Caveolina-1 en la progresión metastásica del Sarcoma de Ewing

El sarcoma de Ewing (SE) es el segundo tumor óseo maligno infantil más frecuente y presenta una alta incidencia de enfermedad metastásica. Este tipo de tumores presentan una translocación génica característica que da origen a una proteína de fusión, normalmente EWS/FLI1. Esta proteína de fusión actúa como factor de transcripción aberrante, regulando la expresión de diferentes genes implicados en la iniciación, mantenimiento y progresión del tumor. Nuestro grupo describió como uno de estos genes diana a caveolina 1 (CAV1), describiendo además su papel determinante en el fenotipo maligno del SE, en la tumorigénesis, en la angiogénesis y en la resistencia a apoptosis inducida por quimioterapia. Para investigar el papel concreto de CAV1 en el proceso metastásico de este sarcoma, se creó un modelo de baja expresión de CAV1 en líneas celulares de SE y se determinaron cambios en su capacidad migratoria, invasiva y metastásica. En los ensayos in vitro se halló una menor capacidad migratoria de las células con silenciamiento de CAV1 y una reducción en la expresión de metaloproteinasa 9 (MMP-9) y en la actividad de MMP-2, ambas MMPs implicadas en el proceso de invasión. La regulación de la actividad de MMP-2 parece estar relacionada con la posible regulación que ejerce CAV1 en la función de la MMP de membrana tipo 1 (MT1-MMP), proteína fundamental para la activación de MMP-2. Por otro lado, en este estudio se propone que CAV1 promueve la expresión de MMP-9 transcripcionalmente a través de la regulación de la activación de la vía de señalización de ERK1/2 (Extracellular signal-regulated kinase 1/2). En nuestro modelo, ERK1/2 activo translocaría al núcleo donde activaría los factores de transcripción responsables de la activación del promotor de MMP-9. Por otro lado, la fosforilación de RSK2 (Ribosomal S6 kinase 2) por parte de ERK1/2 en el citoplasma, produciría la activación de RSK2 que a su vez activaría la proteína ribosomal rpS6, uno de los responsables de la iniciación de la traducción, por lo que también podría estar influyendo a la producción de MMP-9 a este nivel. Según nuestros resultados, CAV1 estaría influyendo en la capacidad migratoria de las células de SE mediante dos mecanismos: 1) a través de la activación de la vía de ERK1/2 y 2) mediante la unión a diferentes proteínas con dominio SH2 a través de la fosforilación de CAV1 en la tirosina 14. ERK1/2 influye en la regulación de la capacidad migratoria de una forma de pendiente o independiente de RSK1. Además, en los ensayos de metástasis experimental in vivo las células con inhibición de CAV1 presentaron una menor incidencia de metástasis pulmonar, hecho que correlacionó con una disminución en la expresión de SPARC (Secreted protein acidic and rich in cysteine), una proteína de adhesión importante en procesos metastásicos. En resumen, nuestros resultados evidencian la importancia de CAV1 en el proceso metastásico del SE.Ewing’s sarcoma (ES) is the second most common bone tumor in childhood and occurs with a high incidence of metastatic disease. Such tumors have a characteristic gene translocation that gives rise to a fusion protein, most commonly EWS/FLI1. This fusion protein acts as an aberrant transcription factor regulating the expression of different target genes involved in the initiation, maintenance and progression of the tumor. Our group described caveolin 1 (CAV1) as one of these target genes, describing its role in the malignant phenotype, tumorigenicity and resistance to chemotherapy-induced apoptosis of ES cell lines. To investigate the specific role of CAV1 in the metastatic process of this sarcoma, we established a model of low expression of CAV1 in cell lines of ES. Then, we measured changes in their migratory capacity, invasiveness and metastatic potential. In vitro, we found a lower migratory capability of CAV1 knockdown cells and a reduction in MMP-9 expression and MMP-2 activity. The regulation of MMP-2 activity seems to be related to the possible regulation that CAV1 exerts on the function of MT1- MMP, an essential protein for the activation of MMP-2. On the other hand, we suggest that CAV1 promotes the expression of MMP-9, both transcriptionaly and post-transcriptionaly, through regulating ERK1/2 signaling pathway. In our model, activated ERK1/2 would translocate to the nucleus where it would activate the transcription factors responsible for MMP-9 promoter activation. At the cytoplasm, activated ERK1/2 would phosphorylate and activate RSK2, which, in turn, would promote rpS6 activation, leading to protein translation initiation. Our results indicate that CAV1 is regulating migratory capability of ES cells by two different mechanisms; 1) through ERK1/2 pathway activation, 2) by linking several proteins bearing SH2 domains trough phosphorylated Tyr14 of CAV1. ERK1/2 seems to regulate cell migration in both RSK1-dependent and independent manner. In addition, experimental metastasis assays in vivo showed that, CAV1 knockdown cells had a lower incidence of pulmonary metastasis, a fact that correlated with a decrease in the expression of SPARC, a major adhesion protein in metastatic processes. In summary, our results demonstrate the importance of CAV1 in the metastatic process on ES tumors

Critical appraisal of international guidelines for the management of fecal incontinence in adults: is it possible to define what to do in different clinical scenarios?

Faecal incontinence; Treatment algorithm; Guidelines; ReviewIncontinencia fecal; Algoritmo de tratamiento; Guias; RevisiónIncontinència fecal; Algorisme de tractament; Guies; RevisióFecal incontinence (FI) is a complex often multifactorial functional disorder which is associated with a significant impact on patients’ quality of life. There is a broad spectrum of symptoms, and degrees of severity and diverse patient backgrounds. Several treatment algorithms from different professional societies and experts are available in the literature. However, no consensus has been reached on several aspects of FI management. We performed a critical review of the most recently pub- lished guidelines on FI, emphasising the lack of consensus, highlighting specific topics mentioned in each of the guidelines that are not covered in the others and defining the treatment proposed in different clinical scenarios

Targeted Therapies in Sarcomas: Challenging the Challenge

Sarcomas are a heterogeneous group of mesenchymal malignancies that very often lead to death. Nowadays, chemotherapy is the only available treatment for most sarcomas but there are few active drugs and clinical results still remain very poor. Thus, there is an imperious need to find new therapeutic alternatives in order to improve sarcoma patient's outcome. During the last years, there have been described a number of new molecular pathways that have allowed us to know more about cancer biology and tumorigenesis. Sarcomas are one of the tumors in which more advances have been made. Identification of specific chromosomal translocations, some important pathways characterization such as mTOR pathway or the insulin-like growth factor pathway, the stunning development in angiogenesis knowledge, and brand new agents like viruses have lead to the development of new therapeutic options with promising results. This paper makes an exhaustive review of preclinical and clinical evidence of the most recent targeted therapies in sarcomas and provides a future view of treatments that may lead to improve prognosis of patients affected with this disease

Caveolin-1 in sarcomas: friend or foe?

Sarcomas represent a heterogeneous group of tumors with a complex and difficult reproducible classification. Their pathogenesis is poorly understood and there are few effective treatment options for advanced disease. Caveolin-1 is a multifunctional scaffolding protein with multiple binding partners that regulates multiple cancer-associated processes including cellular transformation, tumor growth, cell death and survival, multidrug resistance, angiogenesis, cell migration and metastasis. However, ambiguous roles have been ascribed to caveolin-1 in signal transduction and cancer, including sarcomas. In particular, evidence indicating that caveolin-1 function is cell context dependent has been repeatedly reported. Caveolin-1 appears to act as a tumor suppressor protein at early stages of cancer progression. In contrast, a growing body of evidence indicates that caveolin-1 is up-regulated in several multidrug-resistant and metastatic cancer cell lines and human tumor specimens. This review is focused on the role of caveolin-1 in several soft tissue and bone sarcomas and discusses the use of this protein as a potential diagnostic and prognostic marker and as a therapeutic target

The importance of being dead: cell death mechanisms assessment in anti-sarcoma therapy

Cell death can occur through different mechanisms, defined by their nature and physiological implications. Correct assessment of cell death is crucial for cancer therapy success. Sarcomas are a large and diverse group of neoplasias from mesenchymal origin. Among cell death types, apoptosis is by far the most studied in sarcomas. Albeit very promising in other fields, regulated necrosis and other cell death circumstances (as so-called "autophagic cell death" or "mitotic catastrophe") have not been yet properly addressed in sarcomas. Cell death is usually quantified in sarcomas by unspecific assays and in most cases the precise sequence of events remains poorly characterized. In this review, our main objective is to put into context the most recent sarcoma cell death findings in the more general landscape of different cell death modalities

EphA2-induced angiogenesis in ewing sarcoma cells works through bFGF production and is dependent on caveolin-1

Angiogenesis is the result of the combined activity of the tumor microenvironment and signaling molecules. The angiogenic switch is represented as an imbalance between pro- and anti-angiogenic factors and is a rate-limiting step in the development of tumors. Eph receptor tyrosine kinases and their membrane-anchored ligands, known as ephrins, constitute the largest receptor tyrosine kinase (RTK) subfamily and are considered a major family of pro-angiogenic RTKs. Ewing sarcoma (EWS) is a highly aggressive bone and soft tissue tumor affecting children and young adults. As other solid tumors, EWS are reliant on a functional vascular network for the delivery of nutrients and oxygen and for the removal of waste. Based on the biological roles of EphA2 in promoting angiogenesis, we explored the functional role of this receptor and its relationship with caveolin-1 (CAV1) in EWS angiogenesis. We demonstrated that lack of CAV1 results in a significant reduction in micro vascular density (MVD) on 3 different in vivo models. In vitro, this phenomenon correlated with inactivation of EphA2 receptor, lack of AKT response and downregulation of bFGF. We also demonstrated that secreted bFGF from EWS cells acted as chemoattractant for endothelial cells. Furthermore, interaction between EphA2 and CAV1 was necessary for the right localization and signaling of the receptor to produce bFGF through AKT and promote migration of endothelial cells. Finally, introduction of a dominant-negative form of EphA2 into EWS cells mostly reproduced the effects occurred by CAV1 silencing, strongly suggesting that the axis EphA2-CAV1 participates in the promotion of endothelial cell migration toward the tumors favoring EWS angiogenesis

Caveolin-1 is down-regulated in alveolar habdomyosarcomas and negatively regulates tumor growth

Rhabdomyosarcoma is the most common soft tissue sarcoma of childhood and adolescence. Despite advances in therapy, patients with histological variant of rhabdomyosarcoma known as alveolar rhabdomyosarcoma (ARMS) have a 5-year survival of less than 30%. Caveolin-1 (CAV1), encoding the structural component of cellular caveolae, is a suggested tumor suppressor gene involved in cell signaling. In the present study we report that compared to other forms of rhabdomyosarcoma (RMS) CAV1 expression is either undetectable or very low in ARMS cell lines and tumor samples. DNA methylation analysis of the promoter region and azacytidine-induced re-expression suggest the involvement of epigenetic mechanisms in the silencing of CAV1. Reintroduction of CAV1 in three of these cell lines impairs their clonogenic capacity and promotes features of muscular differentiation. In vitro, CAV1-expressing cells show high expression of Caveolin-3 (CAV3), a muscular differentiation marker. Blockade of MAPK signaling is also observed. In vivo, CAV1-expressing xenografts show growth delay, features of muscular differentiation and increased cell death. In summary, our results suggest that CAV1 could function as a potent tumor suppressor in ARMS tumors. Inhibition of CAV1 function therefore, could contribute to aberrant cell proliferation, leading to ARMS development

Caveolin-1 promotes Ewing sarcoma metastasis regulating MMP-9 expression through MAPK/ERK pathway

Ewing sarcoma (ES) is a bone and soft tissue sarcoma affecting mostly children and young adults. Caveolin-1 (CAV1) is a well-known target of EWS/FLI1, the main driver of ES, with an oncogenic role in ES. We have previously described how CAV1 is able to induce metastasis in ES via matrix metalloproteinase-9 (MMP-9). In the present study we showed how CAV1 silencing in ES reduced MEK1/2 and ERK1/2 phosphorylation. Accordingly, chemical inhibition of MEK1/2 resulted in reduction in MMP-9 expression and activity that correlated with reduced migration and invasion. IQ Motif Containing GTPase Activating Protein 1 (IQGAP1) silencing reduced MEK1/2 and ERK1/2 phosphorylation and MMP-9 expression. Furthermore, IQGAP1 silenced cells showed a marked decrease in their migratory and invasive capacity. We demonstrated that CAV1 and IQGAP1 localize in close proximity at the cellular edge, thus IQGAP1 could be the connecting node between CAV1 and MEK/ERK in ES metastatic phenotype. Analysis of the phosphorylation profile of CAV1-silenced cells showed a decrease of p-ribosomal protein S6 (RPS6). RPS6 can be phosphorylated by p90 ribosomal S6 kinases (RSK) proteins. CAV1-silenced cells showed reduced levels of p-RSK1 and treatment with U0126 provoked the same effect. Despite not affecting ERK1/2 and RPS6 phosphorylation status neither MMP-9 expression nor activity, RSK1 silencing resulted in a reduced migratory and invasive capacity in vitro and reduced incidence of metastases in vivo in a novel orthotopic model. The present work provides new insights into CAV1-driven metastatic process in ES unveiling novel key nodes

Caveolina-1 en la progresión metastásica del Sarcoma de Ewing

[spa] El sarcoma de Ewing (SE) es el segundo tumor óseo maligno infantil más frecuente y presenta una alta incidencia de enfermedad metastásica. Este tipo de tumores presentan una translocación génica característica que da origen a una proteína de fusión, normalmente EWS/FLI1. Esta proteína de fusión actúa como factor de transcripción aberrante, regulando la expresión de diferentes genes implicados en la iniciación, mantenimiento y progresión del tumor. Nuestro grupo describió como uno de estos genes diana a caveolina 1 (CAV1), describiendo además su papel determinante en el fenotipo maligno del SE, en la tumorigénesis, en la angiogénesis y en la resistencia a apoptosis inducida por quimioterapia. Para investigar el papel concreto de CAV1 en el proceso metastásico de este sarcoma, se creó un modelo de baja expresión de CAV1 en líneas celulares de SE y se determinaron cambios en su capacidad migratoria, invasiva y metastásica. En los ensayos in vitro se halló una menor capacidad migratoria de las células con silenciamiento de CAV1 y una reducción en la expresión de metaloproteinasa 9 (MMP-9) y en la actividad de MMP-2, ambas MMPs implicadas en el proceso de invasión. La regulación de la actividad de MMP-2 parece estar relacionada con la posible regulación que ejerce CAV1 en la función de la MMP de membrana tipo 1 (MT1-MMP), proteína fundamental para la activación de MMP-2. Por otro lado, en este estudio se propone que CAV1 promueve la expresión de MMP-9 transcripcionalmente a través de la regulación de la activación de la vía de señalización de ERK1/2 (Extracellular signal-regulated kinase 1/2). En nuestro modelo, ERK1/2 activo translocaría al núcleo donde activaría los factores de transcripción responsables de la activación del promotor de MMP-9. Por otro lado, la fosforilación de RSK2 (Ribosomal S6 kinase 2) por parte de ERK1/2 en el citoplasma, produciría la activación de RSK2 que a su vez activaría la proteína ribosomal rpS6, uno de los responsables de la iniciación de la traducción, por lo que también podría estar influyendo a la producción de MMP-9 a este nivel. Según nuestros resultados, CAV1 estaría influyendo en la capacidad migratoria de las células de SE mediante dos mecanismos: 1) a través de la activación de la vía de ERK1/2 y 2) mediante la unión a diferentes proteínas con dominio SH2 a través de la fosforilación de CAV1 en la tirosina 14. ERK1/2 influye en la regulación de la capacidad migratoria de una forma de pendiente o independiente de RSK1. Además, en los ensayos de metástasis experimental in vivo las células con inhibición de CAV1 presentaron una menor incidencia de metástasis pulmonar, hecho que correlacionó con una disminución en la expresión de SPARC (Secreted protein acidic and rich in cysteine), una proteína de adhesión importante en procesos metastásicos. En resumen, nuestros resultados evidencian la importancia de CAV1 en el proceso metastásico del SE.[eng] Ewing’s sarcoma (ES) is the second most common bone tumor in childhood and occurs with a high incidence of metastatic disease. Such tumors have a characteristic gene translocation that gives rise to a fusion protein, most commonly EWS/FLI1. This fusion protein acts as an aberrant transcription factor regulating the expression of different target genes involved in the initiation, maintenance and progression of the tumor. Our group described caveolin 1 (CAV1) as one of these target genes, describing its role in the malignant phenotype, tumorigenicity and resistance to chemotherapy-induced apoptosis of ES cell lines. To investigate the specific role of CAV1 in the metastatic process of this sarcoma, we established a model of low expression of CAV1 in cell lines of ES. Then, we measured changes in their migratory capacity, invasiveness and metastatic potential. In vitro, we found a lower migratory capability of CAV1 knockdown cells and a reduction in MMP-9 expression and MMP-2 activity. The regulation of MMP-2 activity seems to be related to the possible regulation that CAV1 exerts on the function of MT1- MMP, an essential protein for the activation of MMP-2. On the other hand, we suggest that CAV1 promotes the expression of MMP-9, both transcriptionaly and post-transcriptionaly, through regulating ERK1/2 signaling pathway. In our model, activated ERK1/2 would translocate to the nucleus where it would activate the transcription factors responsible for MMP-9 promoter activation. At the cytoplasm, activated ERK1/2 would phosphorylate and activate RSK2, which, in turn, would promote rpS6 activation, leading to protein translation initiation. Our results indicate that CAV1 is regulating migratory capability of ES cells by two different mechanisms; 1) through ERK1/2 pathway activation, 2) by linking several proteins bearing SH2 domains trough phosphorylated Tyr14 of CAV1. ERK1/2 seems to regulate cell migration in both RSK1-dependent and independent manner. In addition, experimental metastasis assays in vivo showed that, CAV1 knockdown cells had a lower incidence of pulmonary metastasis, a fact that correlated with a decrease in the expression of SPARC, a major adhesion protein in metastatic processes. In summary, our results demonstrate the importance of CAV1 in the metastatic process on ES tumors