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1.
J Pediatr Hematol Oncol ; 43(2): e180-e183, 2021 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-31876779

RESUMEN

WT1-mutant Wilms tumors exhibit a high rate of concomitant CTNNB1 mutations, associated with activated Wnt signaling. Here, we show by laser and manual microdissection of different histologic cell types from 6 WT1-mutant tumor samples that 1 patient's tumor can contain up to 4 distinct mutations in CTNNB1 and/or WTX. Consecutive sections may also harbor different CTNNB1 mutations. The variability of activating CTNNB1 mutations demonstrates the multifocal nature of WT1-mutant Wilms tumors. As multiple independent tumors can occur in patients with constitutional WT1 mutations, they need to be surveyed more closely for tumor development.


Asunto(s)
Evolución Clonal , Neoplasias Renales/patología , Mutación , Proteínas WT1/genética , Tumor de Wilms/patología , beta Catenina/genética , Progresión de la Enfermedad , Humanos , Neoplasias Renales/genética , Pronóstico , Tumor de Wilms/genética
2.
Int J Cancer ; 143(11): 2828-2837, 2018 12 01.
Artículo en Inglés | MEDLINE | ID: mdl-30132831

RESUMEN

A previous genome-wide association study (GWAS) identified common variation at the BARD1 locus as being highly associated with susceptibility to high-risk neuroblastoma, but the mechanisms underlying this association have been not extensively investigated. Here, we performed a fine mapping analysis of BARD1 locus (2q35) using GWAS data from 556 high-risk neuroblastoma patients and 2,575 controls of European-American ancestry, and identified two independent genome-wide neuroblastoma-associated loci. Functional single-nucleotide polymorphism (SNP) prioritization identified two causative variants that independently contributed to neuroblastoma risk, and each replicated robustly in multiple independent cohorts comprising 445 high-risk cases and 3,170 controls (rs17489363: combined p = 1.07 × 10-31 , OR:1.79, 95% CI:1.62-1.98 and rs1048108: combined p = 7.27 × 10-14 , OR:0.65, 95% CI:0.58-0.73). Particularly, the T risk allele of rs17489363 in the canonical promoter region of full-length BARD1 altered binding site of the transcription factor HSF1 and correlated with low expression of full-length BARD1 mRNA and protein. Low-level expression of full-length BARD1 associated with advanced neuroblastoma. In human neuroblastoma cells, attenuating full-length BARD1 increased proliferation and invasion capacity. In conclusion, we have identified two potentially causative SNPs at the BARD1 locus associated with predisposition to high-risk neuroblastoma, and have shown that full-length BARD1 may act as tumor suppressor.


Asunto(s)
Predisposición Genética a la Enfermedad/genética , Neuroblastoma/genética , Polimorfismo de Nucleótido Simple/genética , Proteínas Supresoras de Tumor/genética , Ubiquitina-Proteína Ligasas/genética , Alelos , Estudios de Casos y Controles , Línea Celular Tumoral , Proliferación Celular/genética , Genes Supresores de Tumor/fisiología , Estudio de Asociación del Genoma Completo/métodos , Genotipo , Células HEK293 , Humanos , Lactante , Invasividad Neoplásica/genética , Regiones Promotoras Genéticas/genética , ARN Mensajero/genética
3.
Mol Carcinog ; 56(4): 1281-1289, 2017 04.
Artículo en Inglés | MEDLINE | ID: mdl-27862333

RESUMEN

We have previously reported that the calcium-sensing receptor (CaSR) is expressed in benign, differentiated neuroblastic tumors, and epigenetically silenced in undifferentiated, malignant cases. Furthermore, cinacalcet, an allosteric activator of the CaSR, reduces neuroblastoma tumor growth in preclinical models. However, to identify patients that might benefit from this treatment, a complete understanding of mechanisms governing CaSR expression in these tumors would be required. We have now analyzed two polymorphisms in the promoter region of the CASR gene (rs7652589 and rs1501899) by allelic discrimination in neuroblastoma patients and cell lines. Association of genotypes and haplotypes with CaSR mRNA levels and CASR promoter P2 methylation status was determined. Data presented show that minor alleles rs7652589 and rs1501899, present either in homo- or heterozygosis, were correlated with reduced CaSR mRNA levels in matching primary tumors and this association was independent of CASR promoter P2 hypermethylation. Haplotype AA was independently associated with reduced CaSR expression after adjusting by promoter P2 methylation status. These polymorphisms were identified in some ganglioneuromas in which CaSR expression is low despite exhibiting a high degree of differentiation. Furthermore, homozygous variants rs7652589 and rs1501899 were detected in SH-SY5Y cells, which are devoid of CaSR expression in the absence of hypermethylation of CASR promoter P2. In summary, minor alleles rs7652589 and rs1501899 are associated with reduced CaSR expression in neuroblastic tumors and neuroblastoma cell lines in which the CASR gene promoter P2 is not hypermethylated. Therefore, they potentially represent an additional mechanism of CASR transcriptional regulation in this group of developmental malignancies. © 2016 Wiley Periodicals, Inc.


Asunto(s)
Regulación hacia Abajo , Regulación Neoplásica de la Expresión Génica , Neuroblastoma/genética , Neoplasias del Sistema Nervioso Periférico/genética , Polimorfismo de Nucleótido Simple , Receptores Sensibles al Calcio/genética , Línea Celular Tumoral , Metilación de ADN , Supervivencia sin Enfermedad , Variación Genética , Haplotipos , Humanos , Lactante , Neuroblastoma/epidemiología , Neoplasias del Sistema Nervioso Periférico/epidemiología , Regiones Promotoras Genéticas
4.
Proc Natl Acad Sci U S A ; 111(51): E5564-73, 2014 Dec 23.
Artículo en Inglés | MEDLINE | ID: mdl-25512523

RESUMEN

Osteosarcoma is the most common primary bone tumor, yet there have been no substantial advances in treatment or survival in three decades. We examined 59 tumor/normal pairs by whole-exome, whole-genome, and RNA-sequencing. Only the TP53 gene was mutated at significant frequency across all samples. The mean nonsilent somatic mutation rate was 1.2 mutations per megabase, and there was a median of 230 somatic rearrangements per tumor. Complex chains of rearrangements and localized hypermutation were detected in almost all cases. Given the intertumor heterogeneity, the extent of genomic instability, and the difficulty in acquiring a large sample size in a rare tumor, we used several methods to identify genomic events contributing to osteosarcoma survival. Pathway analysis, a heuristic analytic algorithm, a comparative oncology approach, and an shRNA screen converged on the phosphatidylinositol 3-kinase/mammalian target of rapamycin (PI3K/mTOR) pathway as a central vulnerability for therapeutic exploitation in osteosarcoma. Osteosarcoma cell lines are responsive to pharmacologic and genetic inhibition of the PI3K/mTOR pathway both in vitro and in vivo.


Asunto(s)
Neoplasias Óseas/metabolismo , Genoma Humano , Osteosarcoma/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Neoplasias Óseas/genética , Neoplasias Óseas/patología , Línea Celular Tumoral , Proliferación Celular , Heterogeneidad Genética , Mutación de Línea Germinal , Humanos , Osteosarcoma/genética , Osteosarcoma/patología , Proteína p53 Supresora de Tumor/genética
5.
Nanomedicine ; 12(1): 53-61, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-26492976

RESUMEN

Parathyroid hormone-like hormone (PTHLH) exerts relevant roles in progression and dissemination of several tumors. However, factors influencing its production and secretion have not been fully characterized. The main limitation is the lack of specific, sensitive and widely available techniques to detect and quantify PTHLH. We have developed a lateral flow immunoassay using gold nanoparticles label for the fast and easy detection of PTHLH in lysates and culture media of three human cell lines (HaCaT, LA-N-1, SK-N-AS). Levels in culture media and lysates ranged from 11 to 20 ng/mL and 0.66 to 0.87 µg/mL respectively. Results for HaCaT are in agreement to the previously reported, whereas LA-N-1 and SK-N-AS have been evaluated for the first time. The system also exhibits good performance in human serum samples. This methodology represents a helpful tool for future in vitro and in vivo studies of mechanisms involved in PTHLH production as well as for diagnostics. From the Clinical Editor: Parathyroid Hormone-like Hormone (PTHLH) is known to be secreted by some tumors. However, the detection of this peptide remains difficult. The authors here described their technique of using gold nanoparticles as label for the detection of PTHLH by Lateral-flow immunoassays (LFIAs). The positive results may also point a way to using the same technique for the rapid determination of other relevant cancer proteins.


Asunto(s)
Inmunoensayo/instrumentación , Nanopartículas del Metal/química , Técnicas Analíticas Microfluídicas/instrumentación , Neoplasias Experimentales/diagnóstico , Neoplasias Experimentales/inmunología , Hormona Paratiroidea/análisis , Biomarcadores de Tumor/inmunología , Línea Celular Tumoral , Diseño de Equipo , Análisis de Falla de Equipo , Oro/química , Humanos , Nanopartículas del Metal/ultraestructura , Hormona Paratiroidea/inmunología , Reproducibilidad de los Resultados , Sensibilidad y Especificidad
6.
J Neurosci ; 33(7): 2773-83, 2013 Feb 13.
Artículo en Inglés | MEDLINE | ID: mdl-23407937

RESUMEN

Neuroblastoma is an embryonic tumor derived from cells of the neural crest. Taking advantage of a newly developed neural crest lineage tracer and based on the hypothesis that the molecular mechanisms that mediate neural crest delamination are also likely to be involved in the spread of neuroblastoma, we were able to identify genes that are active both in neural crest development and neuroblastoma tumor formation. A subsequent search of the neuroblastoma gene server for human orthologues of genes differentially expressed in the chick embryo neural crest screen retrieved the LIM domain only protein 4 (LMO4), which was expressed in both cell types analyzed. Functional experiments in these two model systems revealed that LMO4 activity is required for neuroblastoma cell invasion and neural crest delamination. Moreover, we identified LMO4 as an essential cofactor in Snail2-mediated cadherin repression and in the epithelial-to-mesenchymal transition of both neural crest and neuroblastoma cells. Together, our results suggest that the association of high levels of LMO4 with aggressive neuroblastomas is dependent on LMO4 regulation of cadherin expression and hence, tumor invasiveness.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/fisiología , Neoplasias Encefálicas/patología , Transición Epitelial-Mesenquimal/genética , Transición Epitelial-Mesenquimal/fisiología , Proteínas con Dominio LIM/genética , Proteínas con Dominio LIM/fisiología , Cresta Neural/patología , Neuroblastoma/patología , Factores de Transcripción/genética , Factores de Transcripción/fisiología , Animales , Western Blotting , Cadherinas/biosíntesis , Cadherinas/fisiología , Línea Celular Tumoral , Embrión de Pollo , ADN/genética , Electroforesis en Gel de Poliacrilamida , Citometría de Flujo , Vectores Genéticos , Humanos , Inmunohistoquímica , Hibridación in Situ , Lentivirus/genética , Luciferasas/fisiología , Análisis por Micromatrices , Invasividad Neoplásica/genética , Interferencia de ARN , ARN Interferente Pequeño/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Factores de Transcripción de la Familia Snail , Timidina/metabolismo
7.
Carcinogenesis ; 34(2): 268-76, 2013 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-23108190

RESUMEN

Neuroblastic tumors (NTs) include the neuroblastomas, ganglioneuroblastomas and ganglioneuromas. We have reported previously that the calcium-sensing receptor is expressed in differentiated, favorable NTs but almost undetectable in unfavorable neuroblastomas. We have now detected hypermethylation of a particular region within the CpG island encompassing the CaSR gene promoter 2 in neuroblastoma cell lines and 25% primary neuroblastomas. Hypermethylation of this region was associated with reduced CaSR messenger RNA expression and several predictors of poor outcome in neuroblastomas, including MYCN amplification. Treatment with 5'aza-2-deoxycitidine and/or trichostatin A restored CaSR expression in MYCN-amplified cell lines. Following 5'aza-2-deoxycitidine exposure, decreased percentages of methylated CpG sites were observed at the above-mentioned region. By interphase fluorescence in situ hybridization, variable percentages of nuclei with monosomy of chromosome 3, where the human CaSR gene resides, were observed in more than 90% of primary NTs of all subgroups. Nuclei harboring this alteration were heterogeneously distributed among tumor cells. Ectopic overexpression of the calcium-sensing receptor in two MYCN-amplified neuroblastoma cell lines in which this gene is silenced by promoter hypermethylation significantly reduced their in vitro proliferation rates and almost abolished their capacity to generate xenografts in immunocompromised mice. Finally, upon acute exposure to calcium, the primary activator of this receptor, calcium-sensing receptor-overexpressing neuroblastoma cells underwent apoptosis, a process dependent on sustained activation of ERK1/2. These data would support the hypothesis that epigenetic silencing of the CaSR gene is neither an in vitro artefact in neuroblastoma cell lines nor an irrelevant, secondary event in primary NTs, but a significant mechanism for neuroblastoma survival.


Asunto(s)
Apoptosis , Metilación de ADN , Epigénesis Genética , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Neuroblastoma/patología , Receptores Sensibles al Calcio/genética , Glándulas Suprarrenales/metabolismo , Glándulas Suprarrenales/patología , Animales , Western Blotting , Proliferación Celular , Islas de CpG , Femenino , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica , Humanos , Técnicas para Inmunoenzimas , Inmunoprecipitación , Hibridación Fluorescente in Situ , Lactante , Ratones , Ratones Desnudos , Monosomía , Proteína Proto-Oncogénica N-Myc , Estadificación de Neoplasias , Neuroblastoma/genética , Neuroblastoma/mortalidad , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Fosforilación , Regiones Promotoras Genéticas/genética , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores Sensibles al Calcio/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto
8.
Hum Mol Genet ; 19(9): 1651-68, 2010 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-20106868

RESUMEN

Wilms tumors (WTs) are genetically heterogeneous kidney tumors whose cells of origin are unknown. Tumors with WT1 mutations and concomitant loss of the wild-type allele represent a distinct subgroup, frequently associated with mutations in CTNNB1. Here, we describe the establishment and characterization of long-term cell cultures derived from five individual WTs with WT1 mutations. Three of these tumor cell lines also had CTNNB1 mutations and an activated canonical Wnt signaling pathway as measured by beta-catenin/T cell-specific transcription factor (TCF) transcriptional activity. Four of the five Wilms cell lines had a stable normal karyotype for at least 25 passages, and four lines showed loss of heterozygosity of chromosome 11p due to mitotic recombination in 11p11. Gene expression profiling revealed that the WT cell lines are highly similar to human mesenchymal stem cells (MSCs) and FACS analysis demonstrated the expression of MSC-specific surface proteins CD105, CD90 and CD73. The stem cell like nature of the WT cells is further supported by their adipogenic, chondrogenic, osteogenic and myogenic differentiation potentials. By generating multipotent mesenchymal precursors from paraxial mesoderm (PAM) in tissue culture using embryonal stem cells, gene expression profiles of PAM and MSCs were described. Using these published gene sets, we found coexpression of a large number of genes in WT cell lines, PAM and MSCs. Lineage plasticity is indicated by the simultaneous expression of genes from the mesendodermal and neuroectodermal lineages. We conclude that WTs with WT1 mutations have specific traits of PAM, which is the source of kidney stromal cells.


Asunto(s)
Línea Celular Tumoral/citología , Regulación Neoplásica de la Expresión Génica/genética , Genes del Tumor de Wilms , Neoplasias Renales/genética , Células Madre Mesenquimatosas/citología , Mesodermo/metabolismo , Tumor de Wilms/genética , Secuencia de Aminoácidos , Linaje de la Célula/genética , Cromosomas Humanos Par 11/genética , Citometría de Flujo , Perfilación de la Expresión Génica , Humanos , Cariotipificación , Pérdida de Heterocigocidad , Factor de Unión 1 al Potenciador Linfoide/metabolismo , Mesodermo/citología , Datos de Secuencia Molecular , Mutación/genética , beta Catenina/genética
9.
Pediatr Blood Cancer ; 58(4): 532-8, 2012 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-21994054

RESUMEN

BACKGROUND: In Ewing sarcoma (EWS) most of the research on signaling pathways has been performed on cell lines or animal models. The objective of the current study was to determine the relation between clinical outcome and the expression of proteins involved in active growth signaling pathways. METHODS: A paraffin-embedded microarray of 45 human primary EWS tissue specimens was stained with the antibodies against c-KIT, AKT, p-AKT, p-mTOR, IGF-1R, IGFBP-3, MAPK, p27(KIP1) , and p70S6 kinase. Immunohistochemical staining was correlated with patient overall survival (OS). RESULTS: In the univariate analysis 3 variables showed statistical significance to predict survival: presence of metastasis, p-mTOR, and p27(KIP1). A positive stain for p-mTOR (hazard ratio of 4.74 [95% CI (57, 121)]) was significantly (log-rank test with a P = 0.029) associated with better OS. Also, a positive stain for p27(KIP1) (hazard ratio of 6.87 [95% CI (77, 136)] was significantly (log-rank test with a P = 0.009) associated with better OS. Multivariate analysis showed metastasis (HR: 4.3; 95% CI: 0.99, 19; P = 0.05), p-mTOR (HR: 4.8 with 95% CI: 0.6, 38; P = 0.13) and p27 (HR: 5.3; 95% CI: 1.37, 20; P = 0.01) as independent prognostic factors of outcome. CONCLUSIONS: In our series, p-mTOR and p27(KIP1) protein overexpression were independently associated with better survival.


Asunto(s)
Regulación Neoplásica de la Expresión Génica , Proteínas de Neoplasias/biosíntesis , Sarcoma de Ewing/metabolismo , Sarcoma de Ewing/mortalidad , Transducción de Señal , Adolescente , Niño , Preescolar , Supervivencia sin Enfermedad , Femenino , Humanos , Lactante , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/biosíntesis , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Masculino , Metástasis de la Neoplasia , Proteínas Quinasas/biosíntesis , Sarcoma de Ewing/tratamiento farmacológico , Sarcoma de Ewing/patología , Tasa de Supervivencia
10.
Cancer Res ; 82(7): 1193-1207, 2022 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-35101866

RESUMEN

Noncoding cis-regulatory variants have gained interest as cancer drivers, yet progress in understanding their significance is hindered by the numerous challenges and limitations of variant prioritization. To overcome these limitations, we focused on active cis-regulatory elements (aCRE) to design a customized panel for the deep sequencing of 56 neuroblastoma tumor and normal DNA sample pairs. To search for driver mutations, aCREs were defined by reanalysis of H3K27ac chromatin immunoprecipitation sequencing peaks in 25 neuroblastoma cell lines. These regulatory genomic regions were tested for an excess of somatic mutations and assessed for statistical significance using a global approach that accounted for chromatin accessibility and replication timing. Additional validation was provided by whole genome sequence analysis of 151 neuroblastomas. Analysis of HiC data determined the presence of candidate target genes interacting with mutated regions. An excess of somatic mutations in aCREs of diverse genes were identified, including IPO7, HAND2, and ARID3A. CRISPR-Cas9 editing was utilized to assess the functional consequences of mutations in the IPO7-aCRE. Patients with noncoding mutations in aCREs showed inferior overall and event-free survival independent of age at diagnosis, stage, risk stratification, and MYCN status. Expression of aCRE-interacting genes correlated strongly with negative prognostic markers and low survival rates. Moreover, a convergence between the biological functions of aCRE target genes and transcription factors with mutated binding motifs was associated with embryonic development and immune system response. Overall, this strategy enabled the identification of somatic mutations in regulatory elements that collectively promote neuroblastoma tumorigenesis. SIGNIFICANCE: Assessment of noncoding cis-regulatory variants and long-range interaction data highlight the combined effect of somatic mutations in regulatory elements in driving neuroblastoma.


Asunto(s)
Regulación Neoplásica de la Expresión Génica , Neuroblastoma , Proteínas de Unión al ADN/genética , Desarrollo Embrionario , Humanos , Sistema Inmunológico/patología , Mutación , Neuroblastoma/patología , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
11.
Pediatr Blood Cancer ; 57(1): 69-75, 2011 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-21384537

RESUMEN

BACKGROUND: Reported overall survival (OS) rates of patients with localized Ewing sarcoma family of tumors (ESFT) are >80% when treated with the MSKCC P6 protocol. However, it has been associated with a 5.8% incidence of secondary leukemias. A modified P6 (mP6) protocol with reduced exposure to chemotherapy is presented. PROCEDURE: Thirty-one newly diagnosed ESFT patients were enrolled onto this phase II, single-arm, non-randomized protocol. Courses 1, 2 and 4 consisted of cyclophosphamide 4.2 g/m², doxorubicin 75 mg/m², and vincristine 2 mg/m² (CDV). Cycles 3 and 5 consisted of ifosfamide 9 g/m² and etoposide 500 mg/m² (IE). Course 5 ifosfamide was 14 g/m² if necrosis was <90%. RESULTS: Twenty-four patients had loco-regional disease and seven had metastases. The 4-year event-free survival (EFS) rate for patients with localized tumors is 83% and overall survival (OS) is 92%. The 3-year EFS rate for patients with distant metastases is 28% and OS rate is 42%. EWS-FLI1 fusion genes were detected in 17 cases (74%) and EWS-ERG in six cases (26%). Type 1 EWS-FLI1 variant was present in 6/7 metastatic patients and 3/16 loco-regional cases (P = 0.001). None of the patients experienced tumor progression before remission. All relapses occurred within 2 years from the end of treatment and local relapses (n = 3) happened in patients who did not receive radiation therapy. No secondary malignancies have been observed, median follow-up of 4.3 years for surviving patients. CONCLUSIONS: In this pilot study, the mP6 protocol produced a complete remission rate of 83% at 4 years in non-metastatic ESFT reducing the risk of secondary malignancies.


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Ciclofosfamida/administración & dosificación , Doxorrubicina/administración & dosificación , Etopósido/administración & dosificación , Ifosfamida/administración & dosificación , Sarcoma de Ewing/tratamiento farmacológico , Vincristina/administración & dosificación , Adolescente , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Niño , Preescolar , Ciclofosfamida/efectos adversos , Supervivencia sin Enfermedad , Doxorrubicina/efectos adversos , Etopósido/efectos adversos , Femenino , Estudios de Seguimiento , Humanos , Ifosfamida/efectos adversos , Lactante , Masculino , Metástasis de la Neoplasia , Proteínas de Fusión Oncogénica/metabolismo , Proteína Proto-Oncogénica c-fli-1/metabolismo , Proteína EWS de Unión a ARN/metabolismo , Recurrencia , Sarcoma de Ewing/metabolismo , Sarcoma de Ewing/mortalidad , Sarcoma de Ewing/patología , Tasa de Supervivencia , Vincristina/efectos adversos
12.
Nat Commun ; 12(1): 1749, 2021 03 19.
Artículo en Inglés | MEDLINE | ID: mdl-33741928

RESUMEN

Sonic hedgehog medulloblastoma encompasses a clinically and molecularly diverse group of cancers of the developing central nervous system. Here, we use unbiased sequencing of the transcriptome across a large cohort of 250 tumors to reveal differences among molecular subtypes of the disease, and demonstrate the previously unappreciated importance of non-coding RNA transcripts. We identify alterations within the cAMP dependent pathway (GNAS, PRKAR1A) which converge on GLI2 activity and show that 18% of tumors have a genetic event that directly targets the abundance and/or stability of MYCN. Furthermore, we discover an extensive network of fusions in focally amplified regions encompassing GLI2, and several loss-of-function fusions in tumor suppressor genes PTCH1, SUFU and NCOR1. Molecular convergence on a subset of genes by nucleotide variants, copy number aberrations, and gene fusions highlight the key roles of specific pathways in the pathogenesis of Sonic hedgehog medulloblastoma and open up opportunities for therapeutic intervention.


Asunto(s)
Neoplasias Cerebelosas/genética , Regulación Neoplásica de la Expresión Génica , Proteínas Hedgehog/genética , Meduloblastoma/genética , Transcriptoma , Adolescente , Adulto , Niño , Preescolar , Femenino , Redes Reguladoras de Genes , Variación Genética , Humanos , Lactante , Masculino , Persona de Mediana Edad , Transducción de Señal/genética , Adulto Joven
13.
Mol Cancer ; 9: 277, 2010 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-20950435

RESUMEN

BACKGROUND: The chromodomain, helicase DNA-binding protein 5 (CHD5) is a potential tumor suppressor gene located on chromosome 1p36, a region recurrently deleted in high risk neuroblastoma (NB). Previous data have shown that CHD5 mRNA is present in normal neural tissues and in low risk NB, nevertheless, the distribution of CHD5 protein has not been explored. The aim of this study was to investigate CHD5 protein expression as an immunohistochemical marker of outcome in NB. With this purpose, CHD5 protein expression was analyzed in normal neural tissues and neuroblastic tumors (NTs). CHD5 gene and protein expression was reexamined after induction chemotherapy in a subset of high risk tumors to identify potential changes reflecting tumor response. RESULTS: We provide evidence that CHD5 is a neuron-specific protein, absent in glial cells, with diverse expression amongst neuron types. Within NTs, CHD5 immunoreactivity was found restricted to differentiating neuroblasts and ganglion-like cells, and absent in undifferentiated neuroblasts and stromal Schwann cells. Correlation between protein and mRNA levels was found, suggesting transcriptional regulation of CHD5. An immunohistochemical analysis of 90 primary NTs highlighted a strong association of CHD5 expression with favorable prognostic variables (age at diagnosis <12 months, low clinical stage, and favorable histology; P < 0.001 for all), overall survival (OS) (P < 0.001) and event-free survival (EFS) (P < 0.001). Multivariate analysis showed that CHD5 prognostic value is independent of other clinical and biologically relevant parameters, and could therefore represent a marker of outcome in NB that can be tested by conventional immunohistochemistry. The prognostic value of CHD5 was confirmed in an independent, blinded set of 32 NB tumors (P < 0.001).Reactivation of CHD5 expression after induction chemotherapy was observed mainly in those high risk tumors with induced tumor cell differentiation features. Remarkably, these NB tumors showed good clinical response and prolonged patient survival. CONCLUSIONS: The neuron-specific protein CHD5 may represent a marker of outcome in NB that can be tested by conventional immunohistochemistry. Re-establishment of CHD5 expression induced by chemotherapy could be a surrogate marker of treatment response.


Asunto(s)
ADN Helicasas/metabolismo , Regulación Neoplásica de la Expresión Génica , Proteínas del Tejido Nervioso/metabolismo , Neuroblastoma/metabolismo , Western Blotting , Línea Celular Tumoral , Cerebelo/metabolismo , Corteza Cerebral/metabolismo , ADN Helicasas/genética , Ganglios Simpáticos/metabolismo , Humanos , Técnicas In Vitro , Proteínas del Tejido Nervioso/genética , Neuroblastoma/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
14.
Pediatr Blood Cancer ; 54(3): 480-2, 2010 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-19927293

RESUMEN

Axenfeld-Rieger (AR) ocular anomaly might be due to deletions of different chromosomes. No association between AR, mental retardation, and retinoblastoma has been described. We report a 2-month-old female with general development delay and dysmorphic features. AR anomaly was detected, and a retinoblastoma (RB) was diagnosed in a very early stage. De novo 13q deletion was identified. Systemic chemotherapy, focal cryotherapy, transpupillary thermotherapy, brachytherapy, and intra-arterial chemotherapy were needed to control the RB. This is the first report of an association of AR, 13q deletion, and retinoblastoma, to be disclosed in patients born with such ocular and dysmorphic features.


Asunto(s)
Deleción Cromosómica , Cromosomas Humanos Par 13 , Anomalías del Ojo/genética , Retinoblastoma/genética , Femenino , Humanos , Lactante , Síndrome
15.
Cancer Res ; 80(3): 382-393, 2020 02 01.
Artículo en Inglés | MEDLINE | ID: mdl-31784426

RESUMEN

The contribution of coding mutations to oncogenesis has been largely clarified, whereas little is known about somatic mutations in noncoding DNA and their role in driving tumors remains controversial. Here, we used an alternative approach to interpret the functional significance of noncoding somatic mutations in promoting tumorigenesis. Noncoding somatic mutations of 151 neuroblastomas were integrated with ENCODE data to locate somatic mutations in regulatory elements specifically active in neuroblastoma cells, nonspecifically active in neuroblastoma cells, and nonactive. Within these types of elements, transcription factors (TF) were identified whose binding sites were enriched or depleted in mutations. For these TFs, a gene expression signature was built to assess their implication in neuroblastoma. DNA- and RNA-sequencing data were integrated to assess the effects of those mutations on mRNA levels. The pathogenicity of mutations was significantly higher in transcription factor binding site (TFBS) of regulatory elements specifically active in neuroblastoma cells, as compared with the others. Within these elements, there were 18 over-represented TFs involved mainly in cell-cycle phase transitions and 15 under-represented TFs primarily regulating cell differentiation. A gene expression signature based on over-represented TFs correlated with poor survival and unfavorable prognostic markers. Moreover, recurrent mutations in TFBS of over-represented TFs such as EZH2 affected MCF2L and ADP-ribosylhydrolase like 1 expression, among the others. We propose a novel approach to study the involvement of regulatory variants in neuroblastoma that could be extended to other cancers and provide further evidence that alterations of gene expression may have relevant effects in neuroblastoma development. SIGNIFICANCE: These findings propose a novel approach to study regulatory variants in neuroblastoma and suggest that noncoding somatic mutations have relevant implications in neuroblastoma development.


Asunto(s)
Biomarcadores de Tumor/metabolismo , Carcinogénesis/patología , ADN de Neoplasias/metabolismo , Regulación Neoplásica de la Expresión Génica , Mutación , Neuroblastoma/patología , Factores de Transcripción/metabolismo , Sitios de Unión , Biomarcadores de Tumor/genética , Carcinogénesis/genética , Carcinogénesis/metabolismo , ADN de Neoplasias/genética , Humanos , Neuroblastoma/genética , Neuroblastoma/metabolismo , Unión Proteica , Secuencias Reguladoras de Ácidos Nucleicos , Factores de Transcripción/genética , Secuenciación Completa del Genoma
16.
BMC Dev Biol ; 9: 12, 2009 Feb 12.
Artículo en Inglés | MEDLINE | ID: mdl-19216736

RESUMEN

BACKGROUND: Neuroblastic tumors (NBT) derive from neural crest stem cells (NCSC). Histologically, NBT are composed by neuroblasts and Schwannian cells. In culture, neuroblastic (N-), substrate-adherent (S-) and intermediate phenotype (I-) cell subtypes arise spontaneously. METHODS: Here, neuroblastoma (NB) cell line subtypes were characterized according to embryonic peripheral nervous system development markers (GAP43, Phox2b, Sox10, c-kit, GD2, NF68, vimentin, S100beta, calcyclin and ABCG2), morphological features, gene expression and differentiation potential. I-type cells were investigated as a bipotential (neuronal and glial) differentiation stage. RESULTS: Positive immunostaining of NCSC (GAP43, c-kit, NF68, vimentin and Phox2b) and undifferentiated cell (ABCG2) markers was observed in all NB subtypes. N- and I-type cells displayed cytoplasmic membrane GD2 staining, while nuclear calcyclin was restricted to S-type. N- and I-type cells showed similar phenotype and immunoreactivity pattern. Differential gene expression was associated with each cell subtype. N- and I-type cells displayed similar differentiation capacity towards neuronal and glial lineage fates. S-type cells, upon induction, did not show a neuronal-like phenotype, despite gene expression changes. CONCLUSION: Results suggest that N- and I-type NB cell subtypes represent an immature bilineage stage, able to progress towards neuronal and glial fates upon induction of differentiation. S-type cells appear irreversibly committed to a glial lineage fate.


Asunto(s)
Linaje de la Célula/fisiología , Cresta Neural/citología , Neuroblastoma/metabolismo , Neuroblastoma/patología , Células Madre/citología , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP/metabolismo , Bromodesoxiuridina/farmacología , Proteínas de Ciclo Celular/genética , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Linaje de la Célula/genética , Proteína GAP-43/metabolismo , Proteínas de Homeodominio/metabolismo , Humanos , Inmunohistoquímica , N-Acetilgalactosaminiltransferasas/genética , Proteínas de Neoplasias/metabolismo , Proteínas de Neurofilamentos/metabolismo , Proteínas Proto-Oncogénicas c-kit/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína A6 de Unión a Calcio de la Familia S100 , Proteínas S100/genética , Factores de Transcripción SOXE/genética , Células Madre/metabolismo , Factores de Transcripción/metabolismo , Tretinoina/farmacología , Vimentina/metabolismo
17.
BMC Cancer ; 9: 44, 2009 Feb 03.
Artículo en Inglés | MEDLINE | ID: mdl-19192278

RESUMEN

BACKGROUND: Neuroblastoma (NB) tumours have the highest incidence of spontaneous remission, especially among the stage 4s NB subgroup affecting infants. Clinical distinction of stage 4s from lethal stage 4 can be difficult, but critical for therapeutic decisions. The aim of this study was to investigate chromosomal alterations and differential gene expression amongst infant disseminated NB subgroups. METHODS: Thirty-five NB tumours from patients diagnosed at < 18 months (25 stage 4 and 10 stage 4s), were evaluated by allelic and gene expression analyses. RESULTS: All stage 4s patients underwent spontaneous remission, only 48% stage 4 patients survived despite combined modality therapy. Stage 4 tumours were 90% near-diploid/tetraploid, 44% MYCN amplified, 77% had 1p LOH (50% 1p36), 23% 11q and/or 14q LOH (27%) and 47% had 17q gain. Stage 4s were 90% near-triploid, none MYCN amplified and LOH was restricted to 11q. Initial comparison analyses between stage 4s and 4 < 12 months tumours revealed distinct gene expression profiles. A significant portion of genes mapped to chromosome 1 (P < 0.0001), 90% with higher expression in stage 4s, and chromosome 11 (P = 0.0054), 91% with higher expression in stage 4. Less definite expression profiles were observed between stage 4s and 4 < 18m, yet, association with chromosomes 1 (P < 0.0001) and 11 (P = 0.005) was maintained. Distinct gene expression profiles but no significant association with specific chromosomal region localization was observed between stage 4s and stage 4 < 18 months without MYCN amplification. CONCLUSION: Specific chromosomal aberrations are associated with distinct gene expression profiles which characterize spontaneously regressing or aggressive infant NB, providing the biological basis for the distinct clinical behaviour.


Asunto(s)
Aberraciones Cromosómicas , Regulación Neoplásica de la Expresión Génica , Procesos Neoplásicos , Neuroblastoma/genética , Neuroblastoma/patología , Femenino , Perfilación de la Expresión Génica , Humanos , Lactante , Masculino , Estadificación de Neoplasias
18.
Pediatr Blood Cancer ; 53(4): 663-5, 2009 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-19405123

RESUMEN

A male with metastatic paraganglioma received different chemotherapy regimens and 14 arterial embolizations with no or short-lasting clinical benefit. He was started on gemcitabine and docetaxel and, after two cycles, remission of all clinical signs occurred over 2 months. A complete biochemical response was achieved and angiographic signs of portal hypertension disappeared. He received 18 cycles of therapy and no limiting side effects were observed. More than 2 years after gemcitabine and docetaxel treatment, the patient remains symptom free. Gemcitabine and docetaxel could be a potential therapeutic strategy for this patient.


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Paraganglioma/tratamiento farmacológico , Adolescente , Desoxicitidina/administración & dosificación , Desoxicitidina/análogos & derivados , Docetaxel , Humanos , Masculino , Taxoides/administración & dosificación , Gemcitabina
19.
J Pediatr Hematol Oncol ; 31(10): 723-9, 2009 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-19727011

RESUMEN

AIM OF THE STUDY: In this report we describe experience with gemcitabine-docetaxel in pediatric patients with relapsed or refractory sarcomas. PATIENTS AND METHODS: Ten relapsed/refractory pediatric sarcoma patients including 6 Ewing sarcoma, 2 synovial sarcoma, 1 osteosarcoma, and 1 undifferentiated sarcoma, were treated prospectively, in an outpatient setting, with gemcitabine 1000 mg/m over 90 minutes on day 1 and 8, and docetaxel 100 mg/m over 2 to 4 hours on day 8 of a 21-day cycle, as an investigational rescue therapy. RESULTS: The patients (ages 4 to 18) received a total of 70 cycles of therapy (median 6 cycles; range: 4 to 10 y). All symptomatic patients responded clinically to the new regimen. By Response Evaluation Criteria in Solid Tumors criteria, 4 (40%) patients had a complete response (CR), 1 (10%) had a partial response (PR), 3 (30%) had stable disease (SD), and 2 (20%) had a progressive disease (PD), which provides an objective response rate (CR+PR) of 50%. Median duration of response (CR+PR+SD) was 10 months (range: 6 to 32+ mo). Five out of the 10 patients (50%) are alive, with a median follow-up of 48 months from diagnosis. Mild toxicities (no grades 3 to 4) were encountered and managed in the ambulatory setting. CONCLUSIONS: The gemcitabine-docetaxel regimen demonstrated antitumor activity against advanced pediatric (mainly Ewing) sarcomas, allowing for good quality of life. Evaluation in a large, formal phase 2 trials for Ewing patients is ongoing.


Asunto(s)
Desoxicitidina/análogos & derivados , Terapia Recuperativa/métodos , Sarcoma/tratamiento farmacológico , Taxoides/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Niño , Preescolar , Desoxicitidina/administración & dosificación , Progresión de la Enfermedad , Docetaxel , Femenino , Humanos , Masculino , Osteosarcoma/tratamiento farmacológico , Pacientes Ambulatorios , Inducción de Remisión , Sarcoma/mortalidad , Sarcoma de Ewing/tratamiento farmacológico , Sarcoma Sinovial/tratamiento farmacológico , Análisis de Supervivencia , Resultado del Tratamiento , Gemcitabina
20.
Mol Oncol ; 13(9): 1959-1975, 2019 09.
Artículo en Inglés | MEDLINE | ID: mdl-31293052

RESUMEN

We have previously reported the expression of parathyroid hormone-like hormone (PTHLH) in well-differentiated, Schwannian stroma-rich neuroblastic tumors. The aim of this study was to functionally assess the role of PTHLH and its receptor, PTH1R, in neuroblastoma. Stable knockdown of PTHLH and PTH1R was conducted in neuroblastoma cell lines to investigate the succeeding phenotype induced both in vitro and in vivo. Downregulation of PTHLH reduced MYCN expression and subsequently induced cell cycle arrest, senescence, and migration and invasion impairment in a MYCN-amplified, TP53-mutated neuroblastoma cell line. These phenotypes were associated with reduced tumorigenicity in a murine model. We also show that PTHLH expression is not under the control of the calcium-sensing receptor in neuroblastoma. Conversely, its production is stimulated by epidermal growth factor receptor (EGFR). Accordingly, irreversible EGFR inhibition with canertinib abolished PTHLH expression. The oncogenic role of PTHLH appeared to be a consequence of its intracrine function, as downregulation of its receptor, PTH1R, increased anchorage-independent growth and induced a more undifferentiated, invasive phenotype. Respectively, high PTH1R mRNA expression was found in MYCN nonamplified primary tumors and also significantly associated with other prognostic factors of good outcome. This study provides the first evidence of the dual role of PTHLH in the behavior of neuroblastomas. Moreover, the identification of EGFR as a transcriptional regulator of PTHLH in neuroblastoma provides a novel therapeutic opportunity to promote a less aggressive tumor phenotype through irreversible inhibition of EGFR tyrosine kinase activity.


Asunto(s)
Regulación Neoplásica de la Expresión Génica , Neuroblastoma/metabolismo , Proteína Relacionada con la Hormona Paratiroidea/metabolismo , Receptor de Hormona Paratiroídea Tipo 1/biosíntesis , Animales , Línea Celular Tumoral , Femenino , Células HEK293 , Humanos , Ratones , Ratones Desnudos , Mutación , Neuroblastoma/genética , Neuroblastoma/patología , Proteína Relacionada con la Hormona Paratiroidea/genética , Receptor de Hormona Paratiroídea Tipo 1/genética , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo
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