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1.
Mol Oncol ; 13(9): 1959-1975, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31293052

RESUMO

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.

2.
Childs Nerv Syst ; 35(5): 865-869, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30707305

RESUMO

Medulloblastoma is the most common malignant brain tumor in children. Approximately 30% of children with medulloblastoma will progress or relapse despite being treated. New therapies have been proposed in recent years, including high-dose chemotherapy, immunotherapy, and targeted therapy. However, the best treatment for these patients remains unclear, and in this situation prognosis is poor. Oral etoposide has been used as a single agent or in combination for treating relapsed brain tumors since the 1990s. We report an 8-year-old patient with recurrent metastatic medulloblastoma who had an excellent response after treatment with oral etoposide, maintaining a great quality of life. As clinicians, we must always try to include our patients in clinical trials; however, when this is not possible, we should not forget that "old drugs" such as oral etoposide may work in some patients, with a good response of the tumor, and what is most important, providing the patient with a good quality of life.

3.
Sci Transl Med ; 11(476)2019 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-30674657

RESUMO

Retinoblastoma is a pediatric solid tumor of the retina activated upon homozygous inactivation of the tumor suppressor RB1 VCN-01 is an oncolytic adenovirus designed to replicate selectively in tumor cells with high abundance of free E2F-1, a consequence of a dysfunctional RB1 pathway. Thus, we reasoned that VCN-01 could provide targeted therapeutic activity against even chemoresistant retinoblastoma. In vitro, VCN-01 effectively killed patient-derived retinoblastoma models. In mice, intravitreous administration of VCN-01 in retinoblastoma xenografts induced tumor necrosis, improved ocular survival compared with standard-of-care chemotherapy, and prevented micrometastatic dissemination into the brain. In juvenile immunocompetent rabbits, VCN-01 did not replicate in retinas, induced minor local side effects, and only leaked slightly and for a short time into the blood. Initial phase 1 data in patients showed the feasibility of the administration of intravitreous VCN-01 and resulted in antitumor activity in retinoblastoma vitreous seeds and evidence of viral replication markers in tumor cells. The treatment caused local vitreous inflammation but no systemic complications. Thus, oncolytic adenoviruses targeting RB1 might provide a tumor-selective and chemotherapy-independent treatment option for retinoblastoma.

4.
Nat Med ; 25(1): 176-187, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30531922

RESUMO

Neurofibromatosis type 1 (NF1) is a common tumor predisposition syndrome in which glioma is one of the prevalent tumors. Gliomagenesis in NF1 results in a heterogeneous spectrum of low- to high-grade neoplasms occurring during the entire lifespan of patients. The pattern of genetic and epigenetic alterations of glioma that develops in NF1 patients and the similarities with sporadic glioma remain unknown. Here, we present the molecular landscape of low- and high-grade gliomas in patients affected by NF1 (NF1-glioma). We found that the predisposing germline mutation of the NF1 gene was frequently converted to homozygosity and the somatic mutational load of NF1-glioma was influenced by age and grade. High-grade tumors harbored genetic alterations of TP53 and CDKN2A, frequent mutations of ATRX associated with Alternative Lengthening of Telomere, and were enriched in genetic alterations of transcription/chromatin regulation and PI3 kinase pathways. Low-grade tumors exhibited fewer mutations that were over-represented in genes of the MAP kinase pathway. Approximately 50% of low-grade NF1-gliomas displayed an immune signature, T lymphocyte infiltrates, and increased neo-antigen load. DNA methylation assigned NF1-glioma to LGm6, a poorly defined Isocitrate Dehydrogenase 1 wild-type subgroup enriched with ATRX mutations. Thus, the profiling of NF1-glioma defined a distinct landscape that recapitulates a subset of sporadic tumors.


Assuntos
Neoplasias Encefálicas/complicações , Neoplasias Encefálicas/genética , Glioma/complicações , Glioma/genética , Neurofibromatose 1/complicações , Neurofibromatose 1/genética , Adolescente , Adulto , Antígenos de Neoplasias/metabolismo , Neoplasias Encefálicas/imunologia , Criança , Pré-Escolar , Estudos de Coortes , Metilação de DNA/genética , Feminino , Mutação em Linhagem Germinativa/genética , Glioma/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Neurofibromina 1/genética , Reprodutibilidade dos Testes , Linfócitos T/imunologia , Transcriptoma/genética , Proteína Nuclear Ligada ao X/genética , Adulto Jovem
5.
Science ; 360(6386): 331-335, 2018 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-29674595

RESUMO

Gliomas with histone H3 lysine27-to-methionine mutations (H3K27M-glioma) arise primarily in the midline of the central nervous system of young children, suggesting a cooperation between genetics and cellular context in tumorigenesis. Although the genetics of H3K27M-glioma are well characterized, their cellular architecture remains uncharted. We performed single-cell RNA sequencing in 3321 cells from six primary H3K27M-glioma and matched models. We found that H3K27M-glioma primarily contain cells that resemble oligodendrocyte precursor cells (OPC-like), whereas more differentiated malignant cells are a minority. OPC-like cells exhibit greater proliferation and tumor-propagating potential than their more differentiated counterparts and are at least in part sustained by PDGFRA signaling. Our study characterizes oncogenic and developmental programs in H3K27M-glioma at single-cell resolution and across genetic subclones, suggesting potential therapeutic targets in this disease.


Assuntos
Neoplasias Encefálicas/patologia , Carcinogênese/genética , Glioma/patologia , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Oncogenes , Neoplasias Encefálicas/genética , Proliferação de Células , Glioma/genética , Histonas/metabolismo , Humanos , Proteína Quinase 7 Ativada por Mitógeno/genética , Mutação , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos
6.
Clin Cancer Res ; 24(6): 1355-1363, 2018 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-29351917

RESUMO

Purpose: The classification of medulloblastoma into WNT, SHH, group 3, and group 4 subgroups has become of critical importance for patient risk stratification and subgroup-tailored clinical trials. Here, we aimed to develop a simplified, clinically applicable classification approach that can be implemented in the majority of centers treating patients with medulloblastoma.Experimental Design: We analyzed 1,577 samples comprising previously published DNA methylation microarray data (913 medulloblastomas, 457 non-medulloblastoma tumors, 85 normal tissues), and 122 frozen and formalin-fixed paraffin-embedded medulloblastoma samples. Biomarkers were identified applying stringent selection filters and Linear Discriminant Analysis (LDA) method, and validated using DNA methylation microarray data, bisulfite pyrosequencing, and direct-bisulfite sequencing.Results: Using a LDA-based approach, we developed and validated a prediction method (EpiWNT-SHH classifier) based on six epigenetic biomarkers that allowed for rapid classification of medulloblastoma into the clinically relevant subgroups WNT, SHH, and non-WNT/non-SHH with excellent concordance (>99%) with current gold-standard methods, DNA methylation microarray, and gene signature profiling analysis. The EpiWNT-SHH classifier showed high prediction capacity using both frozen and formalin-fixed material, as well as diverse DNA methylation detection methods. Similarly, we developed a classifier specific for group 3 and group 4 tumors, based on five biomarkers (EpiG3-G4) with good discriminatory capacity, allowing for correct assignment of more than 92% of tumors. EpiWNT-SHH and EpiG3-G4 methylation profiles remained stable across tumor primary, metastasis, and relapse samples.Conclusions: The EpiWNT-SHH and EpiG3-G4 classifiers represent a new simplified approach for accurate, rapid, and cost-effective molecular classification of single medulloblastoma DNA samples, using clinically applicable DNA methylation detection methods. Clin Cancer Res; 24(6); 1355-63. ©2018 AACR.


Assuntos
Biomarcadores Tumorais , Neoplasias Cerebelares/diagnóstico , Neoplasias Cerebelares/genética , Estudos de Associação Genética , Predisposição Genética para Doença , Meduloblastoma/diagnóstico , Meduloblastoma/genética , Biópsia , Ilhas de CpG , Metilação de DNA , Epigênese Genética , Epigenômica/métodos , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Reprodutibilidade dos Testes
7.
J Control Release ; 255: 108-119, 2017 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-28412222

RESUMO

Neuroblastoma is a pediatric solid tumor with high expression of the tumor associated antigen disialoganglioside GD2. Despite initial response to induction therapy, nearly 50% of high-risk neuroblastomas recur because of chemoresistance. Here we encapsulated the topoisomerase-I inhibitor SN-38 in polymeric nanoparticles (NPs) surface-decorated with the anti-GD2 mouse mAb 3F8 at a mean density of seven antibody molecules per NP. The accumulation of drug-loaded NPs targeted with 3F8 versus with control antibody was monitored by microdialysis in patient-derived GD2-expressing neuroblastoma xenografts. We showed that the extent of tumor penetration by SN-38 was significantly higher in mice receiving the targeted nano-drug delivery system when compared to non-targeted system or free drug. This selective penetration of the tumor extracellular fluid translated into a strong anti-tumor effect prolonging survival of mice bearing GD2-high neuroblastomas in vivo.


Assuntos
Anticorpos Monoclonais/administração & dosagem , Antineoplásicos Fitogênicos/administração & dosagem , Camptotecina/análogos & derivados , Líquido Extracelular/metabolismo , Imunoglobulina G/administração & dosagem , N-Acetilgalactosaminiltransferases/antagonistas & inibidores , Nanopartículas/administração & dosagem , Neuroblastoma/metabolismo , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais Murinos , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacocinética , Camptotecina/administração & dosagem , Camptotecina/química , Camptotecina/farmacocinética , Linhagem Celular Tumoral , Pré-Escolar , Liberação Controlada de Fármacos , Regulação Neoplásica da Expressão Gênica , Humanos , Imunoglobulina G/química , Irinotecano , Masculino , Camundongos Nus , N-Acetilgalactosaminiltransferases/genética , N-Acetilgalactosaminiltransferases/imunologia , N-Acetilgalactosaminiltransferases/metabolismo , Nanopartículas/química , Neuroblastoma/tratamento farmacológico , Distribuição Tecidual , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Nat Med ; 23(4): 483-492, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28263309

RESUMO

Diffuse intrinsic pontine glioma (DIPG) is an aggressive brain tumor that is located in the pons and primarily affects children. Nearly 80% of DIPGs harbor mutations in histone H3 genes, wherein lysine 27 is substituted with methionine (H3K27M). H3K27M has been shown to inhibit polycomb repressive complex 2 (PRC2), a multiprotein complex responsible for the methylation of H3 at lysine 27 (H3K27me), by binding to its catalytic subunit EZH2. Although DIPGs with the H3K27M mutation show global loss of H3K27me3, several genes retain H3K27me3. Here we describe a mouse model of DIPG in which H3K27M potentiates tumorigenesis. Using this model and primary patient-derived DIPG cell lines, we show that H3K27M-expressing tumors require PRC2 for proliferation. Furthermore, we demonstrate that small-molecule EZH2 inhibitors abolish tumor cell growth through a mechanism that is dependent on the induction of the tumor-suppressor protein p16INK4A. Genome-wide enrichment analyses show that the genes that retain H3K27me3 in H3K27M cells are strong polycomb targets. Furthermore, we find a highly significant overlap between genes that retain H3K27me3 in the DIPG mouse model and in human primary DIPGs expressing H3K27M. Taken together, these results show that residual PRC2 activity is required for the proliferation of H3K27M-expressing DIPGs, and that inhibition of EZH2 is a potential therapeutic strategy for the treatment of these tumors.


Assuntos
Neoplasias do Tronco Encefálico/genética , Proliferação de Células/genética , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Glioma/genética , Histonas/genética , Animais , Benzamidas/farmacologia , Neoplasias Encefálicas/genética , Sistemas CRISPR-Cas , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Cromatografia Líquida , Inibidor p16 de Quinase Dependente de Ciclina/efeitos dos fármacos , Inibidor p16 de Quinase Dependente de Ciclina/genética , Modelos Animais de Doenças , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidores , Técnicas de Inativação de Genes , Glioblastoma/genética , Humanos , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Indazóis/farmacologia , Camundongos , Camundongos SCID , Terapia de Alvo Molecular , Mutação , Transplante de Neoplasias , Células-Tronco Neurais , Complexo Repressor Polycomb 2/genética , Piridonas/farmacologia , Espectrometria de Massas em Tandem , Proteína Supressora de Tumor p14ARF/efeitos dos fármacos , Proteína Supressora de Tumor p14ARF/genética
9.
Mol Carcinog ; 56(4): 1281-1289, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27862333

RESUMO

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.


Assuntos
Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Neuroblastoma/genética , Neoplasias do Sistema Nervoso Periférico/genética , Polimorfismo de Nucleotídeo Único , Receptores de Detecção de Cálcio/genética , Linhagem Celular Tumoral , Metilação de DNA , Intervalo Livre de Doença , Variação Genética , Haplótipos , Humanos , Lactente , Neuroblastoma/epidemiologia , Neoplasias do Sistema Nervoso Periférico/epidemiologia , Regiões Promotoras Genéticas
10.
Rev. neurol. (Ed. impr.) ; 63(9): 411-414, 1 nov., 2016. ilus
Artigo em Espanhol | IBECS | ID: ibc-157605

RESUMO

Introducción. El glioblastoma multiforme congénito representa sólo el 3% de los tumores congénitos del sistema nervioso central, y su ubicación infrantentorial es excepcional. Caso clínico. Recién nacido con un glioblastoma multiforme congénito sin mutación en el gen TP53 ni inmunorreactividad nuclear p53, que infiltraba prácticamente todo el tronco cerebral e invadía también estructuras supratentoriales. Conclusiones. Hasta donde sabemos, sólo se han referido previamente cuatro casos de localización infratentorial, tres en el cerebelo y uno en el tronco del encéfalo. La biología del glioblastoma multiforme congénito no se conoce bien y, a diferencia del glioblastoma multiforme en la edad adulta, las mutaciones en el gen TP53 son poco frecuentes, sin que eso parezca implicar un mejor pronóstico. Estas observaciones sugieren que el glioblastoma multiforme con origen en la vida fetal tiene una biología diferente del que se presenta en otras etapas de la vida (AU)


Introduction. Congenital glioblastoma multiforme represents only 3% of congenital central nervous system tumours and an infratentorial location is unusual. Case report. A newborn with congenital glioblastoma multiforme with no mutation in the TP53 gene or p53 nuclear immunoreactivity that infiltrated practically the whole brainstem and also invaded supratentorial structures. Conclusions. As far as we know, only four cases with an infratentorial location have been reported previously, three in the cerebellum and one in the brainstem. The biology of congenital glioblastoma multiforme is not well known and, unlike glioblastoma multiforme in adults and children, mutations in the TP53 gene are uncommon. However, this is not associated with a more favourable prognosis. These observations suggest that specific biological processes underlie fetal glioblastoma multiforme development (AU)


Assuntos
Humanos , Masculino , Recém-Nascido , Glioblastoma/congênito , Neoplasias Infratentoriais/congênito , Neoplasias Encefálicas/congênito , Genes p53/genética , Megalencefalia/genética
11.
Oncotarget ; 7(29): 46283-46300, 2016 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-27317769

RESUMO

Ewing sarcoma (ES) is an aggressive tumor defined by EWSR1 gene fusions that behave as an oncogene. Here we demonstrate that RING1B is highly expressed in primary ES tumors, and its expression is independent of the fusion oncogene. RING1B-depleted ES cells display an expression profile enriched in genes functionally involved in hematological development but RING1B depletion does not induce cellular differentiation. In ES cells, RING1B directly binds the SCN8A sodium channel promoter and its depletion results in enhanced Nav1.6 expression and function. The signaling pathway most significantly modulated by RING1B is NF-κB. RING1B depletion results in enhanced p105/p50 expression, which sensitizes ES cells to apoptosis by FGFR/SHP2/STAT3 blockade. Reduced NaV1.6 function protects ES cells from apoptotic cell death by maintaining low NF-κB levels. Our findings identify RING1B as a trait of the cell-of-origin and provide a potential targetable vulnerability.


Assuntos
Neoplasias Ósseas/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.6/metabolismo , NF-kappa B/metabolismo , Complexo Repressor Polycomb 1/metabolismo , Sarcoma de Ewing/metabolismo , Transdução de Sinais , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Humanos , Sarcoma de Ewing/genética , Sarcoma de Ewing/patologia , Transdução de Sinais/fisiologia , Células Tumorais Cultivadas
12.
Cancer Lett ; 380(1): 10-9, 2016 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-27319373

RESUMO

Translational research in retinoblastoma - a pediatric tumor that originates during the development of the retina - would be improved by the creation of new patient-derived models. Using tumor samples from enucleated eyes we established a new battery of preclinical models that grow in vitro in serum-free medium and in vivo in immunodeficient mice. To examine whether the new xenografts recapitulate human disease and disseminate from the retina to the central nervous system, we evaluated their histology and the presence of molecular markers of dissemination that are used in the clinical setting to detect extraocular metastases. We evaluated GD2 synthase and CRX as such markers and generated a Taqman real-time quantitative PCR method to measure CRX mRNA for rapid, sensitive and specific quantification of local and metastatic tumor burden. This approach was able to detect 1 human retinoblastoma cell in 100.000 mouse brain cells. Our research adds novel preclinical tools for the discovery of new retinoblastoma treatments for clinical translation.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/enzimologia , Movimento Celular , Proteínas de Homeodomínio/metabolismo , N-Acetilgalactosaminiltransferases/metabolismo , Neoplasias Experimentais/enzimologia , Neoplasias da Retina/enzimologia , Retinoblastoma/enzimologia , Transativadores/metabolismo , Animais , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/secundário , Linhagem Celular Tumoral , Pré-Escolar , Feminino , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Proteínas de Homeodomínio/genética , Humanos , Lactente , Camundongos Nus , N-Acetilgalactosaminiltransferases/genética , Micrometástase de Neoplasia , Transplante de Neoplasias , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias da Retina/genética , Neoplasias da Retina/patologia , Retinoblastoma/genética , Retinoblastoma/secundário , Transdução de Sinais , Transativadores/genética , Células Tumorais Cultivadas
13.
Oncotarget ; 7(13): 16112-29, 2016 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-26893368

RESUMO

The calcium-sensing receptor is a G protein-coupled receptor that exerts cell-type specific functions in numerous tissues and some cancers. We have previously reported that this receptor exhibits tumor suppressor properties in neuroblastoma. We have now assessed cinacalcet, an allosteric activator of the CaSR approved for clinical use, as targeted therapy for this developmental tumor using neuroblastoma cell lines and patient-derived xenografts (PDX) with different MYCN and TP53 status. In vitro, acute exposure to cinacalcet induced endoplasmic reticulum stress coupled to apoptosis via ATF4-CHOP-TRB3 in CaSR-positive, MYCN-amplified cells. Both phenotypes were partially abrogated by phospholipase C inhibitor U73122. Prolonged in vitro treatment also promoted dose- and time-dependent apoptosis in CaSR-positive, MYCN-amplified cells and, irrespective of MYCN status, differentiation in surviving cells. Cinacalcet significantly inhibited tumor growth in MYCN-amplified xenografts and reduced that of MYCN-non amplified PDX. Morphology assessment showed fibrosis in MYCN-amplified xenografts exposed to the drug. Microarrays analyses revealed up-regulation of cancer-testis antigens (CTAs) in cinacalcet-treated MYCN-amplified tumors. These were predominantly CTAs encoded by genes mapping on chromosome X, which are the most immunogenic. Other modulated genes upon prolonged exposure to cinacalcet were involved in differentiation, cell cycle exit, microenvironment remodeling and calcium signaling pathways. CTAs were up-regulated in PDX and in vitro models as well. Moreover, progressive increase of CaSR expression upon cinacalcet treatment was seen both in vitro and in vivo. In summary, cinacalcet reduces neuroblastoma tumor growth and up-regulates CTAs. This effect represents a therapeutic opportunity and provides surrogate circulating markers of neuroblastoma response to this treatment.


Assuntos
Antígenos de Neoplasias/biossíntese , Antineoplásicos/farmacologia , Cinacalcete/farmacologia , Neuroblastoma/patologia , Animais , Antígenos de Neoplasias/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Camundongos , Camundongos Nus , Proteína Proto-Oncogênica N-Myc/genética , Neuroblastoma/metabolismo , Receptores de Detecção de Cálcio/agonistas , Receptores de Detecção de Cálcio/efeitos dos fármacos , Proteína Supressora de Tumor p53/genética , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Genom Data ; 5: 360-3, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26484286

RESUMO

Neuroblastoma (NB) is one of the most frequently occurring extracranial solid tumors of childhood (Maris et al., 2007 [1]; Brodeur, 2003 [2]). Probability of cure varies according to patient's age, extent of disease and tumor biology (Maris et al., 2007 [1]; Brodeur, 2003 [2]; Cohn et al., 2009 [3]). However, the etiology of this developmental tumor is unknown. Recent evidence has shown that pediatric solid tumors, including NB, harbor a paucity of recurrent genetic mutations, with a significant proportion of recurrent events converging on epigenetic mechanisms (Cheung et al., 2012 [4]; Molenaar et al., 2012 [5]; Pugh et al., 2013 [6]; Sausen et al., 2013 [7]. We have analyzed the DNA methylome of neuroblastoma using high-density microarrays (Infinium Human Methylation 450k BeadChip) to define the epigenetic landscape of this pediatric tumor and its potential clinicopathological impact. Here, we provide the detail of methods and quality control parameters of the microarray data used for the study. Methylation data has been deposited at NCBI Gene Expression Omnibus data repository, accession number GSE54719; superseries record GSE54721.

15.
Epigenomics ; 7(7): 1137-53, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26067621

RESUMO

AIM: To define the DNA methylation landscape of neuroblastoma and its clinicopathological impact. MATERIALS & METHODS: Microarray DNA methylation data were analyzed and associated with functional/regulatory genome annotation data, transcriptional profiles and clinicobiological parameters. RESULTS: DNA methylation changes in neuroblastoma affect not only promoters but also intragenic and intergenic regions at cytosine-phosphate-guanine (CpG) and non-CpG sites, and target functional chromatin domains of development and cancer-related genes such as CCND1. Tumors with diverse clinical risk showed differences affecting CpG and, remarkably, non-CpG sites. Non-CpG methylation observed essentially in clinically favorable cases was associated with the differentiation status of neuroblastoma and expression of key genes such as ALK. CONCLUSION: This epigenetic fingerprint of neuroblastoma provides new insights into the pathogenesis and clinical behavior of this pediatric tumor.


Assuntos
Neoplasias Encefálicas/genética , Ciclina D1/genética , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Neuroblastoma/genética , Receptores Proteína Tirosina Quinases/genética , Quinase do Linfoma Anaplásico , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/mortalidade , Linhagem Celular Tumoral , Criança , Pré-Escolar , Cromatina/química , Cromatina/metabolismo , Ilhas de CpG , Ciclina D1/metabolismo , Impressões Digitais de DNA , Metilação de DNA , DNA Intergênico , Feminino , Perfilação da Expressão Gênica , Genoma Humano , Humanos , Lactente , Masculino , Neuroblastoma/diagnóstico , Neuroblastoma/metabolismo , Neuroblastoma/mortalidade , Análise de Sequência com Séries de Oligonucleotídeos , Prognóstico , Regiões Promotoras Genéticas , Receptores Proteína Tirosina Quinases/metabolismo , Análise de Sobrevida
16.
Pharm Res ; 32(9): 2889-900, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25773723

RESUMO

PURPOSE: To develop a reproducible microdialysis-tumor homogenate method for the study of the intratumor distribution of a highly hydrophobic anticancer drug (SN-38; 7-ethyl-10-hydroxycamptothecin) in neuroblastoma patient-derived xenografts. METHODS: We studied the nonspecific binding of SN-38 to the microdialysis tubing in the presence of 2-hydroxypropyl-beta-cyclodextrin (HPBCD) in the perfusate. We calibrated the microdialysis probes by the zero flow rate (ZFR) method and calculated the enhancement factor (f = extrapolated SN-38 concentration at the ZFR / SN-38 concentration in the dialysed solution) of HPBCD. We characterized the extravasation of HPBCD to tumors engrafted in mice. In vivo microdialysis and terminal homogenate data at the steady state (subcutaneous pump infusions) were used to calculate the volume of distribution of unbound SN-38 (Vu,tumor) in neuroblastoma. RESULTS: HPBCD (10% w/v) in the perfusate prevented the nonspecific binding of SN-38 to the microdialysis probe and enhanced SN-38 recovery (f = 1.86). The extravasation of HPBCD in the tumor during microdialysis was lower than 1%. Vu,tumor values were above 3 mL/g tumor for both neuroblastoma models and suggested efficient cellular penetration of SN-38. CONCLUSIONS: The method contributes to overcome the limitations of the microdialysis technique in hydrophobic drugs and provides a powerful tool to characterize compartmental anticancer drug distribution in xenografts.


Assuntos
Antineoplásicos/metabolismo , Xenoenxertos/metabolismo , Neuroblastoma/metabolismo , 2-Hidroxipropil-beta-Ciclodextrina , Animais , Antineoplásicos/farmacologia , Camptotecina/análogos & derivados , Camptotecina/metabolismo , Camptotecina/farmacologia , Humanos , Interações Hidrofóbicas e Hidrofílicas , Irinotecano , Camundongos , Camundongos Nus , Microdiálise/métodos , Neuroblastoma/tratamento farmacológico , beta-Ciclodextrinas/metabolismo , beta-Ciclodextrinas/farmacologia
17.
Proc Natl Acad Sci U S A ; 111(51): E5564-73, 2014 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-25512523

RESUMO

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.


Assuntos
Neoplasias Ósseas/metabolismo , Genoma Humano , Osteossarcoma/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Heterogeneidade Genética , Mutação em Linhagem Germinativa , Humanos , Osteossarcoma/genética , Osteossarcoma/patologia , Proteína Supressora de Tumor p53/genética
18.
Cancer Discov ; 4(11): 1326-41, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25186949

RESUMO

UNLABELLED: Pediatric Ewing sarcoma is characterized by the expression of chimeric fusions of EWS and ETS family transcription factors, representing a paradigm for studying cancers driven by transcription factor rearrangements. In this study, we describe the somatic landscape of pediatric Ewing sarcoma. These tumors are among the most genetically normal cancers characterized to date, with only EWS-ETS rearrangements identified in the majority of tumors. STAG2 loss, however, is present in more than 15% of Ewing sarcoma tumors; occurs by point mutation, rearrangement, and likely nongenetic mechanisms; and is associated with disease dissemination. Perhaps the most striking finding is the paucity of mutations in immediately targetable signal transduction pathways, highlighting the need for new therapeutic approaches to target EWS-ETS fusions in this disease. SIGNIFICANCE: We performed next-generation sequencing of Ewing sarcoma, a pediatric cancer involving bone, characterized by expression of EWS-ETS fusions. We found remarkably few mutations. However, we discovered that loss of STAG2 expression occurs in 15% of tumors and is associated with metastatic disease, suggesting a potential genetic vulnerability in Ewing sarcoma.


Assuntos
Antígenos Nucleares/genética , Neoplasias Ósseas/genética , Sarcoma de Ewing/genética , Antígenos Nucleares/metabolismo , Neoplasias Ósseas/metabolismo , Linhagem Celular Tumoral , Criança , DNA de Neoplasias/genética , Feminino , Rearranjo Gênico , Genômica , Humanos , Masculino , Mutação , Sarcoma de Ewing/metabolismo , Análise de Sequência de DNA
19.
Dev Neurobiol ; 73(11): 815-27, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23776185

RESUMO

Neuroblastoma, the most common extracranial tumor in children, is caused by genetic lesions in neural crest precursors of the peripheral nervous system. However, since neural crest cells are neither present after birth and nor are they readily accessible for analysis, very little is known about the genetic networks they might share with neuroblastoma cells during their development, despite their common embryonic origin. Here we have developed a novel resource for lineage tracing and for the isolation of neural crest cells in the chick embryo, enabling us to perform a genome-wide expression screen in neural crest progenitors. In this analysis, we efficiently retrieved known neural crest specific genes that validate our screening strategy and we identified new genes that participate in diverse cell activities, yet with a strong representation of genes associated to cell signaling and cell mobility, two hallmarks of migratory cells. We crossed this transcriptome data with that in the neuroblastoma gene server to search for the human orthologues of these genes associated with neuroblastoma. Accordingly, we retrieved 54 genes expressed strongly in both populations, from which we were able to validate a total of 27 genes expressed in the neural crest that are relevant to neuroblastoma formation. We propose that neural crest and neuroblastoma tumor cells share a common genetic signature that might serve to characterize neuroblastoma cancer stem cells, thereby contributing to the identification of specific targets against which new therapeutic strategies can be designed.


Assuntos
Crista Neural/citologia , Neuroblastoma/genética , Transcriptoma , Animais , Embrião de Galinha , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Hibridização In Situ , Análise de Sequência com Séries de Oligonucleotídeos
20.
PLoS One ; 8(3): e59762, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23533647

RESUMO

BACKGROUND: Neuroblastic tumors include the neuroblastomas, ganglioneuroblastomas, and ganglioneuromas. Clinical behavior of these developmental malignancies varies from regression to aggressive growth with metastatic dissemination. Several clinical, histological, genetic, and biological features are associated with this diversity of clinical presentations. The calcium-sensing receptor (CaSR) is a G-protein coupled receptor with a key role in calcium homeostasis. We have previously reported that it is expressed in benign, differentiated neuroblastic tumors, but silenced by genetic and epigenetic events in unfavorable neuroblastomas. We have now analyzed three functionally relevant polymorphisms clustered at the signal transduction region of the CaSR (rs1801725, rs1042636 and rs1801726) to assess if genetic variants producing a less active receptor are associated with more aggressive disease course. METHODS: Polymorphisms were analyzed in DNA samples from 65 patients using specific Taqman Genotyping Assays. RESULTS: Mildly inactivating variant rs1801725 was associated with clinical stage 4 (P = 0.002) and the histological subgroup of undifferentiated neuroblastomas (P = 0.046). Patients harboring this polymorphism had significantly lower overall (P = 0.022) and event-free survival (P = 0.01) rates than those who were homozygous for the most common allele among Caucasians. However, this single locus genotype was not independently associated with outcome in multivariate analyses. Conversely, the tri-locus haplotype TAC was independently associated with an increased risk of death in the entire cohort (Hazard Ratio = 2.45; 95% Confidence Interval [1.14-5.29]; P = 0.022) and also in patients diagnosed with neuroblastomas (Hazard Ratio = 2.74; 95% Confidence Interval [1.20-6.25]; P = 0.016). CONCLUSIONS: The TAC haplotype includes the moderately inactivating variant rs1801725 and absence of the gain-of-function rs1042636 polymorphism. Thus, its association with metastatic disease and poor outcome would add to our previous data and further support that inactivation of the CaSR gene is a mechanism associated with neuroblastoma malignant behavior.


Assuntos
Neuroblastoma/genética , Polimorfismo Genético/genética , Receptores de Detecção de Cálcio/genética , Adolescente , Adulto , Feminino , Genótipo , Haplótipos , Humanos , Masculino , Neuroblastoma/patologia , Adulto Jovem
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