Wnt, notch, and TGF-β pathways impinge on hedgehog signaling complexity: an open window on cancer

Constitutive activation of the Hedgehog (Hh) signaling pathway is associated with increased risk of developing several malignancies. The biological and pathogenic importance of Hh signaling emphasizes the need to control its action tightly, both physiologically and therapeutically. Evidence of crosstalk between Hh and other signaling pathways is reported in many tumor types. Here, we provide an overview of the current knowledge about the communication between Hh and major signaling pathways, such as Notch, Wnt, and transforming growth factor beta (TGF-beta), which play critical roles in both embryonic and adult life. When these pathways are unbalanced, impaired crosstalk contributes to disease development. It is reported that more than one of these pathways are active in different type of tumors, at the same time. Therefore, starting from a plethora of stimuli that activate multiple signaling pathways, we describe the signals that preferentially converge on the Hh signaling cascade that influence its activity. Moreover, we highlight several connection points between Hh and Notch, Wnt, or TGF-beta pathways, showing a reciprocal synergism that contributes to tumorigenesis, supporting a more malignant behavior by tumor cells, such as in leukemia and brain tumors. Understanding the importance of these molecular interlinking networks will provide a rational basis for combined anticancer drug development

Optimización de la extracción de compuestos bioactivos a partir de pétalos de rosas

El presente trabajo tuvo como objetivo la optimización de las condiciones de extracción para maximizar la obtención de compuestos polifenólicos y con actividad antioxidante a partir de pétalos de rosas. Éstos se obtuvieron de viveros de la región de San Pedro, Provincia de Buenos Aires, seleccionándose para el estudio los cultivares Malu (M), de color rosa y Lovely Red (LR), de color rojo. Con el fin de evaluar distintas condiciones de extracción se aplicó un diseño experimental de superficie de respuesta (Diseño de Box-Behnken) en dos bloques, con tres factores experimentales a dos niveles y un punto central realizando un total de 30 experimentos por cultivar de rosa. Las variables y los niveles fueron: solvente de extracción (agua/etanol); temperatura (30, 90 °C) y tiempo (30, 120 min). Los pétalos fueron deshidrataron por liofilización, molidos y tamizados. El contenido de polifenoles totales en los extractos se determinó por el método colorimétrico de Folin-Ciocalteu expresando el resultado en mg de ácido gálico (AG) / g muestra seca (ms). La evaluación de la actividad antioxidante se realizó a través de la reducción del radical 2,2-difenil-1-picrilhidracilo (DPPH) y los resultados se expresaron en mg de trolox / g ms. Todas las extracciones se realizaron por duplicado y las determinaciones por triplicado. Se aplicó el diseño experimental a los parámetros obtenidos y se calcularon las ecuaciones de los modelos predictivos. Se observó que el solvente fue la variable que afectó en forma más significativa la respuesta, presentando valores máximos en el rango cercano al nivel medio, agua/etanol 50:50, tanto para el contenido de fenoles totales como para la actividad antioxidante. Se observó además, que el aumento de temperatura maximizó la extracción de los compuestos analizados. El tiempo no ejerció un efecto significativo. Los valores más elevados para las condiciones de extracción ensayadas fueron: solvente agua/etanol 50:50 y temperatura 90 °C para cualquiera de los tiempos de extracción ensayados. El contenido de polifenoles obtenido fue de 202 mg AG / g ms para LR y 141,3 mg AG / g ms para M. En el caso de la actividad antioxidante, los valores para estas mismas condiciones fueron 544,6 mg trolox / g ms para la variedad roja y 339,6 mg trolox / g ms para la de color rosa. Además, se observó un muy buen ajuste entre los valores experimentales y los predichos por el modelo. Es de destacar que el contenido de polifenoles totales mostró una muy buena correlación con la actividad antioxidante, siendo los coeficientes de 2 determinación (R2) 0,98 (M) y 0,96 (LR). Los resultados comentados permiten concluir que los pétalos ensayados son una matriz rica en polifenoles y en compuestos con actividad antioxidante pudiéndose maximizar la extracción de dichos compuestos mediante la adecuada selección del solvente.EEA San PedroFil: Baibuch, Sabrina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Industrias. Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Universidad de Buenos Aires. Instituto de Tecnologia de Alimentos y Procesos Quimicos (ITAPROQ); Argentina.Fil: Zema, Paula. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento Química Orgánica; ArgentinaFil: Gabilondo, Julieta. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria San Pedro; ArgentinaFil: Malec, Laura. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento Química Orgánica; ArgentinaFil: Campos, Carmen. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Industrias. Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Universidad de Buenos Aires. Instituto de Tecnología de Alimentos y Procesos Químicos (ITAPROQ); Argentin

Maml1 acts cooperatively with Gli proteins to regulate Sonic hedgheog signaling pathway

Sonic hedgehog (Shh) signaling is essential for proliferation of cerebellar granule cell progenitors (GCPs) and its misregulation is linked to various disorders, including cerebellar cancer medulloblastoma. The effects of Shh pathway are mediated by the Gli family of transcription factors, which controls the expression of a number of target genes, including Gli1. Here, we identify Mastermind-like 1 (Maml1) as a novel regulator of the Shh signaling since it interacts with Gli proteins, working as a potent transcriptional coactivator. Notably, Maml1 silencing results in a significant reduction of Gli target genes expression, with a negative impact on cell growth of NIH3T3 and Patched1−/− mouse embryonic fibroblasts (MEFs), bearing a constitutively active Shh signaling. Remarkably, Shh pathway activity results severely compromised both in MEFs and GCPs deriving from Maml1−/− mice with an impairment of GCPs proliferation and cerebellum development. Therefore Maml1−/− phenotype mimics aspects of Shh pathway deficiency, suggesting an intrinsic requirement for Maml1 in cerebellum development. The present study shows a new role for Maml1 as a component of Shh signaling, which plays a crucial role in both development and tumorigenesis

Notch signalling in development and disease: Maml1 and Jagged1 not always on the shadow of Notch

The Notch signalling is an evolutionary conserved pathway from flies to vertebrates, that regulates a large spectrum of cellular processes, such as proliferation, differentiation and cell death. The intracellular domain of Notch receptors controls the transcription of the target genes through the presence of different factors such as ligands and transcriptional co-activators, whose presence permits the activation of the pathway. Moreover, different studies have highlighted the importance of Notch pathway components, that play a fundamental role in development and the onset of several diseases, often with Notch-independent mechanisms. In this thesis, I report novel observations about Maml1 and Jagged1 proteins directly involved in development/differentiation processes and neoplastic transformation in a Notch-independent manner. In mammals, Maml1 belongs to a family of proteins, also including Maml2 and Maml3, which act as transcriptional coactivators for Notch signalling. Maml1 has been recently shown to act as a coactivator in other cell signalling pathways. Of note, we have demonstrated that Maml1 empowers Sonic Hedgehog signalling pathway, regulating the transcriptional activity of Gli proteins, via a novel Notch-independent mechanism. So far, scientific research on Maml1 has been generally focused on its activity as a transcriptional coactivator, while overlooking its role in the post-transcriptional regulation. Interestingly, our preliminary data suggest a novel role for Maml1 in the post- translational regulation of Gli1, being able to prevent its degradation mediated by Itch, an E3 ubiquitin-protein ligase. Therefore, we propose a dual role for Maml1, both as transcriptional co-activator of important transcriptional factors and post-translational regulator of target proteins, regulating Itch activity directly. This study tries to shed light on the molecular mechanism that regulates the stability and activity of Gli1 mediated by Maml1 and seeks to provide a new integrated level of regulation in Shh/Gli pathway, that might lead to future therapeutic approaches directed against Shh-driven tumours. Moreover, several evidences reported that, similarly to Notch receptors, the DSL ligands undergo the same proteolytic cleavages that result in the release of an intracellular fragment. Jagged1 is a substrate for catalytic activity of the Metalloprotease ADAM17 (A Disintegrin and Metalloproteinase) and presenilin/γ-secretase complex activity that releases a soluble intracellular fragment (Jag1-ICD), which translocates into the nucleus. Recently, several studies linked the expression levels of Jagged1 with the development and/or progression of solid tumours. Jagged1 role in Colorectal Cancer (CRC) is linked to the development and progression of the tumour. CRC is characterized by well-known genetic defects and about 50% of the cases harbour oncogenic RAS mutations. Herein, we demonstrate that Jagged1 is constitutively processed in CRC tumours with mutant Kras, ultimately triggering an intrinsic reverse signalling via its nuclear-targeted intracellular domain (Jag1-ICD). We provide evidence that the processing occurs when a Kras/Erk/ADAM17 signalling axis is switched on, demonstrating that Jagged1 is a novel target of Kras signalling pathway. Notably, we show that Jag1-ICD promotes tumour growth and epithelial-mesenchymal transition, enhancing CRC progression and chemoresistance both in vitro and in vivo. Our data pinpoint a novel role for Jagged1 in CRC tumour biology that may go beyond its effect on canonical Notch activation and suggest that Jag1-ICD may behave as a novel oncogenic driver, able to sustain tumour pathogenesis and to confer chemoresistance, through a non-canonical mechanism. By unveiling the Kras/Erk/ADAM17/Jagged1 signalling axis, we provide new mechanistic insights on CRC tumour biology and highlight a novel attractive target for CRC therapy

5FU/Oxaliplatin-induced Jagged1 cleavage counteracts apoptosis induction in colorectal cancer: a novel mechanism of intrinsic drug resistance

Colorectal cancer (CRC) is characterized by early metastasis, resistance to anti-cancer therapy, and high mortality rate. Despite considerable progress in the development of new treatment options that improved survival benefits in patients with early-stage or advanced CRC, many patients relapse due to the activation of intrinsic or acquired chemoresistance mechanisms. Recently, we reported novel findings about the role of Jagged1 in CRC tumors with Kras signatures. We showed that Jagged1 is a novel proteolytic target of Kras signaling, which induces Jagged1 processing/activation resulting in Jag1-ICD release, which favors tumor development in vivo, through a non-canonical mechanism. Herein, we demonstrate that OXP and 5FU cause a strong accumulation of Jag1-ICD oncogene, through ERK1/2 activation, unveiling a surviving subpopulation with an enforced Jag1-ICD expression, presenting the ability to counteract OXP/5FU-induced apoptosis. Remarkably, we also clarify the clinical ineffectiveness of gamma-secretase inhibitors (GSIs) in metastatic CRC (mCRC) patients. Indeed, we show that GSI compounds trigger Jag1-ICD release, which promotes cellular growth and EMT processes, functioning as tumor-promoting agents in CRC cells overexpressing Jagged1. We finally demonstrate that Jagged1 silencing in OXP- or 5FU-resistant subpopulations is enough to restore the sensitivity to chemotherapy, confirming that drug sensitivity/resistance is Jag1-ICD-dependent, suggesting Jagged1 as a molecular predictive marker for the outcome of chemotherapy

Kras/ADAM17-dependent Jag1-ICD reverse signalling sustains CRC progression and chemoresistance

Colorectal cancer (CRC) is characterized by well-known genetic defects and about 50% of the cases harbour oncogenic RAS mutations. Increased expression of Notch-ligand Jagged1 occurs in several human malignancies, including CRC, and correlates with cancer progression, poor prognosis and recurrence. Herein, we demonstrate that Jagged1 is constitutively processed in CRC tumours with mutant Kras, ultimately triggering an intrinsic reverse signalling via its nuclear-targeted intracellular domain (Jag1-ICD). We provide evidence that the processing occurs when a Kras/Erk/ADAM17 signalling axis is switched on, demonstrating that Jagged1 is a novel target of Kras signalling pathway. Notably, we show that Jag1-ICD promotes tumour growth and epithelial mesenchymal transition, enhancing CRC progression and chemoresistance both in vitro and in vivo. Our data pinpoint a novel role for Jagged1 in CRC tumour biology that may go beyond its effect on canonical Notch activation and suggest that Jag1-ICD may behave as a novel oncogenic driver, able to sustain tumour pathogenesis and to confer chemoresistance, through a non-canonical mechanism. By unveiling the Kras/Erk/ADAM17/Jagged1 signalling axis, we provide new mechanistic insights on CRC tumour biology and highlight a novel attractive target for CRC therapy