18 results on '"Álvaro Otero"'
Search Results
2. SARS-CoV-2 B.1.1.7 Decline Is Not Driven by the Introduction of a More Successful Variant
- Author
-
Pilar Catalán, Luis Alcalá, Darío García de Viedma, Sergio Buenestado-Serrano, Víctor Manuel de la Cueva García, Carmen Losada, Cristina Rodríguez-Grande, Pedro J Sola-Campoy, Javier Adán-Jiménez, Patricia Muñoz, Laura Pérez-Lago, Álvaro Otero-Sobrino, Andrea Molero-Salinas, Carla Rico-Luna, and Jorge Rodríguez-Grande
- Subjects
Microbiology (medical) ,Delta ,2019-20 coronavirus outbreak ,Coronavirus disease 2019 (COVID-19) ,Physiology ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,Zoology ,Alpha (ethology) ,Observation ,Biology ,variants of concern ,Microbiology ,Genetics ,Humans ,Phylogeny ,Whole Genome Sequencing ,General Immunology and Microbiology ,Ecology ,SARS-CoV-2 ,COVID-19 ,Genomics ,Cell Biology ,QR1-502 ,Infectious Diseases ,Spain ,Mutation - Abstract
The SARS-CoV-2 variant of concern (VOC) Delta (B.617.2 lineage) displaced the predominant VOC Alpha (B.1.1.7 lineage) in the United Kingdom. In Madrid, recent start of the decline of predominant VOC Alpha suggested an equivalent phenomenon. However, 11 different variants, none overrepresented in frequency, occupied progressively over a period of 7 weeks the niche previously dominated by VOC Alpha. Only after these 7 weeks, VOC Delta started to emerge. Viral competition due to the entry of VOC Delta is not the major force driving the start of VOC Alpha decline in Madrid. IMPORTANCE Our data indicate that the dynamics of SARS-CoV-2 VOCs turnover in our setting differ from those proposed for other countries. A systematic genomic analysis, updated on a weekly basis, of representative randomly selected samples of SARS-CoV-2 circulating variants allowed us to define a lapse of 7 weeks between the start of VOC Alpha decline and the final emergence of VOC Delta. During this period, VOC Alpha showed a sustained decline, while 11 VOCs, variants of interest (VOIs), and other identified variants, none overrepresented, occupied the niche left by VOC Alpha. Only after these 7 weeks, emergence of VOC Delta occurred, indicating that viral competition involving VOC Delta was not the exclusive direct driving force behind the starting of VOC Alpha decline.
- Published
- 2021
3. Shared Mutations in Emerging SARS-CoV-2 Circulating Variants May Lead to Reverse Transcription-PCR (RT-PCR)-Based Misidentification of B.1.351 and P.1 Variants of Concern
- Author
-
Darío García de Viedma, Laura Pérez-Lago, Carla Rico-Luna, Ana Candela, Pilar Catalán, Álvaro Otero-Sobrino, Patricia Muñoz, Andrea Molero-Salinas, Carmen Losada, and Sergio Buenestado-Serrano
- Subjects
Microbiology (medical) ,2019-20 coronavirus outbreak ,Coronavirus disease 2019 (COVID-19) ,Physiology ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,RT-PCR ,Observation ,Genome, Viral ,Biology ,Microbiology ,reverse transcriptase PCR ,Genetics ,Humans ,Diagnostic Errors ,Whole genome sequencing ,General Immunology and Microbiology ,Ecology ,Whole Genome Sequencing ,Reverse Transcriptase Polymerase Chain Reaction ,SARS-CoV-2 ,VOC ,COVID-19 ,Cell Biology ,QR1-502 ,Reverse transcription polymerase chain reaction ,Infectious Diseases ,Real-time polymerase chain reaction ,whole-genome sequencing ,COVID-19 Nucleic Acid Testing ,Spike Glycoprotein, Coronavirus ,WGS - Abstract
Reverse transcription-PCRs (RT-PCRs) targeting SARS-CoV-2 variant of concern (VOC) mutations have been developed to simplify their tracking. We evaluated an assay targeting E484K/N501Y to identify B.1.351/P1. Whole-genome sequencing (WGS) confirmed only 72 (59.02%) of 122 consecutive RT-PCR P.1/B.1.351 candidates. Prescreening RT-PCRs must target a wider set of mutations, updated from WGS data from emerging variants.
- Published
- 2021
4. Tumor cell and immune cell profiles in primary human glioblastoma: Impact on patient outcome
- Author
-
Javie Ortiz, Laura Ruíz Martín, Alberto Orfao, Luis Torres Carretero, Maria Almeida, María González-Tablas Pimenta, Daniel Ángel Arandia Guzmán, Juan Carlos Roa Montes de Oca, María Dolores Tabernero, Adelaida Nieto, Álvaro Otero, Andoni García-Martín, Pablo Sousa-Casasnovas, Javier Villaseñor-Ledezma, Daniel Pascual-Argente, Instituto de Salud Carlos III, Ministerio de Economía y Competitividad (España), European Commission, and Junta de Castilla y León
- Subjects
0301 basic medicine ,Male ,lymphocytes ,Myeloid ,Neutrophils ,microglia ,0302 clinical medicine ,Tumor-Associated Macrophages ,Tumor Microenvironment ,IL-2 receptor ,Lymphocytes ,Research Articles ,CD20 ,Aged, 80 and over ,education.field_of_study ,biology ,Brain Neoplasms ,General Neuroscience ,Immune cells ,Middle Aged ,medicine.anatomical_structure ,myeloid cells ,Myeloid cells ,Female ,Microglia ,Research Article ,Adult ,Microenvironment ,CD3 ,Population ,CD19 ,Pathology and Forensic Medicine ,03 medical and health sciences ,Immune system ,Lymphocytes, Tumor-Infiltrating ,immune cells ,medicine ,Biomarkers, Tumor ,Humans ,education ,Aged ,business.industry ,Myeloid-Derived Suppressor Cells ,glioblastoma ,microenvironment ,030104 developmental biology ,Cancer research ,biology.protein ,Neurology (clinical) ,business ,Glioblastoma ,030217 neurology & neurosurgery ,CD8 - Abstract
© 2020 The Authors., The distribution and role of tumor-infiltrating leucocytes in glioblastoma (GBM) remain largely unknown. Here, we investigated the cellular composition of 55 primary (adult) GBM samples by flow cytometry and correlated the tumor immune profile with patient features at diagnosis and outcome. GBM single-cell suspensions were stained at diagnosis (n = 44) and recurrence following radiotherapy and chemotherapy (n = 11) with a panel of 8-color monoclonal antibody combinations for the identification and enumeration of (GFAPCD45) tumor and normal astrocytic cells, infiltrating myeloid cells —i.e. microglial and blood-derived tumor-associated macrophages (TAM), M1-like, and M2-like TAM, neutrophils. and myeloid-derived suppressor cells (MDSC)— and tumor-infiltrating lymphocytes (TIL) —i.e. CD3T-cells and their TCD4, TCD8, TCD4CD8, and (CD25CD127) regulatory (T-regs) subsets, (CD19CD20) B-cells, and (CD16) NK-cells—. Overall, GBM samples consisted of a major population (mean ± 1SD) of tumor and normal astrocytic cells (73% ± 16%) together with a significant but variable fraction of immune cells (24% ± 18%). Within myeloid cells, TAM predominated (13% ± 12%) including both microglial cells (10% ± 11%) and blood-derived macrophages (3% ± 5%), in addition to a smaller proportion of neutrophils (5% ± 9%) and MDSC (4% ± 8%). Lymphocytes were less represented and mostly included TCD4 (0.5% ± 0.7%) and TCD8 cells (0.6% ± 0.7%), together with lower numbers of TCD4CD8 T-cells (0.2% ± 0.4%), T-regs (0.1% ± 0.2%), B-lymphocytes (0.1% ± 0.2%) and NK-cells (0.05% ± 0.05%). Overall, three distinct immune profiles were identified: cases with a minor fraction of leucocytes, tumors with a predominance of TAM and neutrophils, and cases with mixed infiltration by TAM, neutrophils, and T-lymphocytes. Untreated GBM patients with mixed myeloid and lymphoid immune infiltrates showed a significantly shorter patient overall survival versus the other two groups, in the absence of gains of the EGFR gene (p = 0.02). Here we show that immune cell infiltrates are systematically present in GBM, with highly variable levels and immune profiles. Patients with mixed myeloid and T-lymphoid infiltrates showed a worse outcome., Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, Madrid, Spain and fondos FEDER, Grant/Award Number: CB16/12/00400 and ISCIII PI16/0476; Consejería de Sanidad Junta de Castilla y León, Gerencia Regional de Salud, Spain, Grant/Award Number: GRS2049/A/19
- Published
- 2020
5. Multivariate analysis reveals differentially expressed genes among distinct subtypes of diffuse astrocytic gliomas: diagnostic implications
- Author
-
Ana Luísa Vital, Ana B. Nieto-Librero, Nerea González-García, Alberto Orfao, Hermínio Tão, Purificación Galindo-Villardón, María González-Tablas, María Tabernero, Álvaro Otero, Instituto de Salud Carlos III, Ministerio de Economía y Competitividad (España), European Commission, and Junta de Castilla y León
- Subjects
Male ,Science ,Astrocytoma ,Biology ,Article ,CHI3L1 ,Medical research ,Genetics ,medicine ,Humans ,Gene ,Cancer ,Oligonucleotide Array Sequence Analysis ,Regulation of gene expression ,Multidisciplinary ,Brain Neoplasms ,Gene Expression Profiling ,Discriminant Analysis ,Middle Aged ,medicine.disease ,Computational biology and bioinformatics ,Gene Expression Regulation, Neoplastic ,Gene expression profiling ,Oncology ,Genetic marker ,Multivariate Analysis ,Mutation ,Linear Models ,Cancer research ,Medicine ,Female ,XIST ,DNA microarray ,Software ,Neuroscience ,Signal Transduction ,Anaplastic astrocytoma - Abstract
Diagnosis and classification of gliomas mostly relies on histopathology and a few genetic markers. Here we interrogated microarray gene expression profiles (GEP) of 268 diffuse astrocytic gliomas—33 diffuse astrocytomas (DA), 52 anaplastic astrocytomas (AA) and 183 primary glioblastoma (GBM)—based on multivariate analysis, to identify discriminatory GEP that might support precise histopathological tumor stratification, particularly among inconclusive cases with II–III grade diagnosed, which have different prognosis and treatment strategies. Microarrays based GEP was analyzed on 155 diffuse astrocytic gliomas (discovery cohort) and validated in another 113 tumors (validation set) via sequential univariate analysis (pairwise comparison) for discriminatory gene selection, followed by nonnegative matrix factorization and canonical biplot for identification of discriminatory GEP among the distinct histological tumor subtypes. GEP data analysis identified a set of 27 genes capable of differentiating among distinct subtypes of gliomas that might support current histological classification. DA + AA showed similar molecular profiles with only a few discriminatory genes overexpressed (FSTL5 and SFRP2) and underexpressed (XIST, TOP2A and SHOX2) in DA vs AA and GBM. Compared to DA + AA, GBM displayed underexpression of ETNPPL, SH3GL2, GABRG2, SPX, DPP10, GABRB2 and CNTN3 and overexpression of CHI3L1, IGFBP3, COL1A1 and VEGFA, among other differentially expressed genes., MG-T, AO and MDT were supported by the following Grants: CB16/12/00400 (CIBER-ONC, Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, Madrid, Spain and FONDOS FEDER), ISCIII PI16/0476 (Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, Madrid, Spain) and GRS2049/A/19 (Consejería de Sanidad Junta de Castilla y León, Gerencia Regional de Salud, Spain).
- Published
- 2020
6. Heterogeneous EGFR, CDK4, MDM4, and PDGFRA gene expression profiles in primary GBM: No association with patient survival
- Author
-
María Jara-Acevedo, María González-Tablas, Nerea González-García, Álvaro Otero, Ana B. Nieto-Librero, Daniel Pascual, Alberto Orfao, Carlos Prieto, María Dolores Tabernero, Laura Ruiz, Purificación Galindo-Villardón, Pablo Sousa, Hermínio Tão, Daniel Arandia, Ana-Luisa Vital, Instituto de Salud Carlos III, Ministerio de Economía y Competitividad (España), and European Commission
- Subjects
0301 basic medicine ,Cancer Research ,Platelet-Derived Growth Factor Receptor Alpha ,PDGFRA ,amplification ,lcsh:RC254-282 ,Article ,03 medical and health sciences ,Exon ,0302 clinical medicine ,Intragenic deletions ,Gene expression ,medicine ,Epidermal growth factor receptor ,Gene ,Chromosome 7 (human) ,Polysomy ,biology ,glioblastoma ,Gene expression profile ,lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens ,medicine.disease ,030104 developmental biology ,intragenic deletions ,Oncology ,030220 oncology & carcinogenesis ,Cancer research ,biology.protein ,gene expression profile ,heterogeneity ,Heterogeneity ,Glioblastoma - Abstract
This article belongs to the Special Issue Brain Tumors., [Background]: The prognostic impact of the expression profile of genes recurrently amplified in glioblastoma multiforme (GBM) remains controversial., [Methods]: We investigated the RNA gene expression profile of epidermal growth factor receptor (EGFR), cyclin-dependent kinase 4 (CDK4), murine doble minute 4 (MDM4), and platelet derived growth factor receptor alpha (PDGFRA) in 83 primary GBM tumors vs. 42 normal brain tissue samples. Interphase FISH (iFISH) analysis for the four genes, together with analysis of intragenic deletions in EGFR and PDGFRA, were evaluated in parallel at the DNA level. As validation cohort, publicly available RNA gene expression data on 293 samples from 10 different GBM patient series were also studied., [Results]: At the RNA level, CDK4 was the most frequently overexpressed gene (90%) followed by EGFR (58%) and PDGFRA (58%). Chromosome 7 copy number alterations, i.e., trisomy (49%) and polysomy (44%), showed no clear association with EGFR gene expression levels. In turn, intragenic EGFR deletions were found in 39 patients (47%), including EGFRvIII (46%) in association with EGFRvIVa (4%), EGFRvII (2%) or other EGFR deletions (3%) and PDGFRA deletion of exons 8–9 was found in only two tumors (2%)., [Conclusions]: Overall, none of the gene expression profiles and/or intragenic EGFR deletions showed a significant impact on overall survival of GBM supporting the notion that other still unraveled features of the disease might play a more relevant prognostic role in GBM, This research was funded by Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación and Ministerio de Economía y Competitividad (RD12/0036/0048, AES PI16/00476-FONDOS FEDER and CB16/12/00400).
- Published
- 2020
7. Exome Sequencing of Plasma DNA Portrays the Mutation Landscape of Colorectal Cancer and Discovers Mutated VEGFR2 Receptors as Modulators of Antiangiogenic Therapies
- Author
-
Maria Mitsi, Estela Vega, Yolanda Duran, Rafael Álvarez, Elena Garralda, María Isabel Albarrán, Sofia Perea, Antonio Cubillo, Manuel Hidalgo, Marta Camacho-Artacho, T. Sánchez-Pérez, Orlando Domínguez, Carmen Blanco-Aparicio, Álvaro Otero, Natalia Baños, Jorge Monsech, Alba de Martino, Jorge L. Martínez-Torrecuadrada, Tirso Pons, Francesca Sarno, Rodrigo A. Toledo, Victoria Bonilla, and Daniel Lietha
- Subjects
Vascular Endothelial Growth Factor A ,0301 basic medicine ,Cancer Research ,Class I Phosphatidylinositol 3-Kinases ,Protein Conformation ,Somatic cell ,Colorectal cancer ,Angiogenesis Inhibitors ,Biology ,medicine.disease_cause ,Germline ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Germline mutation ,Cell Line, Tumor ,Exome Sequencing ,Biomarkers, Tumor ,medicine ,Animals ,Humans ,Anaplastic Lymphoma Kinase ,Exome ,Proto-Oncogene Proteins c-abl ,Exome sequencing ,Cell Proliferation ,Mutation ,Neovascularization, Pathologic ,Kinase ,Cancer ,medicine.disease ,Vascular Endothelial Growth Factor Receptor-2 ,ErbB Receptors ,030104 developmental biology ,Oncology ,030220 oncology & carcinogenesis ,Cancer research ,Heterografts ,Female ,Colorectal Neoplasms ,Cell-Free Nucleic Acids - Abstract
Purpose: Despite the wide use of antiangiogenic drugs in the clinical setting, predictive biomarkers of response to these drugs are still unknown. Experimental Design: We applied whole-exome sequencing of matched germline and basal plasma cell-free DNA samples (WES-cfDNA) on a RAS/BRAF/PIK3CA wild-type metastatic colorectal cancer patient with primary resistance to standard treatment regimens, including inhibitors to the VEGF:VEGFR2 pathway. We performed extensive functional experiments, including ectopic expression of VEGFR2 mutants in different cell lines, kinase and drug sensitivity assays, and cell- and patient-derived xenografts. Results: WES-cfDNA yielded a 77% concordance rate with tumor exome sequencing and enabled the identification of the KDR/VEGFR2 L840F clonal, somatic mutation as the cause of therapy refractoriness in our patient. In addition, we found that 1% to 3% of samples from cancer sequencing projects harbor KDR somatic mutations located in protein residues frequently mutated in other cancer-relevant kinases, such as EGFR, ABL1, and ALK. Our in vitro and in vivo functional assays confirmed that L840F causes strong resistance to antiangiogenic drugs, whereas the KDR hot-spot mutant R1032Q confers sensitivity to strong VEGFR2 inhibitors. Moreover, we showed that the D717V, G800D, G800R, L840F, G843D, S925F, R1022Q, R1032Q, and S1100F VEGFR2 mutants promote tumor growth in mice. Conclusions: Our study supports WES-cfDNA as a powerful platform for portraying the somatic mutation landscape of cancer and discovery of new resistance mechanisms to cancer therapies. Importantly, we discovered that VEGFR2 is somatically mutated across tumor types and that VEGFR2 mutants can be oncogenic and control sensitivity/resistance to antiangiogenic drugs. Clin Cancer Res; 24(15); 3550–9. ©2018 AACR.
- Published
- 2018
8. Prognostic stratification of adult primary glioblastoma multiforme patients based on their tumor gene amplification profiles
- Author
-
Maria do Carmo Lopes, Hermínio Tão, Ana Filipa Guedes, Maria C. Patino-Alonso, Pablo Sousa, María Dolores Tabernero, Olinda Rebelo, María González-Tablas, Ana Luísa Vital, Ana Belen Nieto, Álvaro Otero, Pim J. French, Maria Rosário Almeida, Alberto Orfao, Luis A. Corchete, Marcos Barbosa, Inês Crespo, Red Temática de Investigación Cooperativa en Cáncer (España), Instituto de Salud Carlos III, Ministerio de Economía y Competitividad (España), European Commission, Junta de Castilla y León, and Neurology
- Subjects
0301 basic medicine ,Gene amplification ,Subtypes ,Survival ,genes supresores de tumores ,glioblastoma ,Chromosome ,Biology ,Classification ,DNA sequencing ,Prognostic stratification ,03 medical and health sciences ,3201.01 Oncología ,030104 developmental biology ,Oncology ,Primary Glioblastoma Multiforme ,síntomas de cáncer ,Chromosomal region ,Gene duplication ,Cancer research ,EGFR Gene Amplification ,Genes, Tumor Suppressor ,Gene ,3205.07 Neurología ,Research Paper - Abstract
Several classification systems have been proposed to address genomic heterogeneity of glioblastoma multiforme, but they either showed limited prognostic value and/or are difficult to implement in routine diagnostics. Here we propose a prognostic stratification model for these primary tumors based on tumor gene amplification profiles, that might be easily implemented in routine diagnostics, and potentially improve the patients management. Gene amplification profiles were prospectively evaluated in 80 primary glioblastoma multiforme tumors using single-nucleotide polymorphism arrays and the results obtained validated in publicly available data from 267/347 cases. Gene amplification was detected in 45% of patients, and chromosome 7p11.2 including the EGFR gene, was the most frequently amplified chromosomal region – either alone (18%) or in combination with amplification of DNA sequences in other chromosomal regions (10% of cases). Other frequently amplified DNA sequences included regions in chromosomes 12q(10%), 4q12(7%) and 1q32.1(4%). Based on their gene amplification profiles, glioblastomas were subdivided into: i) tumors with no gene amplification (55%); ii) tumors with chromosome 7p/EGFR gene amplification (with or without amplification of other chromosomal regions) (38%); and iii) glioblastoma multiforme with a single (11%) or multiple (6%) amplified DNA sequences in chromosomal regions other than chromosome 7p. From the prognostic point of view, these amplification profiles showed a significant impact on overall survival of glioblastoma multiforme patients (p>0.001). Based on these gene amplification profiles, a risk-stratification scoring system was built for prognostic stratification of glioblastoma which might be easily implemented in routine diagnostics, and potentially contribute to improved patient management., This work was supported by RETICC RD06/0020/0035, RD06/0020/0059 and RD12/0036/0048 grants from Red Temática de Investigación Cooperativa en Cáncer (RTICC), Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, (Madrid, Spain and FONDOS FEDER), AES PI16/000476 (Instituto de Salud Carlos III, Madrid, Spain and FONDOS FEDER), GRS909A14 (JCYL) and CB16/12/00400 grant (CIBERONC, Instituto de Salud Carlos III, Ministerio de Economía y Competitividad, Madrid, Spain and FONDOS FEDER).
- Published
- 2018
9. Molecular and Genomic Alterations in Glioblastoma Multiforme
- Author
-
Alberto Orfao, Patrícia Domingues, Álvaro Otero, Catarina de Oliveira, Maria do Carmo Lopes, María Dolores Tabernero, Maria del Carmen Patino, Inês Crespo, María González-Tablas, and Ana Luísa Vital
- Subjects
Epigenomics ,Genetics ,Brain Neoplasms ,Retinoblastoma ,Genetic heterogeneity ,Genomics ,Biology ,medicine.disease ,Pathology and Forensic Medicine ,Mice ,Cyclin-dependent kinase ,Mutation ,Genomic Profile ,medicine ,Cancer research ,biology.protein ,Animals ,Humans ,Tensin ,Glioblastoma ,Protein kinase B ,Signal Transduction - Abstract
In recent years, important advances have been achieved in the understanding of the molecular biology of glioblastoma multiforme (GBM); thus, complex genetic alterations and genomic profiles, which recurrently involve multiple signaling pathways, have been defined, leading to the first molecular/genetic classification of the disease. In this regard, different genetic alterations and genetic pathways appear to distinguish primary (eg, EGFR amplification) versus secondary (eg, IDH1/2 or TP53 mutation) GBM. Such genetic alterations target distinct combinations of the growth factor receptor–ras signaling pathways, as well as the phosphatidylinositol 3-kinase/phosphatase and tensin homolog/AKT, retinoblastoma/cyclin-dependent kinase (CDK) N2A-p16 INK4A , and TP53/mouse double minute (MDM) 2/MDM4/CDKN2A-p14 ARF pathways, in cells that present features associated with key stages of normal neurogenesis and (normal) central nervous system cell types. This translates into well-defined genomic profiles that have been recently classified by The Cancer Genome Atlas Consortium into four subtypes: classic, mesenchymal, proneural, and neural GBM. Herein, we review the most relevant genetic alterations of primary versus secondary GBM, the specific signaling pathways involved, and the overall genomic profile of this genetically heterogeneous group of malignant tumors.
- Published
- 2015
10. The protein expression profile of meningioma cells is associated with distinct cytogenetic tumour subgroups
- Author
-
María Dolores Tabernero, Catarina de Oliveira, Pablo Sousa, Álvaro Otero, Cristina Teodosio, Patrícia Domingues, Ana Belen Nieto, Alberto Orfao, Maria do Carmo Lopes, and Jesús María Gonçalves
- Subjects
Regulation of gene expression ,CD53 ,medicine.medical_specialty ,Monosomy ,Pathology ,Histology ,biology ,medicine.diagnostic_test ,Proliferation index ,CD44 ,Cytogenetics ,medicine.disease ,Pathology and Forensic Medicine ,Meningioma ,Neurology ,Physiology (medical) ,biology.protein ,medicine ,Neurology (clinical) ,Fluorescence in situ hybridization - Abstract
Aims Limited information exists about the impact of cytogenetic alterations on the protein expression profiles of individual meningioma cells and their association with the clinicohistopathological characteristics of the disease. The aim of this study is to investigate the potential association between the immunophenotypic profile of single meningioma cells and the most relevant features of the tumour. Methods Multiparameter flow cytometry (MFC) was used to evaluate the immunophenotypic profile of tumour cells (n = 51 patients) and the Affymetrix U133A chip was applied for the analysis of the gene expression profile (n = 40) of meningioma samples, cytogenetically characterized by interphase fluorescence in situ hybridization. Results Overall, a close association between the pattern of protein expression and the cytogenetic profile of tumour cells was found. Thus, diploid tumours displayed higher levels of expression of the CD55 complement regulatory protein, tumours carrying isolated monosomy 22/del(22q) showed greater levels of bcl2 and PDGFRβ and meningiomas carrying complex karyotypes displayed a greater proliferation index and decreased expression of the CD13 ectoenzyme, the CD9 and CD81 tetraspanins, and the Her2/neu growth factor receptor. From the clinical point of view, higher expression of CD53 and CD44 was associated with a poorer outcome. Conclusions Here we show that the protein expression profile of individual meningioma cells is closely associated with tumour cytogenetics, which may reflect the involvement of different signalling pathways in the distinct cytogenetic subgroups of meningiomas, with specific immunophenotypic profiles also translating into a different tumour clinical behaviour.
- Published
- 2015
11. A Short Region of Connexin43 Reduces Human Glioma Stem Cell Migration, Invasion, and Survival through Src, PTEN, and FAK
- Author
-
Alberto Orfao, José M. Medina, Arantxa Tabernero, Myriam Jaraíz-Rodríguez, Álvaro Otero, Ma Dolores Tabernero, María González-Tablas, Ministerio de Economía y Competitividad (España), Junta de Castilla y León, Fundación Ramón Areces, and European Commission
- Subjects
0301 basic medicine ,connexin ,PTEN ,Human Glioma Stem Cell Migration ,migration ,Biochemistry ,CSK Tyrosine-Protein Kinase ,2302.22 Farmacología Molecular ,Transcellular Cell Migration ,Cell Movement ,glioma ,migración transcelular de células ,Tensin ,lcsh:QH301-705.5 ,conexina 43 ,lcsh:R5-920 ,biology ,genes src ,invasion ,Connexin43 (CX43) ,Cell biology ,Genes, src ,src-Family Kinases ,Mixed Tumor, Malignant ,Neoplastic Stem Cells ,cardiovascular system ,biological phenomena, cell phenomena, and immunity ,Stem cell ,CNS ,lcsh:Medicine (General) ,Src ,PTEN fosfohidrolasa ,Proto-oncogene tyrosine-protein kinase Src ,Cell Survival ,glioma stem cell ,Recombinant Fusion Proteins ,Instituto de Investigación Biomédica de Salamanca ,Motility ,Models, Biological ,Article ,Focal adhesion ,Malignant tumor ,03 medical and health sciences ,Cell Line, Tumor ,Glioma ,Genetics ,medicine ,Humans ,Cell Proliferation ,FAK ,Cell growth ,PTEN Phosphohydrolase ,Cell Biology ,Human glioma ,medicine.disease ,Peptide Fragments ,tumor mixto maligno ,030104 developmental biology ,lcsh:Biology (General) ,Focal Adhesion Protein-Tyrosine Kinases ,Connexin 43 ,biology.protein ,Cancer research ,Stem Cell Migration ,sense organs ,Developmental Biology - Abstract
Summary Connexin43 (CX43), a protein that forms gap junction channels and hemichannels in astrocytes, is downregulated in high-grade gliomas. Its relevance for glioma therapy has been thoroughly explored; however, its positive effects on proliferation are counterbalanced by its effects on migration and invasion. Here, we show that a cell-penetrating peptide based on CX43 (TAT-Cx43266-283) inhibited c-Src and focal adhesion kinase (FAK) and upregulated phosphatase and tensin homolog in glioma stem cells (GSCs) derived from patients. Consequently, TAT-Cx43266-283 reduced GSC motility, as analyzed by time-lapse microscopy, and strongly reduced their invasive ability. Interestingly, we investigated the effects of TAT-Cx43266-283 on freshly removed surgical specimens as undissociated glioblastoma blocks, which revealed a dramatic reduction in the growth, migration, and survival of these cells. In conclusion, a region of CX43 (amino acids 266–283) exerts an important anti-tumor effect in patient-derived glioblastoma models that includes impairment of GSC migration and invasion., Highlights • TAT-Cx43266-283 exerts anti-tumor effects in patient-derived glioblastoma models • TAT-Cx43266-283 targets Src, PTEN, and FAK • TAT-Cx43266-283 inhibits glioma stem cell migration and invasion, In this article, Arantxa Tabernero and colleagues show that a short region of Connexin43 exerts an important anti-tumor effect in patient-derived glioblastoma models that includes the impairment of glioma stem cell migration and invasion. This peptide targets important proteins for glioblastoma, such as Src, FAK, and PTEN, highlighting its relevance for therapeutic development against this incurable disease.
- Published
- 2017
12. PO-291 A cell-penetrating peptide based on the connexin43-Src interacting sequence reduces glioma stem cell migration and proliferation by recruiting Src, Csk and PTEN
- Author
-
María Dolores Tabernero, María González-Tablas, Myriam Jaraíz-Rodríguez, A. Orfao, Arantxa Tabernero, José M. Medina, and Álvaro Otero
- Subjects
Cancer Research ,medicine.diagnostic_test ,biology ,Chemistry ,Cell ,medicine.disease ,Focal adhesion ,medicine.anatomical_structure ,Oncology ,Western blot ,Glioma ,medicine ,Cancer research ,biology.protein ,Phosphorylation ,PTEN ,Stem cell ,Proto-oncogene tyrosine-protein kinase Src - Abstract
Introduction The expression of connexin43 (Cx43), the main gap junction channel-forming protein, is down-regulated in glioma stem cells (GSC). Our previous studies showed that a cell-penetrating peptide containing the region of Cx43 that interacts with the oncoprotein c-Src (TAT-Cx43266–283), inhibits its oncogenic activity and up-regulates the tumour suppressor PTEN. Through this pathway, TAT-Cx43266–283 reduces glioma cell proliferation and reverses the GSC phenotype. Our next goal was to uncover the mechanism of action and the effects of TAT-Cx43266–283 on the migration and invasion of human GSCs. Material and methods Human primary GSCs (G9, G12, G13, G15 and G16) were obtained from glioblastoma patients’ biopsies from the Neurosurgery Service of the Hospital Universitario de Salamanca, Spain. The migration was analysed in these primary GSCs by tracking individual cell trajectories in cultures recorded by Time-Lapse Live-cell Imaging and the invasion with Matrigel-treated transwell inserts. The same tumour biopsies were used to study the effect of TAT-Cx43266–283 on fresh undissociated tumour blocks by time-lapse microscopy. The molecular interactions were studied in G166 GSCs by pull-down assays of biotinylated TAT-Cx43266–283 (TAT-Cx43266–283-B) and Western blot analysis. Results and discussions Our results confirmed that TAT-Cx43266–283 strongly reduces the migration and invasion of human primary GSCs by inhibiting c-Src activity and up-regulating PTEN that consequently, lowers the levels of phosphorylation of focal adhesion kinase tyrosines 397, 576 and 577. In addition, fresh undissociated tumour blocks revealed a dramatic reduction on the survival of these glioblastoma cells when exposed to TAT-Cx43266–283. By pull-down assays, we showed that TAT-Cx43266–283-B recruits c-Src together with its inhibitors PTEN and Csk, confirming the proposed mechanism for Cx43. Conclusion In conclusion, TAT-Cx43266–283 exerts an important antitumor effect by inhibiting c-Src and up-regulating PTEN with the subsequent reduction in FAK phosphorylation required to establish appropriate focal adhesions for migration, proliferation and survival. This inhibition is mediated by the recruitment of the c-Src intrinsic inhibitors, PTEN and Csk. All together, these results reinforce the relevance of this sequence of Cx43 that interacts with c-Src for the development of new therapies against glioblastoma.
- Published
- 2018
13. Genetic/molecular alterations of meningiomas and the signaling pathways targeted
- Author
-
María Dolores Tabernero, Laura Ruiz, Alberto Orfao, María González-Tablas, Patrícia Domingues, Jesús María Gonçalves, David Miranda, Daniel Pascual, Álvaro Otero, Maria do Carmo Lopes, Pablo Sousa, Fundação para a Ciência e a Tecnologia (Portugal), Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo (España), Instituto de Investigación Biomédica de Salamanca, and Instituto de Estudios de Ciencias de la Salud de Castilla y León
- Subjects
Monosomy ,signal pathways ,Chromosomes, Human, Pair 22 ,Nf2 gene ,ComputingMilieux_LEGALASPECTSOFCOMPUTING ,Review ,Biology ,meningioma ,Prognostic stratification ,Meningioma ,Kruppel-Like Factor 4 ,medicine ,otorhinolaryngologic diseases ,Meningeal Neoplasms ,Humans ,Genetic Predisposition to Disease ,Genetics ,Neurofibromin 2 ,Signal Pathways ,Models, Genetic ,Genetic molecular ,genetic/molecular alteration ,University hospital ,medicine.disease ,chromosome 22 ,Oncology ,Chromosome 22 ,Signal Transduction - Abstract
This is an open-access article distributed under the terms of the Creative Commons Attribution License.-- et al., Meningiomas are usually considered to be benign central nervous system tumors; however, they show heterogenous clinical, histolopathological and cytogenetic features associated with a variable outcome. In recent years important advances have been achieved in the identification of the genetic/molecular alterations of meningiomas and the signaling pathways involved. Thus, monosomy 22, which is often associated with mutations of the NF2 gene, has emerged as the most frequent alteration of meningiomas; in addition, several other genes (e.g. AKT1, KLF4, TRAF7, SMO) and chromosomes have been found to be recurrently altered often in association with more complex karyotypes and involvement of multiple signaling pathways. Here we review the current knowledge about the most relevant genes involved and the signaling pathways targeted by such alterations. In addition, we summarize those proposals that have been made so far for classification and prognostic stratification of meningiomas based on their genetic/genomic features., This work was partially supported by grants from the Fundação para a Ciência e Tecnologia (PIC/IC/83108/2007, FCT, Portugal), Fondo de Investigaciones Sanitarias (RD12/0036/0048, Instituto de Salud Carlos III (ISCIII/FEDER), Ministerio de Sanidad y Consumo, Madrid, Spain), and Consejeria Sanidad Junta de Castilla y León, Gerencia Regional de Salud: GRS689/A/11, and Proyecto Intramural-IBSAL IB14-05. Patrícia Domingues is partially supported by a grant (SFRH/BD/64799/2009) from FCT. Maria Dolores Tabernero is supported by IECSCYL (Soria, Spain).
- Published
- 2015
14. Proposal for a new risk stratification classification for meningioma based on patient age, WHO tumor grade, size, localization, and karyotype
- Author
-
Alberto Orfao, Patrícia Domingues, Álvaro Otero, Catarina de Oliveira, Maria do Carmo Lopes, Jesús María Gonçalves, María Dolores Tabernero, Pablo Sousa, Laura Ruiz, Junta de Castilla y León, Fundación Mutua Madrileña, Ministerio de Sanidad y Consumo (España), Instituto de Salud Carlos III, Red Temática de Investigación Cooperativa en Cáncer (España), Fundação para a Ciência e a Tecnologia (Portugal), and Instituto de Estudios de Ciencias de la Salud de Castilla y León
- Subjects
Adult ,Male ,Cancer Research ,medicine.medical_specialty ,Adolescent ,Clinical Investigations ,Disease ,Biology ,iFISH ,Meningioma ,Young Adult ,Recurrence ,Risk Factors ,medicine ,Humans ,Child ,Risk stratification ,In Situ Hybridization, Fluorescence ,Aged ,Aged, 80 and over ,Neoplasm Grading ,SNP arrays ,Base of skull ,medicine.diagnostic_test ,Brain Neoplasms ,Cytogenetics ,Age Factors ,Middle Aged ,medicine.disease ,Survival Analysis ,Surgery ,Oncology ,Karyotyping ,Benign Meningioma ,Cytogenetic Analysis ,Histopathology ,Female ,Neurology (clinical) ,Radiology ,Fluorescence in situ hybridization - Abstract
[Background]: Tumor recurrence remains the major clinical complication of meningiomas, the majority of recurrences occurring among WHO grade I/benign tumors. In the present study, we propose a new scoring system for the prognostic stratification of meningioma patients based on analysis of a large series of meningiomas followed for a median of >5 years. [Methods]: Tumor cytogenetics were systematically investigated by interphase fluorescence in situ hybridization in 302 meningioma samples, and the proposed classification was further validated in an independent series of cases (n = 132) analyzed by high-density (500K) single-nucleotide polymorphism (SNP) arrays. [Results]: Overall, we found an adverse impact on patient relapse-free survival (RFS) for males, presence of brain edema, younger patients (50 mm, tumor localization at intraventricular and anterior cranial base areas, WHO grade II/III meningiomas, and complex karyotypes; the latter 5 variables showed an independent predictive value in multivariate analysis. Based on these parameters, a prognostic score was established for each individual case, and patients were stratified into 4 risk categories with significantly different (P, This work was partially supported by grants from the Fundação para a Ciência e Tecnologia (PIC/IC/83108/2007, FCT, Portugal); Fondo de Investigaciones Sanitarias (FIS/FEDER 06/0312 and RETICC RD06/0020/0035, RD06/0020/0059 and RD12/0036/0048, Instituto de Salud Carlos III (ISCIII/FEDER), Ministerio de Sanidad y Consumo, Madrid, Spain), Caja Burgos (Spain), Fundación MMA (exp 75312010 and 87692011, Madrid, Spain), and Consejeria Sanidad Junta de Castilla y León, Gerencia Regional de Salud: GRS689/A/11. Patrícia Domingues is supported by grant (SFRH/BD/64799/2009) from FCT. Maria Dolores Tabernero is supported by IECSCYL (Fundación Instituto de Estudios Ciencias de la Salud de Castilla y León, Soria, Spain).
- Published
- 2014
15. Vestibular compensation after vestibular schwannoma surgery: normalization of the subjective visual vertical and disability
- Author
-
Nicolas Perez-Fernandez, Pablo Sousa, Álvaro Otero, Angel Batuecas-Caletrio, Santiago Santacruz-Ruiz, and Angel Muñoz-Herrera
- Subjects
Adult ,Male ,medicine.medical_specialty ,Weakness ,Time Factors ,Schwannoma ,Dizziness ,Cohort Studies ,Disability Evaluation ,Vertigo ,otorhinolaryngologic diseases ,medicine ,Videonystagmography ,Humans ,Postural Balance ,Vestibular system ,Vestibular areflexia ,biology ,medicine.diagnostic_test ,business.industry ,Caloric theory ,General Medicine ,Neuroma, Acoustic ,Recovery of Function ,Middle Aged ,Vestibular Function Tests ,biology.organism_classification ,medicine.disease ,Neuroma ,humanities ,Surgery ,Otorhinolaryngology ,Visual Perception ,Female ,medicine.symptom ,business - Abstract
The degree of caloric weakness before surgery influences faster or slower recovery of patients undergoing vestibular schwannoma surgery. The Dizziness Handicap Inventory (DHI) is a good index to show the recovery of patients as it relates directly to an improvement or not of the subjective visual vertical (SVV).To evaluate the process of recovery of patients as measured by the SVV and the DHI after surgical removal of vestibular schwannoma.We studied 24 consecutive patients of the University Hospital of Salamanca who underwent vestibular schwannoma surgery. We assessed age, tumour size, degree of canalicular weakness and preoperative SVV, and their relationship with DHI and SVV at discharge and also at 1, 3 and 6 months postoperatively.Patients with lesser degrees of caloric weakness took longer to normalize SVV than those with a higher caloric weakness before surgery (p0.05). There was a significant correlation between DHI and improvements in SVV with time. The differences disappeared in 6 months where all patients, with greater or lesser degree of caloric weakness, had the same results.
- Published
- 2013
16. Association between mutation of the NF2 gene and monosomy 22 in menopausal women with sporadic meningiomas
- Author
-
Pablo Sousa, Jesús María Gonçalves, Patrícia Domingues, Álvaro Otero, Alberto Orfao, MariaDolores Tabernero, Ana Belen Nieto, María Jara-Acevedo, Arancha Rodríguez Caballero, Junta de Castilla y León, Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo (España), Fundación Caja de Burgos, and Instituto de Estudios de Ciencias de la Salud de Castilla y León
- Subjects
Male ,medicine.medical_specialty ,Monosomy ,Mutation rate ,NF2 gene ,Chromosomes, Human, Pair 22 ,Gene Dosage ,Loss of Heterozygosity ,ComputingMilieux_LEGALASPECTSOFCOMPUTING ,Biology ,medicine.disease_cause ,Gene dosage ,Polymorphism, Single Nucleotide ,Meningioma ,Loss of heterozygosity ,Mutation Rate ,Sporadic meningiomas ,medicine ,Genetics ,Meningeal Neoplasms ,otorhinolaryngologic diseases ,Humans ,Genetics(clinical) ,Genetics (clinical) ,Aged ,Mutation ,Neurofibromin 2 ,Cytogenetics ,Exons ,Middle Aged ,medicine.disease ,Cancer research ,Female ,Chromosome Deletion ,Menopause ,Menopausal women ,Monosomy 22 ,Chromosome 22 ,Research Article - Abstract
This is an open-access article distributed under the terms of the Creative Commons Attribution License., [Background]: Meningioma was the first solid tumor shown to contain a recurrent genetic alteration e.g. monosomy 22/del(22q), NF2 being the most relevant gene involved. Although monosomy 22/del(22q) is present in half of all meningiomas, and meningiomas frequently carry NF2 mutations, no study has been reported so far in which both alterations are simultaneously assessed and correlated with the features of the disease. [Methods]: Here, we analyzed the frequency of both copy number changes involving chromosome 22 and NF2 mutations in 20 sporadic meningiomas using high-density SNP-arrays, interphase-FISH and PCR techniques. [Results]: Our results show a significant frequency of NF2 mutations (6/20 patients, 30%), most of which (5/6) had not been previously reported in sporadic meningiomas. NF2 mutations involved five different exons and led to a truncated protein (p.Leu163CysfsX46, p.Phe62LeufsX61, p.Asp281MetfsX15, p.Phe285LeufsX11, p.Gln389ArgfsX37) and an in frame deletion of Phe119. Interestingly, all NF2 mutated cases were menopausal women with monosomy 22 but not del(22q). [Conclusions]: These results confirm and extend on previous observations about the high frequency and heterogeneity of NF2 mutations in sporadic meningiomas and indicate they could be restricted to a well-defined cytogenetic and clinical subgroup of menopausal women. Further studies in large series of patients are required to confirm our observations., This work has been partially supported by grants from Consejeria Sanidad Junta de Castilla y León, Gerencia Regional de Salud: GRS689/A/11. Fondo de Investigaciones Sanitarias, Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo, Madrid, Spain: RTICC RD06/0020/0035, RD06/0020/0059 and RD12/0036/0048. MD Tabernero is supported by IECSCYL (Fundación Instituto de Estudios Ciencias de la Salud de Castilla y León).
- Published
- 2013
17. Immunophenotypic identification and characterization of tumor cells and infiltrating cell populations in meningiomas
- Author
-
Paloma Bárcena, Álvaro Otero, María Dolores Tabernero, Alberto Orfao, Cristina Teodosio, Pablo Sousa, María C. García-Macías, Maria do Carmo Lopes, Angel Maillo, Catarina de Oliveira, Javier Ortiz, and Patrícia Domingues
- Subjects
Adult ,Male ,CD14 ,Cell ,CD16 ,Biology ,Pathology and Forensic Medicine ,Immunophenotyping ,Immune system ,Lymphocytes, Tumor-Infiltrating ,Phagocytosis ,medicine ,Meningeal Neoplasms ,Cytotoxic T cell ,Humans ,RNA, Messenger ,Cell adhesion ,Aged ,Aged, 80 and over ,Gene Expression Profiling ,Middle Aged ,Flow Cytometry ,Molecular biology ,Cell biology ,Cell Compartmentation ,Neoplasm Proteins ,Gene Expression Regulation, Neoplastic ,medicine.anatomical_structure ,Female ,Meningioma ,CD8 - Abstract
Meningiomas are primary tumors of the central nervous system composed of both neoplastic and other infiltrating cells. We determined the cellular composition of 51 meningioma samples by multiparameter flow cytometric (MFC) immunophenotyping and investigated the potential relationship between mRNA and protein expression levels of neoplastic cells. For immunophenotypic, morphologic, and cytogenetic characterization of individual cell populations, a large panel of markers was used together with phagocytic/endocytic functional assays and MFC sorting. Overall, our results revealed coexistence of CD45 − neoplastic cells and CD45 + immune infiltrating cells in all meningiomas. Infiltrating cells included tissue macrophages, with an HLA-DR + CD14 + CD45 + CD68 + CD16 −/+ CD33 −/+ phenotype and high phagocytic/endocytic activity, and a small proportion of cytotoxic lymphocytes (mostly T CD8 + and natural killer cells). Tumor cells expressed multiple cell adhesion proteins, tetraspanins, HLA-I/HLA-DR molecules, complement regulatory proteins, cell surface ectoenzymes, and growth factor receptors. Noteworthy, the relationship between mRNA and protein levels was variable, depending on the proteins evaluated and the level of infiltration by immune cells. In summary, our results indicate that MFC immunophenotyping provides a reliable tool for the characterization of the patterns of protein expression of different cell populations coexisting in meningioma samples, with a more accurate measure of gene expression profiles of tumor cells at the functional/protein level than conventional mRNA microarray, independently of the degree of infiltration of the tumor by immune cells.
- Published
- 2012
18. Delineation of commonly deleted chromosomal regions in meningiomas by high-density single nucleotide polymorphism genotyping arrays
- Author
-
María Tabernero, Ana Belén Espinosa, Maria C. Patino-Alonso, Abel Castrillo, Cristina Diez-Tascón, Monica Lara, Pablo Sousa, Alberto Orfao, Enrique de Alava, Ana Belen Nieto, Carlos Mackintosh, Angel Maillo, Álvaro Otero, Junta de Castilla y León, Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación (España), Red Temática de Investigación Cooperativa en Cáncer (España), Fundación Mutua Madrileña, and Fundación Sociosanitaria de Castilla-La Mancha
- Subjects
Genetics ,Adult ,Male ,Cancer Research ,High density ,Single-nucleotide polymorphism ,Biology ,Middle Aged ,Polymorphism, Single Nucleotide ,Meningeal Neoplasms ,Humans ,Female ,Chromosome Deletion ,Meningioma ,Genotyping ,In Situ Hybridization, Fluorescence ,Aged ,Oligonucleotide Array Sequence Analysis ,Signal Transduction - Abstract
Despite recent advances in the identification of the cytogenetic profiles of meningiomas, a significant group of tumors still show normal karyotypes or few chromosomal changes. The authors analyzed the cytogenetic profile of 50 meningiomas using fluorescence in situ hybridization and high-density (500 K) single nucleotide polymorphism (SNP) arrays. Our results confirm that del(22q) (52%) and del(1p) (16%) (common deleted regions: 22q11.21-22q13.3. and 1p31.2-p36.33) are the most frequent alterations. Additionally, recurrent monosomy 14 (8%), del(6q) (10%), del(7p) (10%), and del(19q) (4%) were observed, while copy number patterns consistent with recurrent chromosomal gains, gene amplification, and copy number neutral loss of heterozygosity (cnLOH) were either absent or rare. Based on their overall SNP profiles, meningiomas could be classified into: (i) diploid cases, (ii) meningiomas with a single chromosomal change [e.g., monosomy 22/del(22q)] and (iii) tumors with ≥2 altered chromosomes. In summary, our results confirm and extend on previous observations showing that the most recurrent chromosomal abnormalities in meningiomas correspond to chromosome losses localized in chromosomes 1, 22 and less frequently in chromosomes 6, 7, 14, and 19, while chromosomal gains and cnLOH are restricted to a small proportion of cases. Finally, a set of cancer-associated candidate genes associated with the TP53, MYC, CASP3, HDAC1, and TERT signaling pathways was identified, in cases with coexisting monosomy 14 and del(1p). © 2012 Wiley Periodicals, Inc., Supported by: Consejería de Sanidad, Grant number: 66A/06; Consejería de Educación Junta de Castilla y León (Valladolid, Spain), Grant number: HUS 05A06; Instituto de Salud Carlos III, Ministerio de Ciencia e Innovación-FEDER, Madrid, Spain (Fondo de Investigaciones Sanitarias), Grant numbers: FIS/ FEDER 06/0312, RTICC RD06/0020/0035, RD06/0020/0059; Fundación MM, Grant number: AP87692011; IECSCYL (Fundación Instituto de Estudios Ciencias de la Salud de Castilla y León).
- Published
- 2011
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.