilation were evaluated in sedentary and exercised rats. We also investigated the effects of exercise on neointimal formation in a rat stent model of vascular injury. Compared with sedentary group, the arteries isolated from trained rats showed higher levels of eNOS protein expression and activity 7 days after balloon dilation. A significant reduction of both neointimal hyperplasia and negative remodeling was observed 14 days after balloon injury in trained compared with sedentary rats. Moreover, we demonstrated that exercise training produced accelerated reendothelialization of the balloon injured arterial segments compared with sedentary. L-NMMA administration eliminated the benefits of physical training on vessel wall after balloon dilation. Finally, a decrease of neointimal hyperplasia as well as of platelet aggregation was observed after stent deployment in trained rats compared with sedentary. In conclusion, physical exercise could favorably affect restenosis after balloon angioplasty and stenting. Increase in eNOS expression and activity might contribute to the potential beneficial effects of exercise on the vessel wall after vascular injury.
Assuntos
Angioplastia com Balão/efeitos adversos , Estenose das Carótidas/prevenção & controle , Oclusão de Enxerto Vascular/prevenção & controle , Óxido Nítrico Sintase/metabolismo , Condicionamento Físico Animal , Difosfato de Adenosina/farmacologia , Animais , Estenose das Carótidas/etiologia , Estenose das Carótidas/patologia , Divisão Celular/fisiologia , Modelos Animais de Doenças , Endotélio Vascular/enzimologia , Endotélio Vascular/lesões , Endotélio Vascular/patologia , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Oclusão de Enxerto Vascular/etiologia , Oclusão de Enxerto Vascular/patologia , Hiperplasia/patologia , Hiperplasia/prevenção & controle , Immunoblotting , Imuno-Histoquímica , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/enzimologia , Músculo Liso Vascular/patologia , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase Tipo III , Esforço Físico , Agregação Plaquetária/efeitos dos fármacos , Ratos , Ratos Wistar , Stents/efeitos adversos , Natação/fisiologia , Túnica Íntima/efeitos dos fármacos , Túnica Íntima/enzimologia , Túnica Íntima/lesões , Túnica Íntima/patologia , ômega-N-Metilarginina/farmacologiaRESUMO
Steroids and growth factors control neuronal development through their receptors under physiological and pathological conditions. We show that PC12 cells harbor endogenous androgen receptor (AR), whose inhibition or silencing strongly interferes with neuritogenesis stimulated by the nonaromatizable synthetic androgen R1881 or NGF. This implies a role for AR not only in androgen signaling, but also in NGF signaling. In turn, a pharmacological TrkA inhibitor interferes with NGF- or androgen-induced neuritogenesis. In addition, androgen or NGF triggers AR association with TrkA, TrkA interaction with PI3-K δ, and downstream activation of PI3-K δ and Rac in PC12 cells. Once associated with AR, filamin A (FlnA) contributes to androgen or NGF neuritogenesis, likely through its interaction with signaling effectors, such as Rac. This study thus identifies a previously unrecognized reciprocal cross-talk between AR and TrkA, which is controlled by ß1 integrin. The contribution of FlnA/AR complex and PI3-K δ to neuronal differentiation by androgens and NGF is also novel. This is the first description of AR function in PC12 cells.