Array CGH of breast cancer cells with distinct metastatic properties
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Xiang Zhang
PROVIDER: E-GEOD-23823 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Using array comparative genomic hybridization (aCGH), a large number of deleted genomic regions have been identified in human cancers. However, subsequent efforts to identify target genes selected for inactivation in these regions have often been challenging. We integrated here genome-wide copy number data with gene expression data and non-sense mediated mRNA decay rates in breast cancer cell lines to prioritize gene candidates that are likely to be tumour suppressor genes inactivated by bi-allelic genetic events. The candidates were sequenced to identify potential mutations. This integrated genomic approach led to the identification of RIC8A at 11p15 as a putative candidate target gene for the genomic deletion in the ZR-75-1 breast cancer cell line. We identified a truncating mutation in this cell line, leading to loss of expression and rapid decay of the transcript. We screened 127 breast cancers for RIC8A mutations, but did not find any pathogenic mutations. No promoter hypermethylation in these tumours was detected either. However, analysis of gene expression data from breast tumours identified a small group of aggressive tumours that displayed low levels of RIC8A transcripts. Real-time PCR analysis of 38 breast tumours showed a strong association between low RIC8A expression and the presence of TP53 mutations (P=0.006). We demonstrate a data integration strategy leading to the identification of RIC8A as a gene undergoing a classical double-hit genetic inactivation in a breast cancer cell line, as well as in vivo evidence of loss of RIC8A expression in a subgroup of aggressive TP53 mutant breast cancers. Gene expression data: Samples GSM388181-GSM388198. The experiment utilized six breast cancer cell lines: MDA-MB-468, MDA-MB-231, ZR-75-1, MCF7, BT-474 and T-47D, and three non-malignant cell lines: HMECs (non-malignant human mammary epithelial cells), IMR90 (normal lung fibroblasts) and WS1 (normal skin fibroblasts). All cell lines were obtained from American Type Culture Collection and grown in accordance with the distributor's instructions. Both malignant and non-malignant cell lines were treated with the translation inhibitor emetine dihydrochloride hydrate. For each cell line, parallel cell cultures were grown in 175 cm2 flasks until 70-80 % confluence. Half of the subconfluent cultures were treated with 100 μg ml-1 of emetine dihydrochloride hydrate while the other half were left as untreated controls. Genome-wide copy number data: Samples GSM388211-GSM388216. The experiment utilized six breast cancer cell lines; MDA-MB-468, MDA-MB-231, ZR-75-1, MCF7, BT-474 and T-47D. All cell lines were obtained from American Type Culture Collection and grown in accordance with the distributor's instructions. All samples were hybridized once on 44k Agilent Human Genome CGH microarrays according to manufacturers instructions. Genomic DNA pooled from healthy female donors was used as a reference in all hybridizations. DNA from cell line samples were labeled with Cy5 and DNA from reference were labeled with Cy3.
2010-06-19 | E-GEOD-15477 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the following subset Series: GSE18139: Array-based gene expression in neuroblastic tumors GSE18143: aCGH of neuroblastic tumors Refer to individual Series
2010-05-05 | E-GEOD-18144 | biostudies-arrayexpress
Project description:Thirteen HER2 positive breast cancer cell lines were screened with 22 commercially available compounds, mainly targeting proteins in the ErbB2 signaling pathway, and the molecular mechanisms related to treatment response were sought. To search for response predictors, genomic and transcriptomic profiling, PIK3CA mutations and PTEN status were associated to the drug responses and several genes involved in the response of the compounds were identified. Array-CGH experiments of HER2+ breast cancer cell lines grown under standard conditions. DNA from four HER2 positive breast cancer cell lines was isolated and hybridized on Agilent arrays.
2015-01-30 | E-GEOD-58886 | biostudies-arrayexpress
Project description:Title: Array-based gene expression, CGH and tissue data define a 12q24 gain in neuroblastic tumors with prognostic implication. Background: Neuroblastoma has successfully served as a model system for the identification of neuroectoderm-derived oncogenes. However, in spite of various efforts, only a few clinically useful prognostic markers have been found. Here, we present a framework, which integrates DNA, RNA and tissue data to identify and prioritize genetic events that represent clinically relevant new therapeutic targets and prognostic biomarkers for neuroblastoma. Methods: A single-gene resolution aCGH profiling was integrated with microarray-based gene expression profiling data to distinguish genetic copy number alterations that were strongly associated with transcriptional changes in two neuroblastoma cell lines. FISH analysis using a hotspot tumour tissue microarray of 37 paraffin-embedded neuroblastoma samples and in silico data mining for gene expression information obtained from previously published studies including up to 445 healthy nervous system samples and 123 neuroblastoma samples were used to evaluate the clinical significance and transcriptional consequences of the detected alterations and to identify subsequently activated gene(s). Results: In addition to the anticipated high-level amplification and subsequent overexpression of MYCN, MEIS1, CDK4 and MDM2 oncogenes, the aCGH analysis revealed numerous other genetic alterations, including microamplifications at 2p and 12q24.11. Most interestingly, we identified and investigated the clinical relevance of a previously poorly characterized amplicon at 12q24.31. FISH analysis showed low-level gain of 12q24.31 in 14 of 33 (42%) neuroblastomas. Patients with the low-level gain had an intermediate prognosis in comparison to patients with MYCN amplification (poor prognosis) and to those with no MYCN amplification or 12q24.31 gain (good prognosis) (P = 0.001). Using the in silico data mining approach, we identified elevated expression of five genes located at the 12q24.31 amplicon in neuroblastoma (DIABLO, ZCCHC8, RSRC2, KNTC1 and MPHOSPH9). Among these, DIABLO showed the strongest activation suggesting a putative role in neuroblastoma progression. Conclusions: The presented systematic and rapid framework, which integrates aCGH, gene expression and tissue data to obtain novel targets and biomarkers for cancer, identified a low-level gain of the 12q24.31 as a potential new biomarker for neuroblastoma progression. Furthermore, results of in silico data mining suggest a new neuroblastoma target gene, DIABLO, within this region, whose functional and therapeutic role remains to be elucidated in follow-up studies. High-resolution aCGH was utilized to identify novel genetic alterations in two neuroblastoma cell lines, NGP and IMR-32. Through the integration of gene copy number and gene expression data, the impact of copy number changes on expression levels was determined. Fluorescence in situ hybridization (FISH) on a tissue microarray (TMA) format was used to assess the clinical significance of the identified copy number increase at 12q24.31 in neuroblastoma patients. Finally, we used in silico data mining of publicly available transcriptomics data, to evaluate the transcriptional consequences of the detected 12q24.31 alteration and to identify subsequently activated gene(s). Here, aCGH of the NGP and IMR-32 cell lines was performed. Male genomic DNA was used as reference.
2010-05-05 | E-GEOD-18143 | biostudies-arrayexpress
Project description:Purpose: Preclinical model systems should faithfully reflect the complexity of the human pathology. In hepatocellular carcinoma (HCC), the tumor vasculature is of particular interest in diagnosis and therapy. By comparing two commonly applied preclinical model systems, diethylnitrosamine induced (DEN) and orthotopically implanted (McA) rat HCC, we aimed to measure tumor biology non-invasively and identify differences between the models.<br>Experimental design: DEN and McA tumor development was monitored by magnetic resonance imaging (MRI) and positron emission tomography (PET). A slice-based correlation of imaging and histopathology was performed. Array CGH analyses were applied to determine genetic heterogeneity. Therapy response to sorafenib was tested in DEN and McA tumors.<br>Results: Histologically and biochemically confirmed liver damage resulted in increased 18F-fluordeoxyglucose (FDG) PET uptake and perfusion in DEN animals only. DEN tumors exhibited G1-3 grading compared to uniform G3 grading of McA tumors. Array comparative genomic hybridization revealed a highly variable chromosomal aberration pattern in DEN tumors. Heterogeneity of DEN tumors was reflected in more variable imaging parameter values. DEN tumors exhibited lower mean growth rates and FDG uptake and higher diffusion and perfusion values compared to McA tumors. To test the significance of these differences, the multikinase inhibitor sorafenib was administered, resulting in reduced volume growth kinetics and perfusion in the DEN group only.<br>Conclusion: This work depicts the feasibility and importance of in depth preclinical tumor model characterization and suggests the DEN model as a promising model system of multifocal nodular HCC in future therapy studies.
2016-03-25 | E-MTAB-3507 | biostudies-arrayexpress
Project description:Multiple commercial microarrays for measuring genome-wide gene expression levels are currently available, including oligonucleotide and cDNA, single- and two-channel formats. This study reports on the results of gene expression measurements generated from identical RNA preparations that were obtained using three commercially available microarray platforms. RNA was collected from PANC-1 cells grown in serum-rich medium and at 24 h following the removal of serum. Three biological replicates were prepared for each condition, and three experimental replicates were produced for the first biological replicate. RNA was labeled and hybridized to microarrays from three major suppliers according to manufacturers’ protocols, and gene expression measurements were obtained using each platform’s standard software. For each platform, gene targets from a subset of 2009 common genes were compared. Correlations in gene expression levels and comparisons for significant gene expression changes in this subset were calculated, and showed considerable divergence across the different platforms, suggesting the need for establishing industrial manufacturing standards, and further independent and thorough validation of the technology. Keywords: Differentiation PANC-1 cells were grown in serum-containing media followed by exposure to serum-free media for 24 hours. In both groups, RNA extracted from the first biological replicate was derived from 3 culture dishes producing enough sample for three technical replicates, whereas RNA from the second and third biological replicates were derived from one culture dish each.
2007-05-15 | E-GEOD-7785 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2015-01-30 | E-GEOD-58887 | biostudies-arrayexpress
Project description:Purpose. Simian virus 40 (SV40)-immortalized human corneal epithelial (HCE-T) cells have been widely used as an in vitro model of human corneal epithelial cells. To better understand the nature of this cell line, we assessed it for genomic aberrations and cellular heterogeneity. Methods. For the quantitative measurement of genomic aberrations, array based comparative genomic hybridization (CGH) analysis was performed. For identification of cellular heterogeneity, cell morphology, growth kinetics, trans-epithelial electrical resistance and transcriptional efficiency were analyzed. We also carried out real-time PCR and chromosomal fluorescent in situ hybridization (cFISH) against some gained or lost loci in order to assess genomic heterogeneity. To assess differences in the gene expression profiles between HCE-T cells and normal corneal epithelial cells, we collected expressed sequence tags (ESTs) for this cell line. Southern blotting and inverse PCR analyses were used to determine the genomic integration site of the SV40 large T antigen gene (LTAG). Results. Array CGH analysis indicated that the genomic content of HCE-T cells is different from the normal healthy genome. Our results from cellular functional assays, real-time PCR and cFISH strongly showed that HCE-T cells consist of a not insignificant number of heterogeneous cell populations. The genomic integration site of the SV40 large T antigen was at p22.1 of chromosome 9. Conclusions. Our results indicate that HCE-T cells have an altered genomic content and that they are composed of heterogeneous cell populations. This should be considered when conducting experiments or interpreting the results of studies which use this cell line. Keywords: comparative genomic hybridization Genomic DNAs from HCE-T cells, immortalized cell line of human corneal epithelial cells and normal female duploid cells were subjected to array CGH analysis to identify genomic alterations in the HCE-T cells. These DNAs were labeled with Cy5 and Cy3, respectively.
2008-04-15 | E-GEOD-11101 | biostudies-arrayexpress
Project description:Understanding the genetic architecture of cancer pathways that distinguishes subsets of human cancer is critical to developing new therapies that better target tumors based upon their molecular expression profiles. In this study, we identify an integrated gene signature from multiple transgenic models of epithelial cancers intrinsic to the functions of the Simean virus 40 T/t-antigens that is associated with the biologic behavior and prognosis for several human epithelial tumors. This genetic signature, composed primarily of genes regulating cell replication, proliferation, DNA repair and apoptosis, is not a general cancer signature. Rather, it is uniquely activated primarily in tumors with aberrant p53, Rb or BRCA1 expression, but not in tumors initiated through the overexpression of myc, ras, her2/neu, or Polyoma middle T oncogenes. Importantly, human breast, lung and prostate tumors expressing this set of genes represent subsets of tumors with the most aggressive phenotype and with poor prognosis. The T/t-antigen signature is highly predictive of human breast cancer prognosis. Since this class of epithelial tumors is generally intractable to currently existing standard therapies, this genetic signature identifies potential targets for novel therapies directed against these lethal forms of cancer. Since the these genetic targets have been discovered using mammary, prostate, and lung T/t-antigen mouse cancer models, these models are rationale candidates for use in pre-clinical testing of therapies focused on these biologically important targets. Keywords: SuperSeries Gene expression profiles from the SV40 T/t-antigen mouse models were compared with respect to specimen type (normal vs. tumor tissue), location of tumor (mammary, lung, prostate, seminal vesicle), and background strain of mice (FVB vs. C57BL/6). A three-way ANOVA model with one interaction effect (type X location) was fitted. Cancer genes that differ among all four tumor locations were identified, as those that had a significant interaction effect at the 0.001 level and showed at least a 2-fold change between the maximal and minimal mean tumor/normal ratio over the different locations (2638 cDNA probes). Based on the ANOVA model, differentially expressed genes between normal and tumor specimens within each tumor location were also identified, as those genes whose expression were significant at the 0.001 level and were at least 2-fold different compared to the mean expression ratio. Overall 3004 unique array features were selected using ANOVA. Further selection was applied based upon identification of differentially expressed genes between normal and tumor tissue for the three epithelial tumors (mammary, lung and prostate). The SV40 T/t-antigen oncogene-specific signature included genes similarly differentially expressed in each epithelial tumor (153 cDNA clones). In contrast, genes were included in a tissue-specific SV40 T/t-antigen tumor signature if they were found to be differentially expressed between the tumor and normal samples exclusively for one location. Two-hundred and eighty three, 220 and 999 cDNA clones were identified as specifically dysregulated in mammary, lung and prostate tumors, respectively).
2007-09-01 | E-GEOD-8666 | biostudies-arrayexpress
Project description:Affymetrix gene expression analysis was carried out to investigate the differences in gene profile of MBP89-101-reactive encephalitogenic T cells before and after co-culture with cerebellar granular neurons (CGNs). Co-culture of MBP89-101-reactive encephalitogenic T cells with CGNs leads to generation of T cells with regulatory T cells phenotype (CD4+CD25+membrane bound TGF-b+ T cells) or a new regulatory phenotype (CD4highPD-L1high T cells). CGN-induced CD4+CD25+membrane bound TGF-b+ T regulatory cells, CD4highPD-L1high T cells were purified by FACSAria. IFN-beta induced T lymphocytes (CD4highPD-L1high T cells) were also FACSAria purified. All these populations were compared to MBP89-101-reactive encephalitogenic T cells. Samples were prepared from biological triplicates for each FACSAria sorted population. There are four different cell types, as determined by cell surface markers, in triplicate in the experiment.
2014-02-16 | E-GEOD-54490 | biostudies-arrayexpress