Hypoxia in the Initiation and Progression of Neuroblastoma Tumours

Neuroblastoma is the most frequent extracranial solid tumour in children, causing 10% of all paediatric oncology deaths. It arises in the embryonic neural crest due to an uncontrolled behaviour of sympathetic nervous system progenitors, giving rise to heterogeneous tumours. Low local or systemic tissue oxygen concentration has emerged as a cellular stimulus with important consequences for tumour initiation, evolution and progression. In neuroblastoma, several evidences point towards a role of hypoxia in tumour initiation during development, tumour cell differentiation, survival and metastatic spreading. However, the heterogeneous nature of the disease, its developmental origin and the lack of suitable experimental models have complicated a clear understanding of the effect of hypoxia in neuroblastoma tumour progression and the molecular mechanisms implicated. In this review, we have compiled available evidences to try to shed light onto this important field. In particular, we explore the effect of hypoxia in neuroblastoma cell transformation and differentiation. We also discuss the experimental models available and the emerging alternatives to study this problem, and we present hypoxia-related therapeutic avenues being explored in the field.Spanish Ministry of Science and Innovation SAF program (grant number SAF2016-80412-P

Trimeric G proteins modulate the dynamic interaction of PKAII with the Golgi complex

The Golgi complex represents a major subcellular location of protein kinase A (PKA) concentration in mammalian cells where it has been previously shown to be involved in vesicle-mediated protein transport processes. We have studied the factors that influence the interaction of PKA typeII subunits with the Golgi complex. In addition to the cytosol, both the catalytic (Cα) and regulatory (RIIα) subunits of PKAII were detected at both sides of the Golgi stack, particularly in elements of the cis- and trans-Golgi networks. PKAII subunits, in contrast, were practically absent from the middle Golgi cisternae. Cell treatment with either brefeldin A, AlF4 − or at low temperature induced PKAII dissociation from the Golgi complex and redistribution to the cytosol. This suggested the existence of a cycle of association/dissociation of PKAII holoenzyme to the Golgi. The interaction of purified RIIα with Golgi membranes was studied in vitro and found not to be affected by brefeldin A while it was sensitive to modulators of heterotrimeric G proteins such as AlF4 −, GTPγS, βγ subunits and mastoparan. RIIα binding was stimulated by recombinant, myristoylated Gαi3 subunit and inhibited by cAMP. Pretreatment of Golgi membranes with bacterial toxins known to catalyze ADP-ribosylation of selected Gα subunits also modified RIIα binding. Taken together the data support a regulatory role for Golgi-associated Gα proteins in PKAII recruitment from the cytosol

CD44-high neural crest stem-like cells are associated with tumour aggressiveness and poor survival in neuroblastoma tumours

BACKGROUND: Neuroblastoma is a paediatric tumour originated from sympathoadrenal precursors and characterized by its heterogeneity and poor outcome in advanced stages. Intra-tumoral cellular heterogeneity has emerged as an important feature in neuroblastoma, with a potential major impact on tumour aggressiveness and response to therapy. CD44 is an adhesion protein involved in tumour progression, metastasis and stemness in different cancers; however, there has been controversies about the significance of CD44 expression in neuroblastoma and its relationship with tumour progression. METHODS: We have performed transcriptomic analysis on patient tumour samples studying the outcome of patients with high CD44 expression. Adhesion, invasion and proliferation assays were performed in sorted CD44high neuroblastoma cells. Tumoursphere cultures have been used to enrich in undifferentiated stem-like cells and to asses self-renewal and differentiation potential. We have finally performed in vivo tumorigenic assays on cell line-derived or Patient-derived xenografts. FINDINGS: We show that high CD44 expression is associated with low survival in high-grade human neuroblastoma, independently of MYCN amplification. CD44 is expressed in a cell population with neural crest stem-like features, and with the capacity to generate multipotent, undifferentiated tumourspheres in culture. These cells are more invasive and proliferative in vitro. CD44 positive cells obtained from tumours are more tumorigenic and metastatic, giving rise to aggressive neuroblastic tumours at high frequency upon transplantation. INTERPRETATION: We describe an unexpected intra-tumoural heterogeneity within cellular entities expressing CD44 in neuroblastoma, and propose that CD44 has a role in neural crest stem-like undifferentiated cells, which can contribute to tumorigenesis and malignancy in this type of cancer. FUNDING: Research supported by grants from the "Asociación Española contra el Cáncer" (AECC), the Spanish Ministry of Science and Innovation SAF program (SAF2016-80412-P), and the European Research Council (ERC Starting Grant to RP).Spanish Ministry of Science and Innovation SAF program (SAF2016-80412-P

A protocol to enrich in undifferentiated cells from neuroblastoma tumor tissue samples and cell lines

The existence of a subpopulation of undifferentiated cells with stem-like properties has been suggested in neuroblastoma tumors, but a definitive biomarker for their successful isolation is missing. Here we describe an in vitro culture system for the enrichment in undifferentiated stem-like tumor cells for subsequent functional assays. We make use of clonal non-adherent cell culture conditions together with cell sorting with specific expression markers. This protocol allows for the differential study of heterogeneous cell population in neuroblastoma tumors. For complete details on the use and execution of this protocol, please refer to Vega et al. (2019)

Oncogenic Sox2 regulates and cooperates with VRK1 in cell cycle progression and differentiation.

Sox2 is a pluripotency transcription factor that as an oncogene can also regulate cell proliferation. Therefore, genes implicated in several different aspects of cell proliferation, such as the VRK1 chromatin-kinase, are candidates to be targets of Sox2. Sox 2 and VRK1 colocalize in nuclei of proliferating cells forming a stable complex. Sox2 knockdown abrogates VRK1 gene expression. Depletion of either Sox2 or VRK1 caused a reduction of cell proliferation. Sox2 up-regulates VRK1 expression and both proteins cooperate in the activation of CCND1. The accumulation of VRK1 protein downregulates SOX2 expression and both proteins are lost in terminally differentiated cells. Induction of neural differentiation with retinoic acid resulted in downregulation of Sox2 and VRK1 that inversely correlated with the expression of differentiation markers such as N-cadherin, Pax6, mH2A1.2 and mH2A2. Differentiation-associated macro histones mH2A1.2and mH2A2 inhibit CCND1 and VRK1 expression and also block the activation of the VRK1 promoter by Sox2. VRK1 is a downstream target of Sox2 and both form an autoregulatory loop in epithelial cell differentiation.Ministerio de Economía y Competitividad [SAF2013-44810R, SAF2014-57791-REDC].Consejería de Educación de la Junta de Castilla y León [CSI002U14 and UIC-017] to P.A.L

A triple action CDK4/6-PI3K-BET inhibitor with augmented cancer cell cytotoxicity

National Institutes of Health GM125195, GM135671, CA192656, FD00511

1,3-diphenylpropan-1-ones as allosteric modulators of α7 nACh receptors with analgesic and antioxidant properties

Nicotine acethylcholine receptors (nAChRs) play critical roles in cognitive processes, neuroprotection and inflammation. Results: According to their substituents, 1,3-diphenylpropan-1-one derivatives act as α7 nAChRs negative allosteric modulators (NAM, OMe) or Type I positive allosteric modulators (PAMs, OH). Compounds 7 and 31 were the most effective (989 and 666% enhancement of ACh-induced currents) and potent (EC: 12.9 and 6.85 μM) PAMs. They exhibited strong radical scavenging values. Compound 31, selective over other neuronal nAChR subtypes and with acceptable pharmacokinetic profile, showed antinociceptive effects in a model of inflammatory pain. Conclusion: Compound 31 is a novel, potent and selective α7 nAChR PAM, displaying antioxidant and analgesic activities. The 1,3-diphenylpropan-1-one scaffold could be the base toward more advanced type I PAMs for the treatment of nAChR-mediated diseases.This work was supported by the Spanish MINECO: CSD2008-00005, The Spanish Ion Channel Initiative-CONSOLIDER INGENIO 2010, SAF2011-22802 and BFU2012-39092-C02. The Instituto de Neurociencias is a “Centre of Excellence Severo Ochoa”. We thank Susana Cámara Garrido for her assistance in the synthesis of some starting compounds and Susana Gerber for technical assistance. BBP thanks the CSIC for a predoctoral fellowship (JAE-Predoc from Junta para la Ampliación de Estudios, co-financed by FSE). Alpha7 nAChRPeer Reviewe

Tenemos cita con el arte: a pilot project of visits and workshops with people affected by Alzheimer's disease in the Prado Museum, the Centro de Arte Reina Sofía Museum and the Faculty of Fine Arts at the Complutense University of Madrid

GIMUPAI is a research group comprising teachers and researchers from the Faculty of Fine Arts (University Complutense of Madrid) and the Department of Social Psychology and Anthropology (Salamanca University) who have been working in art and health projects over the last thirteen years. Recently, we have carried out Tenemos cita con el arte, part of a national research project entitled "Art education in museums and other cultural institutions as a tool for increasing the wellbeing of people affected with Alzheimer" (Ministry of the Economy and Competitiveness-EDU2013-43253-R). The main objective of the program is to make the museum‘s artworks available to people with Alzheimer‘s and their caregivers, at the same time encouraging them to participate in artistic activities and artistic creation through art workshops. Tenemos cita con el arte has been designed as a program of visits of the Prado Museum and the Centro de Arte Reina Sofia Museum. The program also has included participation in workshops on visual arts and artistic creativity in the Faculty of Fine Arts. The program was undertaken between October and December 2015 with a group of 15 participants (Alzheimer‘s patients, caregivers, and other health and social workers)