Mecanismos moleculares que regulan la capacidad oncogénica de AIB 1

La señalización celular por receptores nucleares y otros factores de transcripción es un proceso complejo que implica la actuación de múltiples cofactores que regulan la actividad transcripcional. La expresión alterada o la modificación de estos cofactores puede resultar en cambios en la expresión génica, provocar alteraciones en el ciclo celular y dar lugar a patologías diversas, incluyendo el cáncer. Por eso, es muy importante que la cantidad y función de estos moduladores esté bien regulada en las células. AIB1 (Amplificado en cáncer de mama), miembro de la familia de coactivadores de receptores esteroideos, es un coactivador transcripcional no sólo de receptores nucleares, sino también de otros factores de transcripción, incluyendo E2F1, AP-1, NF-κB, etcétera. Un gran número de estudios revela que AIB1 es un oncogén. La amplificación y/o sobreexpresión de AIB1 existe en una gran variedad de tumores humanos. Experimentos en modelos murinos han demostrado que AIB1 es suficiente para iniciar la tumorigénesis. Por el contrario, la deficiencia de AIB1 disminuye la incidencia tumoral y bloquea las metástasis. Los mecanismos por los cuales AIB1 promueve el inicio, la progresión del cáncer y la resistencia endocrina implican diversas vías de señalización, incluyendo la señalización por el receptor de estrógenos, la vía IGF/PI3K/AKT, la vía HER2, y la señalización por NF-κB, E2F1 y Ets. La regulación de la proliferación, supervivencia y migración celular a través de estas vías sugiere un papel central para AIB1 en la tumorigénesis. Desde la identificación de AIB1 se han realizado diversos estudios para investigar su estructura molecular, sus funciones fisiológicas, los mecanismos intermoleculares que regula, y sus efectos patológicos. Sin embargo, aunque se ha progresado mucho en el entendimiento de la función de AIB1 y en cómo influencia la tumorigénesis integrando varios mecanismos moleculares, todavía quedan por aclarar cuestiones importantes referentes a cómo se consigue su sobreexpresión o exceso de proteína. Un número limitado de estudios ha sugerido que el aumento de los niveles de AIB1 en cáncer se puede atribuir tanto a su activación transcripcional como a la estabilización postranscripcional de este coactivador. No obstante, la regulación de la estabilidad proteica representa otro mecanismo importante para incrementar los niveles de esta molécula en cáncer. En la presente tesis se han estudiado algunos de los factores implicados en la regulación de la estabilidad, degradación y actividad de AIB1. La distribución nuclear de este coactivador correlaciona con su menor vida media y facilita su degradación por el sistema ubicuitina-proteasoma. La inhibición de la vía de señalización PI3K/AKT resulta en una mayor inestabilidad para AIB1. Por el contrario, la deleción del motivo PEST, encontrado en la secuencia de este coactivador, conduce a una mayor estabilidad. Por otra parte, se ha identificado una nueva fosforilación de AIB1 específica de mitosis. El complejo Cdk1-ciclina B1 fosforila a AIB1, y esta modificación es revertida durante la progresión a G1 por alguna fosfatasa sensible a ácido okadaico y a caliculina A. La fosforilación mitótica de AIB1 parece no mediar su proteólisis, sino más bien modifica la localización subcelular y la actividad coactivadora de esta molécula. Los datos presentados en este trabajo representan un punto de apoyo para la comprensión de aspectos relacionados con los niveles y actividad del oncogén AIB1. No obstante, es necesaria más investigación en este campo para determinar los mecanismos exactos por los que AIB1 promueve tumorigénesis y para poder establecer a este coactivador como marcador tumoral y como diana terapéutica en la lucha contra el cáncer.Cell signaling comprises a complex process which involves multiple cofactors that regulate transcriptional activity. Altered expression or modification of these cofactors might result in gene expression changes, provoke alterations in cell cycle and lead to several pathologies, including cancer. Therefore, it is essential that amount and function of these molecules are highly regulated. Amplified in breast cancer (AIB1), a member of the steroid receptor coactivator family, is a transcriptional coactivator, not only for nuclear receptors but also for several transcription factors. Compelling evidences reveal AIB1 as a clear oncogen. AIB1 amplification and/or its overexpression occur in many human tumors. Experiments in rodents have demonstrated that AIB1 alone is sufficient to initiate tumorigenesis. However, AIB1 deficiency protects from tumors. The mechanisms of AIB1 to promote cancer initiation, progression, and endocrine resistance involve a variety of signaling pathways. Regulation of cell proliferation, survival and migration through these pathways suggest a central role of AIB1 in tumorigenesis. Although great progress has been made to understand the structure and function of AIB1, the mechanisms which lead to its overexpression are still unknown. A few studies have suggested that increased levels of AIB1 in cancer might be attributable to both transcriptional activation and post-translational stabilization. However, protein stabilization represents another important mechanism. In this work, factors involved in stability regulation, degradation and activity of AIB1 have been studied. Nuclear localization correlates with lower AIB1 half-life and facilitates its degradation through the ubiquitin-proteasome system. PI3K/AKT signaling pathway inhibition results in higher instability for AIB1. However, deletion of the AIB1 PEST motif leads to greater protein stability. A novel mitotic-specific phosphorylation of AIB1 has been identified. Cdk1-cyclin B1 phosphorylates AIB1, and this modification is reverted during mitosis exit by a phosphatase sensitive to okadaic acid and calyculin A. AIB1 mitotic-phosphorylation seems to affect its subcellular distribution and its coactivation activity. Data presented in this work are essential for the comprehension of aspects linked to AIB1 levels and activity. Nonetheless, more research is required for determining the exact mechanisms by which AIB1 promotes tumorigenesis and for establishing AIB1 as a tumoral marker as well as a therapeutic target in cancer

Ultrastructural and molecular analyzes of insulin-producing cells induced from human hepatoma cells.

Background aims. Diabetes type I is an autoimmune disease characterized by the destruction of pancreatic insulin-producing (beta-) cells and resulting in external insulin dependence for life. Islet transplantation represents a potential treatment for diabetes but there is currently a shortage of suitable organs donors. To augment the supply of donors, different strategies are required to provide a potential source of beta-cells. These sources include embryonic and adult stem cells as well as differentiated cell types. The main goal of this study was to induce the transdifferentiation (or conversion of one type cell to another) of human hepatoma cells (HepG2 cells) to insulin-expressing cells based on the exposure of HepG2 cells to an extract of rat insulinoma cells (RIN). Methods. HepG2 cells were fi rst transiently permeabilized with Streptolysin O and then exposed to a cell extract obtained from RIN cells. Following transient exposure to the RIN extract, the HepG2 cells were cultured for 3 weeks. Results. Acquisition of the insulin-producing cell phenotype was determined on the basis of (i) morphologic and (ii) ultrastructural observations, (iii) immunologic detection and (iv) reverse transcription (RT)- polymerase chain reaction (PCR) analysis. Conclusions. This study supports the use of cell extract as a feasible method for achieve transdifferentiation of hepatic cells to insulin-producing cells

miR-146a-5p impairs melanoma resistance to kinase inhibitors by targeting COX2 and regulating NFkB-mediated inflammatory mediators

BACKGROUND: Targeted therapy with BRAF and MEK inhibitors has improved the survival of patients with BRAF-mutated metastatic melanoma, but most patients relapse upon the onset of drug resistance induced by mechanisms including genetic and epigenetic events. Among the epigenetic alterations, microRNA perturbation is associated with the development of kinase inhibitor resistance. Here, we identified and studied the role of miR-146a-5p dysregulation in melanoma drug resistance.METHODS: The miR-146a-5p-regulated NFkB signaling network was identified in drug-resistant cell lines and melanoma tumor samples by expression profiling and knock-in and knock-out studies. A bioinformatic data analysis identified COX2 as a central gene regulated by miR-146a-5p and NFkB. The effects of miR-146a-5p/COX2 manipulation were studied in vitro in cell lines and with 3D cultures of treatment-resistant tumor explants from patients progressing during therapy.RESULTS: miR-146a-5p expression was inversely correlated with drug sensitivity and COX2 expression and was reduced in BRAF and MEK inhibitor-resistant melanoma cells and tissues. Forced miR-146a-5p expression reduced COX2 activity and significantly increased drug sensitivity by hampering prosurvival NFkB signaling, leading to reduced proliferation and enhanced apoptosis. Similar effects were obtained by inhibiting COX2 by celecoxib, a clinically approved COX2 inhibitor.CONCLUSIONS: Deregulation of the miR-146a-5p/COX2 axis occurs in the development of melanoma resistance to targeted drugs in melanoma patients. This finding reveals novel targets for more effective combination treatment. Video Abstract

KRAS-mutant non-small cell lung cancer (NSCLC) therapy based on tepotinib and omeprazole combination

Background KRAS-mutant non-small cell lung cancer (NSCLC) shows a relatively low response rate to chemotherapy, immunotherapy and KRAS-G12C selective inhibitors, leading to short median progression-free survival, and overall survival. The MET receptor tyrosine kinase (c-MET), the cognate receptor of hepatocyte growth factor (HGF), was reported to be overexpressed in KRAS-mutant lung cancer cells leading to tumor-growth in anchorage-independent conditions. Methods Cell viability assay and synergy analysis were carried out in native, sotorasib and trametinib-resistant KRAS-mutant NSCLC cell lines. Colony formation assays and Western blot analysis were also performed. RNA isolation from tumors of KRAS-mutant NSCLC patients was performed and KRAS and MET mRNA expression was determined by real-time RT-qPCR. In vivo studies were conducted in NSCLC (NCI-H358) cell-derived tumor xenograft model. Results Our research has shown promising activity of omeprazole, a V-ATPase-driven proton pump inhibitor with potential anti-cancer properties, in combination with the MET inhibitor tepotinib in KRAS-mutant G12C and non-G12C NSCLC cell lines, as well as in G12C inhibitor (AMG510, sotorasib) and MEK inhibitor (trametinib)-resistant cell lines. Moreover, in a xenograft mouse model, combination of omeprazole plus tepotinib caused tumor growth regression. We observed that the combination of these two drugs downregulates phosphorylation of the glycolytic enzyme enolase 1 (ENO1) and the low-density lipoprotein receptor-related protein (LRP) 5/6 in the H358 KRAS G12C cell line, but not in the H358 sotorasib resistant, indicating that the effect of the combination could be independent of ENO1. In addition, we examined the probability of recurrence-free survival and overall survival in 40 early lung adenocarcinoma patients with KRAS G12C mutation stratified by KRAS and MET mRNA levels. Significant differences were observed in recurrence-free survival according to high levels of KRAS mRNA expression. Hazard ratio (HR) of recurrence-free survival was 7.291 (p = 0.014) for high levels of KRAS mRNA expression and 3.742 (p = 0.052) for high MET mRNA expression. Conclusions We posit that the combination of the V-ATPase inhibitor omeprazole plus tepotinib warrants further assessment in KRAS-mutant G12C and non G12C cell lines, including those resistant to the covalent KRAS G12C inhibitors

Phosphorylation of AIB1 at Mitosis Is Regulated by CDK1/CYCLIN B

Although the AIB1 oncogene has an important role during the early phase of the cell cycle as a coactivator of E2F1, little is known about its function during mitosis.Mitotic cells isolated by nocodazole treatment as well as by shake-off revealed a post-translational modification occurring in AIB1 specifically during mitosis. This modification was sensitive to the treatment with phosphatase, suggesting its modification by phosphorylation. Using specific inhibitors and in vitro kinase assays we demonstrate that AIB1 is phosphorylated on Ser728 and Ser867 by Cdk1/cyclin B at the onset of mitosis and remains phosphorylated until exit from M phase. Differences in the sensitivity to phosphatase inhibitors suggest that PP1 mediates dephosphorylation of AIB1 at the end of mitosis. The phosphorylation of AIB1 during mitosis was not associated with ubiquitylation or degradation, as confirmed by western blotting and flow cytometry analysis. In addition, luciferase reporter assays showed that this phosphorylation did not alter the transcriptional properties of AIB1. Importantly, fluorescence microscopy and sub-cellular fractionation showed that AIB1 phosphorylation correlated with the exclusion from the condensed chromatin, thus preventing access to the promoters of AIB1-dependent genes. Phospho-specific antibodies developed against Ser728 further demonstrated the presence of phosphorylated AIB1 only in mitotic cells where it was localized preferentially in the periphery of the cell.Collectively, our results describe a new mechanism for the regulation of AIB1 during mitosis, whereby phosphorylation of AIB1 by Cdk1 correlates with the subcellular redistribution of AIB1 from a chromatin-associated state in interphase to a more peripheral localization during mitosis. At the exit of mitosis, AIB1 is dephosphorylated, presumably by PP1. This exclusion from chromatin during mitosis may represent a mechanism for governing the transcriptional activity of AIB1

Signature-driven repurposing of Midostaurin for combination with MEK1/2 and KRASG12C inhibitors in lung cancer

Drug combinations are key to circumvent resistance mechanisms compromising response to single anti-cancer targeted therapies. The implementation of combinatorial approaches involving MEK1/2 or KRASG12C inhibitors in the context of KRAS-mutated lung cancers focuses fundamentally on targeting KRAS proximal activators or effectors. However, the antitumor effect is highly determined by compensatory mechanisms arising in defined cell types or tumor subgroups. A potential strategy to find drug combinations targeting a larger fraction of KRAS-mutated lung cancers may capitalize on the common, distal gene expression output elicited by oncogenic KRAS. By integrating a signature-driven drug repurposing approach with a pairwise pharmacological screen, here we show synergistic drug combinations consisting of multi-tyrosine kinase PKC inhibitors together with MEK1/2 or KRASG12C inhibitors. Such combinations elicit a cytotoxic response in both in vitro and in vivo models, which in part involves inhibition of the PKC inhibitor target AURKB. Proteome profiling links dysregulation of MYC expression to the effect of both PKC inhibitor-based drug combinations. Furthermore, MYC overexpression appears as a resistance mechanism to MEK1/2 and KRASG12C inhibitors. Our study provides a rational framework for selecting drugs entering combinatorial strategies and unveils MEK1/2- and KRASG12C-based therapies for lung cancer.M.R. was supported by a fellowship from MICIU (FPU15/00173), R.E. by a donation from Mauge Burgos de la Iglesia’s family. The Proteomics Platforms of Navarrabiomed led by E.S. and J.F.-I. is a member of Proteored, PRB3 and is supported by grant PT17/0019 of the PE I + D + i 2013–2016, funded by ISCIII and ERDF. E.J.-L. was supported by Foundation of Spanish Association Against Cancer (PROYE18012ROSE), by Centro de Investigación Biomédica en Red (CIBERONC; CB16-12-00350), and by Generalitat Valenciana (AICO/2021/333). F.L. was funded by the Gobierno de Navarra (Ref. 34/2021), the Cancer Research Thematic Network of the Instituto de Salud Carlos III (RTICC RD12/0036/0066), PID2021-122638OB-I00 MCIN/AEI/10.13039/501100011033/ FEDER, UE and by FEDER “Una manera de hacer Europa”. I.F. was funded by FIS PI19/00320 and by the Miguel Servet Program CP21/00052. S.V. was supported by Ministerio de Ciencia, Innovación y Universidades, Convocatoria 2019 para incentivar la Incorporación Estable de Doctores (IED2019-001007-I), by FEDER /Ministerio de Ciencia, Innovación y Universidades - Agencia Estatal de Investigación (SAF2017-89944-R and PID2020‐116344‐RB‐100/MCIN/AEI/10.13039/501100011033, by a Leonardo Grant for Researchers and Cultural Creators 2018 from BBVA Foundation, by a seed grant at the I Convocatoria Proyectos Prueba de Concepto from PRB3-Proteored (Institute of Health Carlos III-ISCIII), by Fundació La Marató de TV3 (474/C/2019), and by Foundation of Spanish Association Against Cancer - Strategic Projects 2020 (PROYE20029VICE). M.P.-S. and S.V. were also funded by Fundación Alberto Palatchi. None of the funding sources were involved in the decision to submit the article for publication.Peer reviewe

Mortality and pulmonary complications in patients undergoing surgery with perioperative SARS-CoV-2 infection: an international cohort study

Background: The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on postoperative recovery needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. This study reports 30-day mortality and pulmonary complication rates in patients with perioperative SARS-CoV-2 infection. Methods: This international, multicentre, cohort study at 235 hospitals in 24 countries included all patients undergoing surgery who had SARS-CoV-2 infection confirmed within 7 days before or 30 days after surgery. The primary outcome measure was 30-day postoperative mortality and was assessed in all enrolled patients. The main secondary outcome measure was pulmonary complications, defined as pneumonia, acute respiratory distress syndrome, or unexpected postoperative ventilation. Findings: This analysis includes 1128 patients who had surgery between Jan 1 and March 31, 2020, of whom 835 (74·0%) had emergency surgery and 280 (24·8%) had elective surgery. SARS-CoV-2 infection was confirmed preoperatively in 294 (26·1%) patients. 30-day mortality was 23·8% (268 of 1128). Pulmonary complications occurred in 577 (51·2%) of 1128 patients; 30-day mortality in these patients was 38·0% (219 of 577), accounting for 81·7% (219 of 268) of all deaths. In adjusted analyses, 30-day mortality was associated with male sex (odds ratio 1·75 [95% CI 1·28–2·40], p\textless0·0001), age 70 years or older versus younger than 70 years (2·30 [1·65–3·22], p\textless0·0001), American Society of Anesthesiologists grades 3–5 versus grades 1–2 (2·35 [1·57–3·53], p\textless0·0001), malignant versus benign or obstetric diagnosis (1·55 [1·01–2·39], p=0·046), emergency versus elective surgery (1·67 [1·06–2·63], p=0·026), and major versus minor surgery (1·52 [1·01–2·31], p=0·047). Interpretation: Postoperative pulmonary complications occur in half of patients with perioperative SARS-CoV-2 infection and are associated with high mortality. Thresholds for surgery during the COVID-19 pandemic should be higher than during normal practice, particularly in men aged 70 years and older. Consideration should be given for postponing non-urgent procedures and promoting non-operative treatment to delay or avoid the need for surgery. Funding: National Institute for Health Research (NIHR), Association of Coloproctology of Great Britain and Ireland, Bowel and Cancer Research, Bowel Disease Research Foundation, Association of Upper Gastrointestinal Surgeons, British Association of Surgical Oncology, British Gynaecological Cancer Society, European Society of Coloproctology, NIHR Academy, Sarcoma UK, Vascular Society for Great Britain and Ireland, and Yorkshire Cancer Research

Proyecto, investigación e innovación en urbanismo, arquitectura y diseño industrial

Actas de congresoLas VII Jornadas de Investigación “Encuentro y Reflexión” y I Jornadas de Investigación de becarios y doctorandos. Proyecto, investigación e innovación en Urbanismo, Arquitectura y Diseño Industrial se centraron en cuatro ejes: el proyecto; la dimensión tecnológica y la gestión; la dimensión social y cultural y la enseñanza en Arquitectura, Urbanismo y Diseño Industrial, sustentados en las líneas prioritarias de investigación definidas epistemológicamente en el Consejo Asesor de Ciencia y Tecnología de esta Universidad Nacional de Córdoba. Con el objetivo de afianzar continuidad, formación y transferencia de métodos, metodología y recursos se incorporó becarios y doctorandos de los Institutos de investigación. La Comisión Honoraria la integraron las tres Secretarias de Investigación de la Facultad, arquitectas Marta Polo, quien fundó y María del Carmen Franchello y Nora Gutiérrez Crespo quienes continuaron la tradición de la buena práctica del debate en la cotidianeidad de la propia Facultad. Los textos que conforman las VII Jornadas son los avances y resultados de las investigaciones realizadas en el bienio 2016-2018.Fil: Novello, María Alejandra. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Repiso, Luciana. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Mir, Guillermo. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Brizuela, Natalia. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Herrera, Fernanda. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Períes, Lucas. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Romo, Claudia. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Gordillo, Natalia. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; ArgentinaFil: Andrade, Elena Beatriz. Universidad Nacional de Córdoba. Facultad de Arquitectura, Urbanismo y Diseño; Argentin

Humanización de la Sanidad y Salud Laboral: Implicaciones, estado de la cuestión y propuesta del Proyecto HU-CI

Resumen La evolución científica y técnica de la atención sanitaria mantienen un desarrollo exponencial que permite cada vez una mejor asistencia a los pacientes, nuevas formas de tratamiento y crecientes índices de supervivencia. Sin embargo, estos importantes avances que enriquecen la atención a la salud, en muchas ocasiones, han ido invadiendo espacio al propio acto humano de la atención a la persona. Potenciar organizaciones más humanizadas implicará establecer acciones dirigidas al ambiente y lugar de trabajo, así como a todos los agentes implicados (pacientes, familiares y profesionales). Con dicha finalidad, las políticas de Humanización y su aplicación al contexto sanitario son una realidad en nuestros días y están concretándose en algunas propuestas específicas en determinadas organizaciones, con el apoyo de algunos organismos oficiales. En el presente trabajo, señalamos la interacción entre los objetivos de dichas políticas de humanización y las metas de la salud laboral en contextos asistenciales. Se exploran áreas comunes de trabajo dirigidas al bienestar del profesional sanitario, posibilidades de gestión integral de ambos conceptos, y se expone un ejemplo concreto como es el Proyecto de Humanización de las Unidades de Cuidados Intensivos (HU-CI)