Dataset Information

Genome wide analysis of DNA copy number neutral loss of heterozygosity (CNNLOH) and its relation to gene expression in esophageal squamous cell carcinoma.

ABSTRACT:

Background

Genomic instability plays an important role in human cancers. We previously characterized genomic instability in esophageal squamous cell carcinomas (ESCC) in terms of loss of heterozygosity (LOH) and copy number (CN) changes in tumors using the Affymetrix GeneChip Human Mapping 500K array in 30 cases from a high-risk region of China. In the current study we focused on copy number neutral (CN = 2) LOH (CNNLOH) and its relation to gene expression in ESCC.

Results

Overall we found that 70% of all LOH observed was CNNLOH. Ninety percent of ESCCs showed CNNLOH (median frequency in cases = 60%) and this was the most common type of LOH in two-thirds of cases. CNNLOH occurred on all 39 autosomal chromosome arms, with highest frequencies on 19p (100%), 5p (96%), 2p (95%), and 20q (95%). In contrast, LOH with CN loss represented 19% of all LOH, occurred in just half of ESCCs (median frequency in cases = 0%), and was most frequent on 3p (56%), 5q (47%), and 21q (41%). LOH with CN gain was 11% of all LOH, occurred in 93% of ESCCs (median frequency in cases = 13%), and was most common on 20p (82%), 8q (74%), and 3q (42%). To examine the effect of genomic instability on gene expression, we evaluated RNA profiles from 17 pairs of matched normal and tumor samples (a subset of the 30 ESCCs) using Affymetrix U133A 2.0 arrays. In CN neutral regions, expression of 168 genes (containing 1976 SNPs) differed significantly in tumors with LOH versus tumors without LOH, including 101 genes that were up-regulated and 67 that were down-regulated.

Conclusion

Our results indicate that CNNLOH has a profound impact on gene expression in ESCC, which in turn may affect tumor development.

SUBMITTER: Hu N

PROVIDER: S-EPMC3091724 | biostudies-literature | 2010 Oct

REPOSITORIES: biostudies-literature

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Publications

Genome wide analysis of DNA copy number neutral loss of heterozygosity (CNNLOH) and its relation to gene expression in esophageal squamous cell carcinoma.

Hu Nan N Clifford Robert J RJ Yang Howard H HH Wang Chaoyu C Goldstein Alisa M AM Ding Ti T Taylor Philip R PR Lee Maxwell P MP

BMC genomics 20101018

<h4>Background</h4>Genomic instability plays an important role in human cancers. We previously characterized genomic instability in esophageal squamous cell carcinomas (ESCC) in terms of loss of heterozygosity (LOH) and copy number (CN) changes in tumors using the Affymetrix GeneChip Human Mapping 500K array in 30 cases from a high-risk region of China. In the current study we focused on copy number neutral (CN = 2) LOH (CNNLOH) and its relation to gene expression in ESCC.<h4>Results</h4>Overall ...[more]

PMID: 20955586

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Project description:Somatic mosaicism denotes the presence of genetically distinct populations of somatic cells in one individual who has developed from a single fertilised oocyte. Mosaicism may result from a mutation that occurs during postzygotic development and is propagated to only a subset of the adult cells. Our aim was to investigate both somatic mosaicism for copy-neutral loss of heterozygosity (cn-LOH) events and DNA copy number variations (CNVs) in fully differentiated tissues.We studied panels of tissue samples (11-12 tissues per individual) from four autopsy subjects using high-resolution Illumina HumanOmniExpress-12 BeadChips to reveal the presence of possible intra-individual tissue-specific cn-LOH and CNV patterns. We detected five mosaic cn-LOH regions >5 Mb in some tissue samples in three out of four individuals. We also detected three CNVs that affected only a portion of the tissues studied in one out of four individuals. These three somatic CNVs range from 123 to 796 kb and are also found in the general population. An attempt was made to explain the succession of genomic events that led to the observed somatic genetic mosaicism under the assumption that the specific mosaic patterns of CNV and cn-LOH changes reflect their formation during the postzygotic embryonic development of germinal layers and organ systems.Our results give further support to the idea that somatic mosaicism for CNVs, and also cn-LOHs, is a common phenomenon in phenotypically normal humans. Thus, the examination of only a single tissue might not provide enough information to diagnose potentially deleterious CNVs within an individual. During routine CNV and cn-LOH analysis, DNA derived from a buccal swab can be used in addition to blood DNA to get information about the CNV/cn-LOH content in tissues of both mesodermal and ectodermal origin. Currently, the real frequency and possible phenotypic consequences of both CNVs and cn-LOHs that display somatic mosaicism remain largely unknown. To answer these questions, future studies should involve larger cohorts of individuals and a range of tissues.

Project description:Background: Insufficient quantities of human genomic DNA are a limiting factor for many clinical applications. Whole genome amplification (WGA) is an approach designed to overcome small amount of DNA for genome-wide genetic tests as it allows amplification of the entire genome from picogram or nanogram quantities of DNA. Various strategies of WGA have been developed; however, none of them can guarantee the absence of amplification bias. High-quality genome-representative amplified DNA is crucial for WGA use in basic research and clinical genetics. Thus, systematic evaluation of WGA effect on downstream methods is necessary. Results: In this paper, 4 multiple displacement amplification (MDA) -based and 2 PCR-based WGA kits were compared in their effect on segmental copy-number changes as well as copy-number neutral loss of heterozygosity detection by high-density oligonucleotide DNA arrays. We described outcomes and limits for each individual WGA; however, the main goal of this study was chiefly to show a general compatibility and features specific for particular WGA strategy. The main outcomes are as follows: 1) MDA-based WGAs showed higher tendency to generate false positive imbalances in contrast to PCR-based WGAs with higher risk of false negativity; 2) the specific risk of false positivity and/or negativity increased with decreasing copy-number segments size; 3) single-cell WGAs showed significantly worse effect on results in comparison to WGAs with nanogram level of DNA as input; 4) PCR-based WGAs were not compatible with copy-number neutral loss of heterozygosity analysis based on single nucleotide polymorphisms in restriction digestion sites and also showed higher risk of copy-number neutral loss of heterozygosity false negativity if combined with analysis based on simple hybridization. Conclusions: This study gives a comprehensive insight into the WGA effect on DNA array analysis. The results of this study help to choose WGA according to individual user requirements and options. Moreover, we show a strategy to verify and validate segmental copy-number changes detection by DNA array protocol including any WGA for any purpose to attain the highest efficiency without an unnecessary WGA bias.

Project description:Esophageal squamous cell carcinoma (ESCC) is one of the most common cancers worldwide with poor prognosis. Thus, identification of predictive biomarkers for early diagnosis and intervention is needed to improve patients' survival. Research shows that heritable mutations, such as single nucleotide polymorphisms (SNPs), contribute to human cancer susceptibility significantly. However, the association of copy number variations (CNVs), another major source of genetic variation, with ESCC risk remains poorly clarified. In this study, we aimed to identify ESCC risk-related CNVs based on candidate-gene strategy in a case-control study. A meta-analysis was first performed to identify the most variable chromosome regions of ESCC tissues. Bioinformatic analysis and dual-luciferase reporter assays were carried out to evaluate the properties of all recorded CNVs located on these regions. Six candidate CNVs located within well-known oncogenes and detoxification-associated enzymes were enrolled in the final analysis. A newly developed multiplex gene copy number quantitation method AccuCopy(TM) was employed to simultaneously genotype all six candidate sites in 404 ESCC patients and 402 cancer-free controls from Southwest China, and in 42 ESCC tissues. qRT-PCR was performed to measure UGT2B28 mRNA in cancerous and corresponding normal tissues. Unconditional logistic regression was applied to test association between germline CNV genotypes and ESCC risk. Relationship between germline copy number variation and somatic copy number alterations was further analyzed. Finally we found that copy number loss of UDP-glucuronosyltransferase family 2, polypeptide B28 (UGT2B28) and gain of plectin (PLEC) conferred increased ESCC risk (Adjusted OR = 2.085, 95% CI = 1.493-2.912, P < 0.001 for UGT2B28. Adjusted OR = 3.725, 95% CI = 1.026-13.533, P = 0.046 for PLEC). mRNA level was lower in UGT2B28 loss genotyped esophageal tissues than in two-copy tissues, indicating that UGT2B28 loss genotypes modify ESCC susceptibility perhaps by decreasing UGT2B28 expression level and enzyme activity. In addition, an association was drawn between germline copy number variations and somatic alterations for PLEC, UGT2B17 and UGT2B28, but not for other candidate loci.

Dataset Information

Genome wide analysis of DNA copy number neutral loss of heterozygosity (CNNLOH) and its relation to gene expression in esophageal squamous cell carcinoma.

Background

Results

Conclusion

Publications

Genome wide analysis of DNA copy number neutral loss of heterozygosity (CNNLOH) and its relation to gene expression in esophageal squamous cell carcinoma.

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