RESUMEN
Using single nucleotide polymorphic (SNP) allele arrays, we analyzed 28 pediatric gliomas consisting of 14 high-grade gliomas and 14 low-grade gliomas. Most of the low-grade gliomas had no detectable loss of heterozygosity (LOH) in any of the 11,562 SNP loci; exceptions were two gangliogliomas (3q and 9p), one astrocytoma (6q), and two subependymal giant cell astrocytomas (16p and 21q). On the other hand, all high-grade gliomas had various degrees of LOH affecting 52 to 2,168 SNP loci on various chromosomes. LOH occurred most frequently in regions located at 4q (54%), 6q (46%), 9p (38%), 10q (38%), 11p (38%), 12 (38%), 13q (69%), 14q (54%), 17 (38%), 18p (46%), and 19q (38%). We also detected amplifications of epidermal growth factor receptor (EGFR) or platelet-derived growth factor receptor alpha (PDGFRalpha) in a few of the 13 cases of glioblastoma multiforme analyzed. Interestingly, the amplified EGFR and PDGFRalpha were located within regions of LOH. SNP loci with LOH and copy number changes were validated by sequencing and quantitative PCR, respectively. Our results indicate that, in some pediatric glioblastoma multiforme, one allele each of EGFR and PDGFRalpha was lost but the remaining allele was amplified. This may represent a new molecular mechanism underlying tumor progression.
Asunto(s)
Genoma Humano/genética , Glioma/patología , Pérdida de Heterocigocidad , Polimorfismo de Nucleótido Simple/genética , Alelos , Secuencia de Bases , Ciclo Celular/genética , Niño , Análisis Mutacional de ADN , Receptores ErbB/genética , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica/genética , Genotipo , Glioblastoma/genética , Glioblastoma/patología , Glioma/genética , Humanos , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/genética , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Juvenile pilocytic astrocytoma (JPA) is one of the most common brain tumors in children. The expression profiles of 21 JPAs, determined using Affymetrix GeneChip U133A, were compared with subjects with normal cerebella. The genes involved in neurogenesis, cell adhesion, synaptic transmission, central nervous system development, potassium ion transport, protein dephosphorylation, and cell differentiation were found to be significantly deregulated in JPA. These 21 JPAs were further clustered into two major groups by unsupervised hierarchical clustering using a set of 848 genes with high covariance (0.5-10). Supervised analysis with Significance Analysis of Microarrays software between these two potential subgroups identified a list of significant differentially expressed genes involved in cell adhesion, regulation of cell growth, cell motility, nerve ensheathment, and angiogenesis. Immunostaining of myelin basic protein on paraffin sections derived from 18 incompletely resected JPAs suggests that JPA without myelin basic protein-positively stained tumor cells may have a higher tendency to progress.
Asunto(s)
Astrocitoma/genética , Neoplasias Encefálicas/genética , Adolescente , Astrocitoma/clasificación , Secuencia de Bases , Neoplasias Encefálicas/clasificación , Niño , Preescolar , Cartilla de ADN , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Besides their use in mRNA expression profiling, oligonucleotide microarrays have also been applied to single-nucleotide polymorphism (SNP) and loss of heterozygosity (LOH) or allelic imbalance studies. In this report, we evaluate the reliability of using whole genome amplified DNA for analysis with an oligonucleotide microarray containing 11 560 SNPs to detect allelic imbalance and chromosomal copy number abnormalities. Whole genome SNP analyses were performed with DNA extracted from osteosarcoma tissues and patient-matched blood. SNP calls were then generated by Affymetrix GeneChip DNA Analysis Software. In two osteosarcoma cases, using unamplified DNA, we identified 793 and 1070 SNP loci with allelic imbalance, respectively. In a parallel experiment with amplified DNA, 78% and 83% of these SNP loci with allelic imbalance was detected. The average false-positive rate is 13.8%. Furthermore, using the Affymetrix GeneChip Chromosome Copy Number Tool to analyze the SNP array data, we were able to detect identical chromosomal regions with gain or loss in both amplified and unamplified DNA at cytoband resolution.
Asunto(s)
Genoma Humano , Pérdida de Heterocigocidad/genética , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Osteosarcoma/genética , Polimorfismo de Nucleótido Simple/genética , Cromosomas Humanos Par 6/genética , Reacciones Falso Positivas , Genómica/métodos , Humanos , Repeticiones de Microsatélite/genética , Técnicas de Amplificación de Ácido Nucleico , Sensibilidad y EspecificidadRESUMEN
Gene amplifications have been observed in many different tumor cells, and many of these changes are related to tumor pathogenesis. Comparative genomic hybridization (CGH) using metaphase chromosomes can detect changes in chromosome copy number with a resolution of 10-20 Mb. Current advances in CGH analysis in a microarray format allow us to refine such changes down to the gene level. We applied microarray technology to detect novel gene amplification in a malignant mixed tumor of salivary gland. Besides detecting previously known gene amplifications (MDM2 and MYC), we identified four other highly amplified genes located at 8q11.2 approximately q13: MGC2177, PLAG1, PSMC6P, and LYN. The amplification was further validated with real-time quantitative polymerase chain reaction.