Identification of novel molecular markers in pituitary tumors: contribution to pathogenesis and therapeutic potential

Los adenomas hipofisarios se han considerado comúnmente como una patología tumoral infrecuente debido, posiblemente, a una subestimación en su diagnóstico y consecuentemente, baja incidencia; sin embargo, resultados de autopsias y estudios de imagen revelan que estos tumores presentan una prevalencia total del 14-22% (1), representando la neoplasia intracraneal más frecuente. A pesar de que los adenomas hipofisarios se han clasificado como benignos, estas lesiones pueden ser altamente invasivas e incluso letales debido a la infiltración de estructuras locales o por complicaciones de los efectos metabólicos de los síndromes causados por una sobreproducción hormonal [como es el caso de la acromegalia, resultado de una producción excesiva de hormona del crecimiento (GH), o la enfermedad de Cushing, caracterizada por una sobreexpresión de adrenocorticotropina (ACTH)] (2). El diagnóstico de los adenomas hipofisarios suele ser complicado y engañoso, debido a sus múltiples síntomas clínicos. La mayoría de los adenomas hipofisarios expresan receptores de somatostatina (sst1-5), junto con receptores de dopamina (D 1-5), sobre todo D2 (3). De hecho, estos tumores son sensibles a somatostatina y dopamina, que reducen la secreción hormonal y/o la proliferación celular (4,5). Por lo tanto, agonistas de somatostatina y dopamina, con afinidad por un receptor o por varios recetores, han sido diseñados para el tratamiento farmacológico de estos tumores. Sin embargo, aunque los ssts y Ds estén muy expresados en estos adenomas, y que la eficacia de compuestos dirigidos para sst2, sst5 y D2 ha sido demostrada para el tratamiento de los tumores hipofisarios, existe un considerable grupo de pacientes que responden pobremente, o incluso que son totalmente resistentes, a las terapias con agonistas de ssts y Ds (6). En línea con esto, el objetivo principal de este estudio fue profundizar en el conocimiento de características moleculares y celulares que definan los tumores hipofisarios con el objetivo de mejorar el conocimiento de la pato-fisiología hipofisaria, y de facilitar la identificación de nuevos marcadores para el diagnóstico, pronóstico y herramientas terapéuticas para tratar estas patologías. En concreto, los objetivos de estos estudios fueron: 1) investigar el papel del sistema de la ghrelina, particularmente la nueva, y recientemente identificada, variante de splicing In-1ghrelina (7); 2) estudiar y comparar el efecto de análogos de somatostatina clásicos (octreótido), nuevos (pasireótido) y quiméricos (BIM-23A760) en hipófisis normales y tumorales; 3) explorar el papel funcional de las variantes de splicing, recientemente descubiertas, del receptor de somatostatina 5, sst5TMD4 y sst5TMD5 (8), particularmente sobre características de agresividad de tumores secretores de GH; 4) dilucidad los mecanismos celulares y moleculares involucrados en el test de desmopresina para la enfermedad de Cushing; y finalmente, 5) establecer la relación concreta entre la melatonina y la función hipofisaria normal

E-cadherin expression is associated with somatostatin analogue response in acromegaly

Acromegaly is a rare disease resulting from hypersecretion of growth hormone (GH) and insulin‐like growth factor 1 (IGF1) typically caused by pituitary adenomas, which is associated with increased mortality and morbidity. Somatostatin analogues (SSAs) represent the primary medical therapy for acromegaly and are currently used as first‐line treatment or as second‐line therapy after unsuccessful pituitary surgery. However, a considerable proportion of patients do not adequately respond to SSAs treatment, and therefore, there is an urgent need to identify biomarkers predictors of response to SSAs. The aim of this study was to examine E‐cadherin expression by immunohistochemistry in fifty‐five GH‐producing pituitary tumours and determine the potential association with response to SSAs as well as other clinical and histopathological features. Acromegaly patients with tumours expressing low E‐cadherin levels exhibit a worse response to SSAs. E‐cadherin levels are associated with GH‐producing tumour histological subtypes. Our results indicate that the immunohistochemical detection of E‐cadherin might be useful in categorizing acromegaly patients based on the response to SSAs.ISCIII‐Subdirección General de Evaluación y Fomento de la Investigación PI13/02043 PI16/00175FEDER PI13/02043 PI16/00175Junta de Andalucía A‐0023‐2015 A‐0003‐2016 CTS‐1406 BIO‐0139Andalusian Ministry of Health C‐0015‐2014CIBERobn PI13/ 02043 PI16/0017

Expression of DDX11 and DNM1L at the 12p11 Locus Modulates Systemic Lupus Erythematosus Susceptibility

This study employed genetic and functional analyses using OASIS meta-analysis of multiple existing GWAS and gene-expression datasets to identify novel SLE genes. Methods: Four hundred and ten genes were mapped using SNIPPER to 30 SLE GWAS loci and investigated for expression in three SLE GEO-datasets and the Cordoba GSE50395-dataset. Blood eQTL for significant SNPs in SLE loci and STRING for functional pathways of differentially expressed genes were used. Confirmatory qPCR on SLE monocytes was performed. The entire 12p11 locus was investigated for genetic association using two additional GWAS. Expression of 150 genes at this locus was assessed. Based on this significance, qPCRs for DNM1L and KRAS were performed. Results: Fifty genes were differentially expressed in at least two SLE GEO-datasets, with all probes directionally aligned. DDX11, an RNA helicase involved in genome stability, was downregulated in both GEO and Cordoba datasets. The most significant SNP, rs3741869 in OASIS locus 12p11.21, containing DDX11, was a cis-eQTL regulating DDX11 expression. DDX11 was found repressed. The entire 12p11 locus showed three association peaks. Gene expression in GEO datasets identified DNM1L and KRAS, besides DDX11. Confirmatory qPCR validated DNM1L as an SLE susceptibility gene. DDX11, DNM1L and KRAS interact with each other and multiple known SLE genes including STAT1/STAT4 and major components of IFN-dependent gene expression, and are responsible for signal transduction of cytokines, hormones, and growth-factors, deregulation of which is involved in SLE-development. Conclusion: A genomic convergence approach with OASIS analysis of multiple GWAS and expression datasets identified DDX11 and DNM1L as novel SLE-genes, the expression of which is altered in monocytes from SLE patients. This study lays the foundation for understanding the pathogenic involvement of DDX11 and DNM1L in SLE by identifying them using a systems-biology approach, while the 12p11 locus harboring these genes was previously missed by four independent GWAS

Epigenetic and post-transcriptional regulation of somatostatin receptor subtype 5 (SST5) in pituitary and pancreatic neuroendocrine tumors

Somatostatin receptor subtype 5 (SST5) is an emerging biomarker and actionable target in pituitary (PitNETs) and pancreatic (PanNETs) neuroendocrine tumors. Transcriptional and epigenetic regulation of SSTR5 gene expression and mRNA biogenesis is poorly understood. Recently, an overlapping natural antisense transcript, SSTR5-AS1, potentially regulating SSTR5 expression, was identified. We aimed to elucidate whether epigenetic processes contribute to the regulation of SSTR5 expression in PitNETs (somatotropinomas) and PanNETs. We analyzed the SSTR5/SSTR5-AS1 human locus in silico to identify CpG islands. SSTR5 and SSTR5-AS1 expression was assessed by quantitative real-time PCR (qPCR) in 27 somatotropinomas, 11 normal pituitaries (NPs), and 15 PanNETs/paired adjacent (control) samples. We evaluated methylation grade in four CpG islands in the SSTR5/SSTR5-AS1 genes. Results revealed that SSTR5 and SSTR5-AS1 were directly correlated in NP, somatotropinoma, and PanNET samples. Interestingly, selected CpG islands were differentially methylated in somatotropinomas compared with NPs. In PanNETs cell lines, SSTR5-AS1 silencing downregulated SSTR5 expression, altered aggressiveness features, and influenced pasireotide response. These results provide evidence that SSTR5 expression in PitNETs and PanNETs can be epigenetically regulated by the SSTR5-AS1 antisense transcript and, indirectly, by DNA methylation, which may thereby impact tumor behavior and treatment response.Junta de AndalucíaMinisterio de EconomíaMinisterio de Ciencia e Innovació

Somatostatin Receptor Splicing Variant sst5TMD4 Overexpression in Glioblastoma Is Associated with Poor Survival, Increased Aggressiveness Features and Somatostatin Analogs Resistance

Glioblastoma (GBM) is the most malignant and lethal brain tumor. Current standard treatment consists of surgery followed by radiotherapy/chemotherapy; however, this is only a palliative approach with a mean post-operative survival of scarcely ~12–15 months. Thus, the identification of novel therapeutic targets to treat this devastating pathology is urgently needed. In this context, the truncated splicing variant of the somatostatin receptor subtype 5 (sst5TMD4), which is produced by aberrant alternative splicing, has been demonstrated to be overexpressed and associated with increased aggressiveness features in several tumors. However, the presence, functional role, and associated molecular mechanisms of sst5TMD4 in GBM have not been yet explored. Therefore, we performed a comprehensive analysis to characterize the expression and pathophysiological role of sst5TMD4 in human GBM. sst5TMD4 was significantly overexpressed (at mRNA and protein levels) in human GBM tissue compared to non-tumor (control) brain tissue. Remarkably, sst5TMD4 expression was significantly associated with poor overall survival and recurrent tumors in GBM patients. Moreover, in vitro sst5TMD4 overexpression (by specific plasmid) increased, whereas sst5TMD4 silencing (by specific siRNA) decreased, key malignant features (i.e., proliferation and migration capacity) of GBM cells (U-87 MG/U-118 MG models). Furthermore, sst5TMD4 overexpression in GBM cells altered the activity of multiple key signaling pathways associated with tumor aggressiveness/progression (AKT/JAK-STAT/NF-κB/TGF-β), and its silencing sensitized GBM cells to the antitumor effect of pasireotide (a somatostatin analog). Altogether, these results demonstrate that sst5TMD4 is overexpressed and associated with enhanced malignancy features in human GBMs and reveal its potential utility as a novel diagnostic/prognostic biomarker and putative therapeutic target in GBMs

Presence of sst5TMD4, a truncated splice variant of the somatostatin receptor subtype 5, is associated to features of increased aggressiveness in pancreatic neuroendocrine tumors

Purpose: Gastroenteropancreatic neuroendocrine tumors (GEP-NETs) are rare and heterogeneous tumors, and their biological behavior is not well known. We studied the presence and potential functional roles of somatostatin receptors (sst1-5), focusing particularly on the truncated variants (sst5TMD4, sst5TMD5) and on their relationships with the angiogenic system (Ang/Tie-2 and VEGF) in human GEP-NETs. Experimental Design: We evaluated 42 tumor tissue samples (26 primary/16 metastatic) from 26 patients with GEP-NETs, and 30 non-tumoral tissues (26 from adjacent non-tumor regions and 4 from normal controls) from a single center. Expression of sst1-5, sst5TMD4, sst5TMD5, Ang1-2, Tie-2 and VEGF was analyzed using real-time qPCR, immunofluorescence and immunohistochemistry. Expression levels were associated with tumor characteristics and clinical outcomes. Functional role of sst5TMD4 was analyzed in GEP-NET cell lines. Results: sst1 exhibited the highest expression in GEP-NET, whilst sst2 was the most frequently observed sst-subtype (90.2%). Expression levels of sst1, sst2, sst3, sst5TMD4, and sst5TMD5 were significantly higher in tumor tissues compared to their adjacent non-tumoral tissue. Lymph-node metastases expressed higher levels of sst5TMD4 than in its corresponding primary tumor tissue. sst5TMD4 was also significantly higher in intestinal tumor tissues from patients with residual disease of intestinal origin compared to those with non-residual disease. Functional assays demonstrated that the presence of sst5TMD4 was associated to enhanced malignant features in GEP-NET cells. Angiogenic markers correlated positively with sst5TMD4, which was confirmed by immunohistochemical/fluorescence studies. Conclusions: sst5TMD4 is overexpressed in GEP-NETs and is associated to enhanced aggressiveness, suggesting its potential value as biomarker and target in GEP-NETs.This work has received the following grants: Proyectos de Investigación en Salud (FIS) PI13-01414, and PIE-0041 (funded by Instituto de Salud Carlos III) and S2011/BMD-2328 TIRONET (funded by Comunidad de Madrid) (to MM). BIO-0139, CTS-5051, CTS-1406, PI-0369-2012, BFU2010-19300, BFU2013-43282-R, PI13/00651, CIBERobn and Ayuda Merck Serono 2013 (to RML and JPC). Fellowship CTS-5051 (to AIC). “Sara Borrell” program CD11/00276 (to MDG

Complement component 3 as biomarker of disease activity and cardiometabolic risk factor in rheumatoid arthritis and spondyloarthritis.

Funder: Europeo de Desarrollo Regional de la Unión EuropeaFunder: Rheumatic Diseases NetworkOBJECTIVE: To analyze the relationship between complement component 3 (C3) and the prevalence of cardiometabolic risk factors and disease activity in the rheumatic diseases having the highest rates of cardiovascular morbidity and mortality: rheumatoid arthritis (RA), psoriatic arthritis (PsA) and axial spondyloarthritis (axSpA). METHODS: This is a cross-sectional study including 200 RA, 80 PsA, 150 axSpA patients and 100 healthy donors. The prevalence of cardiometabolic risk factors [obesity, insulin resistance, type 2 diabetes mellitus, hyperlipidemia, apolipoprotein B/apolipoprotein A (apoB/apoA) and atherogenic risks and hypertension] was analyzed. Serum complement C3 levels, inflammatory markers and disease activity were evaluated. Cluster analysis was performed to identify different phenotypes. Receiver operating characteristic (ROC) curve analysis to assess the accuracy of complement C3 as biomarker of insulin resistance and disease activity was carried out. RESULTS: Levels of complement C3, significantly elevated in RA, axSpA and PsA patients, were associated with the prevalence of cardiometabolic risk factors. Hard clustering analysis identified two distinctive phenotypes of patients depending on the complement C3 levels and insulin sensitivity state. Patients from cluster 1, characterized by high levels of complement C3 displayed increased prevalence of cardiometabolic risk factors and high disease activity. ROC curve analysis showed that non-obesity related complement C3 levels allowed to identify insulin resistant patients. CONCLUSIONS: Complement C3 is associated with the concomitant increased prevalence of cardiometabolic risk factors in rheumatoid arthritis and spondyloarthritis. Thus, complement C3 should be considered a useful marker of insulin resistance and disease activity in these rheumatic disorders

Oncogenic Role of Secreted Engrailed Homeobox 2 (EN2) in Prostate Cancer

Engrailed variant-2 (EN2) has been suggested as a potential diagnostic biomarker; however, its presence and functional role in prostate cancer (PCa) cells is still controversial or unknown. Here, we analyzed 1) the expression/secretion profile of EN2 in five independent samples cohorts from PCa patients and controls (prostate tissues and/or urine) to determine its utility as a PCa biomarker; and 2) the functional role of EN2 in normal (RWPE1) and tumor (LNCaP/22Rv1/PC3) prostate cells to explore its potential value as therapeutic target. EN2 was overexpressed in our two cohorts of PCa tissues compared to control and in tumor cell lines compared with normal-like prostate cells. This profile was corroborated in silico in three independent data sets [The Cancer Genome Atlas(TCGA)/Memorial Sloan Kettering Cancer Center (MSKCC)/Grasso]. Consistently, urine EN2 levels were elevated and enabled discrimination between PCa and control patients. EN2 treatment increased cell proliferation in LNCaP/22Rv1/PC3 cells, migration in RWPE1/PC3 cells, and PSA secretion in LNCaP cells. These effects were associated, at least in the androgen-sensitive LNCaP cells, with increased AKT and androgen-receptor phosphorylation levels and with modulation of key cancer-associated genes. Consistently, EN2 treatment also regulated androgen-receptor activity (full-length and splicing variants) in androgen-sensitive 22Rv1 cells. Altogether, this study demonstrates the potential utility of EN2 as a non-invasive diagnostic biomarker for PCa and provides novel and valuable information to further investigate its putative utility to develop new therapeutic tools in PCa

Derechos Humanos de los Grupos Vulnerables. Manual

La formación que ofrecemos en este manual pretende sensibilizar y dar herramientas de detección y reacción frente a las lesiones de derechos, colocando esas herramientas en manos de nuestros alumnos, que por estar cursando educación superior, se encuentran en unas condiciones óptimas para poder asistir, por sus conocimientos y sus capacidades, a quienes son víctimas de situaciones de extrema vulnerabilidad, facilitándoles los medios adecuados para reforzar su capacidad de reacción y reducir, precisamente, su vulnerabilidad.El Manual es producto de la Red de Derechos Humanos en la Educación Superior (RedDHES), un proyecto ALFA financiado por la Unión EuropeaCap. 1. La Convención sobre los Derechos del Niño y la protección de la infancia en la normativa internacional de derechos humanos / Alejandro Morlachetti. - Cap. 2. La protección de los derechos humanos de niñas y niños en el Sistema Interamericano de Derechos Humanos / Karlos Castilla. - Cap. 3. Asegurando el goce de los derechos en condiciones de igualdad: derechos humanos de las personas con discapacidad – contexto general / José Claudio Monteiro de Brito Filho. - Cap. 4. Entre mantenimiento y cambio: un análisis de los primeros años de la Convención de las Naciones Unidas sobre personas con discapacidad / George Rodrigo Bandeira Galindo. - Cap. 5. La incorporación de la discapacidad en el sistema interamericano: principales regulaciones y estándares post-convención / Renata Bregaglio. - Cap. 6. Género y derechos humanos: revolución de ideas y políticas públicas (contexto general) / Lia Zanotta. - Cap. 7. Sistema internacional de protección de los derechos humanos de las mujeres / Yolanda Gómez. - Cap. 8. La construcción de la igualdad de género en el ámbito regional americano / Yanira Zuñiga. - Cap. 9. La protección de los pueblos indígenas en el sistema internacional / Julian Burger. - Cap. 10. Movimientos, pueblos y ciudadanías indígenas: inscripciones constitucionales y derechos étnicos en latinoamerica / Jane Felipe Beltrão y Assis da Costa Oliveira. - Cap. 11. Los derechos de los pueblos indígenas en América Latina: avances jurídicos y brechas de implementación / José Aylwin. - Cap. 12. La jurisprudencia de la Corte Interamericana de Derechos Humanos en materia de pueblos indígenas y tribales / Rosmerlin Estupiñan Silva y Juana María Ibáñez Rivas. - Cap. 13. Derechos humanos y diversidad sexual: contexto general / Fernando Muñoz León. - Cap. 14. La interdicción de discriminación por razón de orientación sexual e identidad sexual en el ámbito internacional / Ascensión Elvira Perales. - Cap. 15. Derechos humanos y diversidad sexual en el Sistema Interamericano de Protección de los Derechos Humanos / Cristina Figueiredo Terezo. - Cap. 16. El Derecho de los «pueblos y naciones indígena originario campesinos» en la Constitución Política del Estado Plurinacional de Bolivia / Nataly Viviana Vargas Gamboa y Shirley Gamboa Alba. - Cap. 17. La protección del desplazamiento forzado por desastres medioambientales en Colombia: hacia una perspectiva de derechos humanos / Sebastián Rubiano Galvi

In1-ghrelin splicing variant is overexpressed in pituitary adenomas and increases their aggressive features

Pituitary adenomas comprise a heterogeneous subset of pathologies causing serious comorbidities, which would benefit from identification of novel, common molecular/cellular biomarkers and therapeutic targets. The ghrelin system has been linked to development of certain endocrine-related cancers. Systematic analysis of the presence and functional implications of some components of the ghrelin system, including native ghrelin, receptors and the recently discovered splicing variant In1-ghrelin, in human normal pituitaries (n 5 11) and pituitary adenomas (n 5 169) revealed that expression pattern of ghrelin system suffers a clear alteration in pituitary adenomasas comparedwith normal pituitary, where In1-ghrelin is markedly overexpressed. Interestingly, in cultured pituitary adenoma cells In1-ghrelin treatment (acylated peptides at 100 nM; 24–72 h) increasedGHandACTHsecretion, Ca21 and ERK1/2 signaling and cell viability, whereas In1-ghrelin silencing (using a specific siRNA; 100 nM) reduced cell viability. These results indicate that an alteration of the ghrelin system, specially its In1-ghrelin variant, could contribute to pathogenesis of different pituitary adenomas types, and suggest that this variant and its related ghrelin system could provide new tools to identify novel, more general diagnostic, prognostic and potential therapeutic targets in pituitary tumors