Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Genome-Wide Analysis of Somatic Copy Number Alterations (CGH, SNP) and Gene Expression (RNA-seq) in Metastatic Melanoma

ABSTRACT: All cancers are diseases of the genome. A combination of somatic point mutations, focal amplifications and deletions, and chromosome level aberrations conspire to disrupt gene expression and the interplay between signaling pathways that control normal growth and tissue homeostasis. Here we investigate somatic copy number abberations in metastatic melanomas. A metastatic melanoma was assayed on Affymetrix SNP arrays to detect copy number abberations. 7 cutaneous melanomas as well as their matched control ( peripheral blood lymphocytes (PBL) or Epstein-Barr virus transformed lymphoblastoid cell lines ) and 2 control melanocytes were assayed on Illumina SNP arrays. 7 metastatic melanomas were hybridized on Agilent CGH arrays using donor matched control as reference.

ORGANISM(S): Homo sapiens

SUBMITTER: Armand Valsesia

PROVIDER: E-GEOD-22928 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Network-guided analysis of genes with altered somatic copy number and gene expression reveals pathways commonly perturbed in metastatic melanoma.

Valsesia Armand A Rimoldi Donata D Martinet Danielle D Ibberson Mark M Benaglio Paola P Quadroni Manfredo M Waridel Patrice P Gaillard Muriel M Pidoux Mireille M Rapin Blandine B Rivolta Carlo C Xenarios Ioannis I Simpson Andrew J G AJ Antonarakis Stylianos E SE Beckmann Jacques S JS Jongeneel C Victor CV Iseli Christian C Stevenson Brian J BJ

PloS one 20110408 4

Cancer genomes frequently contain somatic copy number alterations (SCNA) that can significantly perturb the expression level of affected genes and thus disrupt pathways controlling normal growth. In melanoma, many studies have focussed on the copy number and gene expression levels of the BRAF, PTEN and MITF genes, but little has been done to identify new genes using these parameters at the genome-wide scale. Using karyotyping, SNP and CGH arrays, and RNA-seq, we have identified SCNA affecting ge ...[more]

PMID: 21494657

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Project description:Circulating tumor cells (CTCs) are critical in the development of distant organ tumor metastasis, and are associated with advanced cancer stage and poor patient outcome. Here, we present the first genome-wide nucleotide-level characterization of CTCs. Our single-nucleotide polymorphism (SNP) analysis in patients with melanoma involved: 1) global comparative genomic analysis of CTCs and matched regional metastases, 2) identification of key genomic aberrations in CTCs, 3) verification of these target genes in aggressive distant tumor metastases, and 4) evidence of selective expression and functional consequence of CTC-associated genes in melanomas. We report 131 aberrant loci in CTCs that are potentially pro-metastatic, and show that such expression of a 5-marker gene panel (CSMD2, CNTNAP5, FLJ14051, ADAM6, TRPM2) in melanomas confers prognostic utility. Successful treatment of melanoma requires understanding of the metastatic process and identification of patients with tumors most likely to develop aggressive metastatic disease. Melanomas are heterogeneous, and CTCs have long been recognized as vehicles for cancer spread, representing particularly aggressive tumor clones that can evolve into successful clinical metastases. Elucidation of genomic aberrations in CTCs will aid in the development of prognostic biomarkers and therapeutic strategies to target CTCs to prevent or control distant cancer spread. This study provides the first detailed genomic confirmation of the close relation between CTCs and tumor metastases, and illustrates how CTCs can be utilized as a novel approach and rational source for identification of pro-metastatic genes in cancer research. Three individual patient cohorts were utilized in the study. CNV and LOH loci were evaluated initially in metastatic melanoma patients (n=13) in a discovery cohort. SNP loci that harbored CNV/LOH in CTCs were then separately verified for: (a) presence in distant organ metastasis (AJCC Stage IV melanoma) (n=27), and (b) relevance to prognosis in regional melanoma metastasis (AJCC Stage III melanoma) (n=35). The first discovery patient cohort group was utilized for capture of CTCs, and consisted of peripheral blood mononuclear cell (PBMC) and tumor specimens from metastatic melanoma patients (n=13). CTC-related loci were verified in a second cohort of patients with Stage IV distant organ metastases (n=27), including 15 brain, 4 lung, and 8 gastrointestinal (bowel, liver) metastases. The third patient cohort consisted of early passage (<12) established melanoma cell lines derived from 35 regional melanoma metastases (Stage III) for evaluation of the prognostic utility of the CTC-associated aberrant loci.

Project description:Development of a new carcinoma cell line (HC-AFW1) derived from a pediatric liver tumor The cell line HC-AFW1 was derived from a 4 year old boy suffering from HCC through culturing and passage into immuno-deficient mice. The cell line is stable now for over 8 months of culture with a doubling time of 40 h. The tumor cells show an epithelial histology and express hepatoma proteins such as Alpha-fetoprotein (AFP), Glypican 3, E-cadherin, CD10, CD326, HepPar1, Vimentin and Desmin. Catenin beta shows a deletion of 49 amino acids in the exon 3 involving the phosphorylation sites of GSK3 and is detectable in the cell nuclei. Cytogenetic analysis revealed large anomalies in the chromosomal map including chromosomal aberrations found in hepatoblastoma as well as in adult HCC. Copy number analysis of two different passages of HC-AFW1 cells Two tumor specimens were used for tissue culture and xenotransplantation into NSG mice. Tumor cells derived from xenografts were cultured and designated as HC-AFW1. DNA was analyzed from the passage 2 (2P2). HC-AFW2 was derived from tissue culture without transfer in to mice. Primary tissue samples were minced into pieces of 3x3 mm and cultured on 6 well plates (Becton Dickenson, Frankfurt, Germany) in DMEM (GIBCO BRL, Carlsbad, CA) supplemented with 10% FCS (growth medium). Cell cultures were maintained in a humidified atmosphere containing 5% CO2 at 37°C. For subculturing cells were detached from the culture surface using accutase in Dulbecco´s PBS containing 0.5mM EDTA (PAA Laboratories GmbH, Cölbe, Germany) for 2-3 minutes at 37°C. A sub-cultivation ratio of 1:4 and 1:6 was performed twice per week. Cells were stored in liquid nitrogen as a suspension in complete growth medium with 10% DMSO. DNA from patient blood and tumor samples was isolated with the QiaAmp DNA mini Kit according to manufacturer's instructions (Qiagen, Hilden, Germany). Single nucleotide polymorphism (SNP) and copy number polymorphism (CNP) genotyping were performed using the Genome-Wide Human SNP Array 6.0 and Genotyping Console (GTC) software (Affymetrix, Santa Clara, CA).

Project description:Human pluripotent stem cells commonly undergo adaptive changes during prolonged passaging in vitro. We have developed a new laminin-521 based method for self-renewal of human pluripotent stem cells. SNP6.0 array was used to assess genetic stability of several human ES cell lines cultured in the new system with two lines (H1 and HS401) cultured on feeders (standard conditions) Two main objectives in this study: To compare the number of the CNVs in human ES cells at early and late passages; for example, samples HS980 p6 and HS980 p31 represent HS980 cells at passages 6 and 31, respectively. The overall conclusion is that the observed number of CNVs is similar in the cells at early and late passages. This analysis can be remade without the reference samples. To compare the number of CNVs in control human ES cells (H1 and HS401) grown under standard protocol with that in cells (HS980, HS983a, HS999, HS916, and HS1001) grown under a new, developed by us, protocol. Because the cells were from different individuals, we could not compare them directly. Instead, we compared them to a set of samples from 75 healthy individuals, and then with each other. For the Reference These samples had been hybridized in the same lab as our samples, but for a different project; and the data were kindly provided by Prof. Kere. However, the same overall conclusion would result from an other sufficiently large dataset of healthy human individuals. Please note that 75 healthy individual samples had been hybridized in the same lab as our samples and the data were kindly provided by Prof. Kere, which is linked as a Series supplementary file ('75_samples_ref.REF'). It is a Reference Model File that Affymetrix Genotyping Console software has created based on data from 75 healthy individuals (i.e. processed file based on the CEL files of the 75 healthy individuals). The file derives from the "CN/LOH Reference Model File Creation and Analysis (Batch Sample Mode)" process according to the Genotyping Console manual. It contains SNP-wise information on the observed distribution of hybridization intensities of a set of samples that are assumed to be diploid in copy number (for most part of their genomes at least). This file is then used to compare the intensities observed in the CEL files of interest to infer deviations from the reference as CNVs. Thus, in combination with the CEL files (from each ES cell line), this file enables the exact replication of the CNV analyses done for the various human ES cell lines study. Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted cultured human ES cells. Several lines were analysed at different time points of the experiment. The analysis was done using default parameters and comparing each sample against a reference set of 75 healthy individuals provided by Juha Kere.

Dataset Information

Genome-Wide Analysis of Somatic Copy Number Alterations (CGH, SNP) and Gene Expression (RNA-seq) in Metastatic Melanoma

Publications

Network-guided analysis of genes with altered somatic copy number and gene expression reveals pathways commonly perturbed in metastatic melanoma.

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