Genome-wide SNP Profiling of 27 Neuroblastoma Cell Lines
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE89968 | GEO | 2017/02/07
SECONDARY ACCESSION(S): PRJNA354536
REPOSITORIES: GEO
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Project description:Neuroblastoma cell lines are an important and cost-effective model used to study oncogenic drivers of the disease. While many of these cell lines have been previously characterized with SNP, methylation, and/or expression microarrays, there has not been an effort to comprehensively sequence these cell lines. Here, we present raw whole transcriptome data generated by RNA sequencing of 39 commonly-used neuroblastoma cell lines. This data can be used to perform differential expression analysis based on a genetic aberration or phenotype in neuroblastoma (eg: MYCN amplification status, ALK mutation status, 11q status, sensitivity to pharmacological pertubation). Additionally, we designed this experiment to enable structural variant and/or long-noncoding RNA analysis across these cell lines. Finally, as more DNase/ATAC and histone/transcription factor ChIP sequencing is performed in these cell lines, our RNA-Seq data will be an important complement to inform transcriptional targets as well as regulatory (enhancer or repressor) elements in neuroblastoma.
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Project description:TERT promoter muatations occur frequently in tumors of various origin. We here accessed the frequency of TERT pomoter mutations in 131 human primary neuroblastomas and 20 human neuroblastoma cell lines. No TERT promoter mutations were found in the 131 primary neuroblastomas. However, in the three cell lines SH-SY5Y, SH-EP and SK-N-SH a TERT promoter mutation was detected. Sanger sequencing indicated a homozygous mutation. To confirm loss of heterozygosity (LOH) we performed Affymetrix CytoScanHD SNP arrays. Indeed LOH could be confirmed. SNP arry experiments were performed according to the standard protocol for Affymetrix CytoScanHD arrays (Affymetrix, Santa Clara). Briefly, a 250 ng sample of genomic DNA was digested with NspI, ligated to adaptors, amplified by PCR, fragmented and biotin-labeled. The labeled samples were hybridized to Affymetrix CytoscanHD arrays, followed by washing, staining and scanning in the Affymetrix GeneChip Scanner 3000. Analysis was performed using the Affymetrix Chromosome Analysis Suite v2.1.
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Project description:Stromal contamination is one of the major confounding factors in the analysis of primary solid tumor samples by single nucleotide polymorphism (SNP) arrays. As we propose to employ genome-wide SNP microarray analysis as a diagnostic platform for neuroblastoma, the sensitivity, specificity, and accuracy of these studies must be optimized. In order to investigate the effects of stroma, we derived early passage cell lines from nine primary tumors and compared their genomic signature with that of the primary tumors by 100K SNP array analysis. The average concordance between tumor and cell line for raw LOH (loss of heterozygosity) calls was 96% (range 91%-99%) and for raw copy number alterations (CNA), 71% (range 43%-87%). In general, there were a larger number of LOH events identified in the cell lines compared to the matched tumor samples (mean increase 3.2% ± 1.9%). We have developed an algorithm that shows that the presence of stroma contributes to under-reporting of LOH and copy number loss (CNL). Notable findings in this sample set were uniparental disomy (UPD) of chromosome arms 11p, 1q, 14q, and 15q and a novel area of amplification on chromosome band 11p15. Our analysis demonstrates that LOH was identified significantly more often in derived cell lines compared to the original tumor samples. While these may in part be due to clonal selection during adaptation to tissue culture, our study indicates contamination by normal stromal elements may be a major contributing factor in underestimation of LOH and CNL events. Keywords: genome wide SNP analysis
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Project description:Stromal contamination is one of the major confounding factors in the analysis of primary solid tumor samples by single nucleotide polymorphism (SNP) arrays. As we propose to employ genome-wide SNP microarray analysis as a diagnostic platform for neuroblastoma, the sensitivity, specificity, and accuracy of these studies must be optimized. In order to investigate the effects of stroma, we derived early passage cell lines from nine primary tumors and compared their genomic signature with that of the primary tumors by 100K SNP array analysis. The average concordance between tumor and cell line for raw LOH (loss of heterozygosity) calls was 96% (range 91%-99%) and for raw copy number alterations (CNA), 71% (range 43%-87%). In general, there were a larger number of LOH events identified in the cell lines compared to the matched tumor samples (mean increase 3.2% ± 1.9%). We have developed an algorithm that shows that the presence of stroma contributes to under-reporting of LOH and copy number loss (CNL). Notable findings in this sample set were uniparental disomy (UPD) of chromosome arms 11p, 1q, 14q, and 15q and a novel area of amplification on chromosome band 11p15. Our analysis demonstrates that LOH was identified significantly more often in derived cell lines compared to the original tumor samples. While these may in part be due to clonal selection during adaptation to tissue culture, our study indicates contamination by normal stromal elements may be a major contributing factor in underestimation of LOH and CNL events. Keywords: genome wide SNP analysis Nine human neuroblastoma tumor samples with paired blood samples and derivative cell lines
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