Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of human short term cultured glioblastoma cell lines treated with 5-aza-2-deoxycitidine to investigate if epigenetic alterations are an alternative means of glioblastoma gene inactivation

ABSTRACT: Glioblastoma, the most aggressive and least treatable form of malignant glioma, is the most common human brain tumor. Although many regions of allelic loss occur in glioblastomas, relatively few tumor suppressor genes have been found mutated at such loci. To address the possibility that epigenetic alterations are an alternative means of glioblastoma gene inactivation, we coupled pharmacological manipulation of methylation with gene profiling to identify potential methylation-regulated, tumor-related genes. Triplicates of three short-term cultured glioblastomas were exposed to 5?M 5-aza-dC for 96 hours followed by cRNA hybridization to an oligonucleotide microarray (Affymetrix U133A). We based candidate gene selection on bioinformatics, RT-PCR, bisulfite sequencing, methylation-specific PCR and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Two genes identified in this manner, RUNX3 and Testin (TES), were subsequently shown to harbor frequent tumor-specific epigenetic alterations in primary glioblastomas. This overall approach therefore provides a powerful means to identify candidate tumor suppressor genes for subsequent evaluation and may lead to the identification of genes whose epigenetic dysregulation is integral to glioblastoma tumorigenesis. Duplicates of three short term cultured glioblastoma cell lines (internal IDs: GLI56;GLI60;GLI72) were either exposed to 5umol 5`aza-dC for 96h or left untreated. Total RNA was extracted of treated and untreated cells after 96h and hybridized to U133A chip. Gene expression profiles of treated and untreated cells were compared. Upregulation of gene expression in the treated (demethylated) samples was interpreted as potentially being regulated by methylation - pointing towards hypermethylation in the related cell line. Identification of novel tumor suppressor genes, regulated by methylation, was the overall goal.

ORGANISM(S): Homo sapiens

SUBMITTER: Wolf Mueller

PROVIDER: E-GEOD-4717 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Downregulation of RUNX3 and TES by hypermethylation in glioblastoma.

Mueller W W Nutt C L CL Ehrich M M Riemenschneider M J MJ von Deimling A A van den Boom D D Louis D N DN

Oncogene 20060814 4

Glioblastoma, the most aggressive and least treatable form of malignant glioma, is the most common human brain tumor. Although many regions of allelic loss occur in glioblastomas, relatively few tumor suppressor genes have been found mutated at such loci. To address the possibility that epigenetic alterations are an alternative means of glioblastoma gene inactivation, we coupled pharmacological manipulation of methylation with gene profiling to identify potential methylation-regulated, tumor-rel ...[more]

PMID: 16909125

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Project description:Epigenetic changes largely contribute to the regulation of gene expression in cancer cells. DNA methylation is part of the epigenetic gene regulation complex which is relevant for the pathogenesis of cancer. We performed a genome-wide search for methylated CpG islands in tumors and corresponding non-malignant lung tissue samples of 101 stage I-III non-small cell lung cancer (NSCLC) patients by combining methylated DNA immunoprecipitation and microarray analysis using NimbleGenM-BM-4s 385K Human CpG Island plus Promoter arrays. By testing for differences in methylation between tumors and corresponding non-malignant lung tissues, we identified 298 tumor-specifically methylated genes. From many of these genes epigenetic regulation was unknown so far. Gene Ontology analysis revealed an over-representation of genes involved in regulation of gene expression and cell adhesion. Expression of 182 of 298 genes was found to be upregulated after 5-aza-2M-BM-4-deoxycytidine (Aza-dC) and/or trichostatin A (TSA) treatment of 3 NSCLC cell lines by Affymetrix microarray analysis. In addition, methylation of selected genes in primary NSCLCs and corresponding non-malignant lung tissue samples were analyzed by methylation-sensitive high resolution melting analysis (MS-HRM). Our results obtained by MS-HRM analysis confirmed our data obtained by MeDIP-chip analysis. Moreover, by comparing methylation results from MeDIP-chip analysis with clinico-pathological parameters of the patients we observed methylation of HOXA2 as potential parameter for shorter disease-free survival of NSCLC patients. In conclusion, using a genome-wide approach we identified a large number of tumor-specifically methylated genes in NSCLC patients. Our results stress the importance of DNA methylation for the pathogenesis of NSCLCs. Overall, samples of 3 untreated, with Aza-dC treated and with Aza-dC/TSA treated NSCLC cell lines were hybridized to Affymetrix HG-U133_plus_2.0 microarrays (18 in total).

Dataset Information

Transcription profiling of human short term cultured glioblastoma cell lines treated with 5-aza-2-deoxycitidine to investigate if epigenetic alterations are an alternative means of glioblastoma gene inactivation

Publications

Downregulation of RUNX3 and TES by hypermethylation in glioblastoma.

Similar Datasets

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