DETECTING AND ANALYZING COPY NUMBER ALTERNATIONS IN ARRAY-BASED CGH DATA

2016 ◽  
Vol 28 (06) ◽  
pp. 1650044 ◽  
Author(s):  
Mariam A. Sheha ◽  
Mai S. Mabrouk ◽  
Mahmoud Elhefnawi
Keyword(s):  
Copy Number ◽  
Genomic Dna ◽  
Array Cgh ◽  
Artificial Chromosome ◽  
Gaussian Mixture ◽  
Comparative Genomic ◽  
Intensity Ratios ◽  
Dna Copy Number Alterations ◽  
Copy Number Changes ◽  
Tissue Specimens

Copy number changes or alterations are a form of genetic variation in the human genome. Genomic DNA copy number alterations (CNAs) are associated with the development and progression of cancers. Array-based comparative genomic hybridization (a-CGH) is a technique used to identify copy number changes in genomic DNA. It yields data consisting of fluorescence intensity ratios of test and reference DNA samples. The intensity ratios provide information about the number of copies in DNA. Practical issues such as the contamination of tumor cells in tissue specimens and normalization errors necessitate the use of automated statistics algorithms for learning about the genomic alterations from array CGH data. Specifically, there is a need for algorithms that can identify gains and losses in the number of copies based on statistical considerations, rather than merely detect trends in the data. For this purpose the proposed study introduces three different approaches; Circular binary segmentation, Bayesian approach, relying on the hidden Markov model and effective Gaussian mixture (GM) clustering for the analysis of array CGH profiles. Publicly available data on pancreatic adenocarcinoma and Coriell cell line bacterial artificial chromosome (BAC) array were used for the analysis to illustrate the reliability and success of the techniques.

Genomic DNA Copy Number Alterations Present in AML Bone Marrow Samples with Normal Cytogenetics.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 142-142 ◽  
Author(s):  
Matthew J. Walter ◽  
Rhonda E. Ries ◽  
Jon Armstrong ◽  
Brian O’Gara ◽  
James W. Vardiman ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Copy Number ◽  
Genomic Dna ◽  
Array Cgh ◽  
Comparative Genomic ◽  
Copy Number Alterations ◽  
Dna Copy Number ◽  
Prognostic Information ◽  
Normal Cytogenetics ◽  
Copy Number Changes

Abstract Cytogenetics and comparative genomic hybridization (CGH) have been used to identify large genomic amplifications and deletions in all subtypes of acute myeloid leukemia (AML). Up to 15–20% of AML patients have a normal karyotype at diagnosis. While cytogenetic abnormalities confer important prognostic information for patients with AML, there remain differences in the therapeutic response and outcome among patients with the same cytogenetic profile, implying that other, more subtle, genetic abnormalities may exist. We hypothesized that a subset of AML patients with normal cytogenetics may contain genomic DNA copy number changes that are too small to be detected using standard cytogenetic techniques. To address this possibility, we used high-resolution bacterial artificial chromosome (BAC) array CGH technology to examine 31 AML patients with normal cytogenetics. The BAC arrays contain 2,464 BAC clones spotted in triplicate on glass slides, and provide a 1 Mb resolution of the entire human genome. Technical generation of the arrays, hybridization parameters, and analysis were similar to that reported for murine BAC array CGH (Nat Genet. 2001 Dec;29(4):459–64). The 31 AML samples included 4 M0, 8 M1, 10 M2, and 9 M4 patients. Array CGH experiments were performed using 500 nanograms of Cyanine 5 labeled genomic DNA from unmanipulated AML bone marrow, mixed with an equal amount of control DNA (a pool of DNA from 4 cancer-free individuals) labeled with Cyanine 3. Using the human 1 Mb BAC arrays, we identified amplifications and deletions from multiple samples that were confirmed with G-banding cytogenetics [del(7)(q31), del(7)(p11.2), +8, del(11)(q13q23), +21, add(21)(q22), −X, −Y, +Y]. In addition, BAC arrays robustly detected copy number alterations that were identified in as few as 4/21 metaphases. We identified 5/31 (16%) patients with normal cytogenetics that contained altered genomic DNA copy numbers using BAC array CGH. Copy number changes were confirmed for several of these genomic loci using a dye-swap experiment, where the AML DNA was labeled with Cyanine 3, and the control DNA with Cyanine 5. Two of the 5 patients with abnormalities detected using array CGH would be reclassified from “intermediate” to “unfavorable” cytogenetics [del(7)(q31.31q34), add(11)(q23.3qter), and 17(p12pter)]. These results suggest that a subset of AML patients with normal cytogenetics contain genomic copy number alterations that may effect treatment and outcome. Patient # FAB subtype Genomic location Gain or loss Size (Megabase) Dye-Swap confirmed 1 M0 7(q31.31q34) loss 2.0 Not done 1 11(q23.3qter) gain 16.5 Not done 2 M1 11(p14) loss 7.4 Yes 3 M1 11(q13.2q14.1) gain 15.8 Yes 3 19(p) gain 64.0 Yes 4 M2 17(p12pter) gain 8.6 Not Done 5 M2 19(p13.1pter) loss 14.8 Yes 5 12(q13) loss 5.0 Yes

Novel DNA Copy Number Changes in Hematological Malignancies: A cDNA-Based CGH Microarray Screening of CML, AML and CLL Cases without Chromosomal Imbalances in G-Banding.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4418-4418
Author(s):  
Tuija Lundan ◽  
Anne Oikarainen ◽  
Lena Hafren ◽  
Maija Wolf ◽  
Erkki Elonen ◽  
...  
Keyword(s):  
Copy Number ◽  
Myeloid Leukemia ◽  
Array Cgh ◽  
Reference Sample ◽  
Copy Number Alterations ◽  
Dna Copy Number ◽  
Chromosomal Imbalances ◽  
Single Allele ◽  
Dna Copy Number Alterations ◽  
Copy Number Changes

Abstract Copy number changes, such as small single allele losses and gains, have important roles in the mechanisms of cancer development. These alterations also have often prognostic significance. Genome wide screening of DNA copy number losses previously conducted by extensive LOH analyses can now be performed with array-based comparative genomic hybridization (array CGH). We assessed the utility of array CGH in the detection of single allele deletions and gains in a cohort of seven patients with chronic myeloid leukemia (CML), seven patients with chronic lymphatic leukemia (CLL) and three patients with acute myeloid leukemia (AML). All the CLL and AML patients had a normal karyotype as assessed by standard G-banding. In CML patients the only clonal abnormality detected by cytogenetics was the reciprocal Philadelphia translocation, t(9;22)(q34;q11). The derivative chromosome 9 [der(9)] deletion status of the CML patients was determined using fluorescence in situ hybridization (FISH) analysis. Four patients did not have the deletion, two had a der(9) deletion spanning both 5′ABL and 3′BCR regions and one patient had a deletion of the 5′ABL region alone. The array CGH experiments were performed using Agilent Technologies Human 1 cDNA microarray slides consisting of 13,000 clones. A total of 6 ug of fluorescently labeled DNA extracted from bone marrow samples was hybridized on cDNA array. Normal male or female DNA was used as the reference sample in the hybridization. The slides were scanned with the Agilent fluorescent scanner and intensity ratio data between the tumor and reference sample was processed using Feature Extraction software. The data was filtered and analyzed using SPSS (version 11) and Origin 7.0 softwares. The processed, untransformed red-to-green fluorescence signal ratio was used for evaluating gene dosage. Ratios greater than 1.1 were considered to indicate DNA copy number gains and ratios below 0.9 DNA copy number losses in tumor samples. In two CML patients who had deletions covering both the 5′ABL and 3′BCR regions in the translocation breakpoint of der(9), the deletion was detectable with the array CGH. In four patients with no deletion the red-to-green ratio profile for der(9) was 1. However, in one patient with an isolated 5′ABL deletion, the deletion was not visible in array CGH. No other obvious DNA copy number alterations were seen in CML patients. Array CGH detected deletions in three of the seven CLL patients. Deletions were found in 13q14, 2q32-33 and 14q24. One of the three AML patients studied showed an amplification in chromosome 9p. No alterations were seen in the other two AML patients. The FISH and array studies are being done on larger set of patient samples to confirm the results. We conclude that array CGH provides new information in patients without chromosomal imbalances in standard cytogenetics and enables the detection of novel small submicroscopic copy number alterations. Furthermore, a cDNA-based array platform can be used both for studies of DNA copy number alterations and gene expression analyses.

scholarly journals Characteristics of Highly Polymorphic Segmental Copy-Number Variations Observed in Japanese by BAC-Array-CGH

2011 ◽  
Vol 2011 ◽  
pp. 1-11 ◽  
Author(s):  
Norio Takahashi ◽  
Yasunari Satoh ◽  
Keiko Sasaki ◽  
Yuko Shimoichi ◽  
Keiko Sugita ◽  
...  
Keyword(s):  
Genetic Variation ◽  
Copy Number ◽  
Array Cgh ◽  
Artificial Chromosome ◽  
Copy Number Variations ◽  
Comparative Genomic ◽  
Bac Clones ◽  
Japanese Cohort ◽  
The Individual ◽  
Chromosome Microarray

Segmental copy-number variations (CNVs) may contribute to genetic variation in humans. Reports of the existence and characteristics of CNVs in a large Japanese cohort are quite limited. We report the data from a large Japanese population. We conducted population screening for 213 unrelated Japanese individuals using comparative genomic hybridization based on a bacterial artificial chromosome microarray (BAC-aCGH). We summarize the data by focusing on highly polymorphic CNVs in ≥5.0% of the individual, since they may be informative for demonstrating the relationships between genotypes and their phenotypes. We found a total of 680 CNVs at 16 different BAC-regions in the genome. The majority of the polymorphic CNVs presented on BAC-clones that overlapped with regions of segmental duplication, and the majority of the polymorphic CNVs observed in this population had been previously reported in other publications. Some of the CNVs contained genes which might be related to phenotypic heterogeneity among individuals.

High-Resolution Genomic Profiling of Myelodysplasia (MDS) by Microarray Comparative Genomic Hybridization (CGH).

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2645-2645
Author(s):  
Noor Esoof ◽  
Aristoteles Giagounidis ◽  
Mario Cazzola ◽  
Luca Malcovati ◽  
Carlo Aul ◽  
...  
Keyword(s):  
Copy Number ◽  
Genomic Dna ◽  
Normal Karyotype ◽  
Comparative Genomic Hybridisation ◽  
Comparative Genomic ◽  
Bac Clones ◽  
Conventional Cytogenetic Analysis ◽  
Copy Number Changes ◽  
Normal Controls ◽  
Microarray Cgh

Abstract Myelodysplasia (MDS) is a heterogeneous group of clonal disorders of hematopoietic stem cells characterised by ineffective hematopoiesis and a variable risk of transformation to acute myelogenous leukaemia. We have used Comparative Genomic Hybridisation (CGH) microarray analysis, a technology that represents a significant improvement in resolution over conventional cytogenetic analysis, to screen genomic DNA from MDS patients for the identification of genome-wide Copy-Number Changes (CNCs). We have studied genomic DNA obtained from the neutrophil population of 48 MDS patients and 40 normal controls. Of the 48 MDS patients 10 had the 5q- syndrome, 32 were assigned normal karyotype and 6 had complex karyotypes. Comparative Genomic Hybridisation (CGH) microarray analysis was performed using microarrays containing 3500 BAC clones at 1Mb intervals over the whole human genome. Furthermore we used a whole genome tiling-path (27 000 overlapping BAC clones) array to profile 9 5q-syndrome patients and for 3 of those patients the T-cell DNA were also profiled to act as constitutional control. The patient DNA and a pool of normal reference DNA was labelled with different fluorochromes and cohybridised to the microarray. The normalised ratio of signal intensities was calculated and log2 ratios between −0.4 and 0.4 were considered normal. Ratios below or above the normal range were interpreted as loss or gain of genetic material, respectively. The deletions on chromosome 5q were precisely mapped by array-CGH in the patients with the 5q- syndrome but no additional CNCs were detected. One of the 5q deletions, however, displayed a discontiguous pattern with the tiling resolution array. Copy-number changes (CNCs) that escaped conventional cytogenetic detection were identified in the MDS patients originally reported with normal bone marrow karyotypes. 8 out of those 32 patients displayed CNCs that were not detected in the 40 normal controls and as such were considered as disease-related changes (non-polymorphic). Many of those CNCs were single-clone abberrations that were validated by dye-swap experiments and some were confirmed by quantitative PCR. Microarray CGH data confirmed all abnormalities reported by conventional cytogenetic analysis in the MDS patients with complex karyotypes and previously undetected abnormalities were uncovered. Several genes involved in either the initiation or progression of hematological malignancies are known to map within the cryptic abnormalities identified in the patients studied. For example, one patient with an apparently normal karyotype showed a small deletion at 17q11 which encompasses the NF1 gene. Further work will determine whether particular abnormalities detected by microarray CGH are recurrent and the nature of the genes involved. However, the promise of microarray CGH in the diagnostic work up of MDS particularly in those patients with normal karyotypes is clear.

Quantitation of sequence copy-number changes in genomic DNA through GenoSensor-based comparative genomic hybridization

10.1038/14372 ◽  
1999 ◽  
Vol 23 (S3) ◽  
pp. 64-65 ◽  
Author(s):  
U.R. Müller ◽  
Y.P. Bao ◽  
D. Che ◽  
N. Lermer ◽  
W.R. Li ◽  
...  
Keyword(s):  
Comparative Genomic Hybridization ◽  
Copy Number ◽  
Genomic Dna ◽  
Comparative Genomic ◽  
Genomic Hybridization ◽  
Copy Number Changes

scholarly journals Array comparative genomic hybridization reveals genomic copy number changes associated with outcome in diffuse large B-cell lymphomas

Blood ◽  
2006 ◽  
Vol 107 (6) ◽  
pp. 2477-2485 ◽  
Author(s):  
Weiyi Chen ◽  
Jane Houldsworth ◽  
Adam B. Olshen ◽  
Gouri Nanjangud ◽  
Seeta Chaganti ◽  
...  
Keyword(s):  
B Cell ◽  
Comparative Genomic Hybridization ◽  
Copy Number ◽  
Array Comparative Genomic Hybridization ◽  
Array Cgh ◽  
B Cell Lymphoma ◽  
Comparative Genomic ◽  
Genomic Hybridization ◽  
Copy Number Changes ◽  
Large B Cell

Abstract To identify, in high-resolution regions of DNA, the copy number changes associated with outcome in patients with diffuse large B-cell lymphoma (DLBCL), a disease with an approximately 50% mortality rate, we performed array comparative genomic hybridization (array-CGH) on specimens from 64 patients with newly diagnosed DLBCL treated with anthracycline-based chemotherapy. For the entire cohort, 55 commonly gained/lost regions, ranging in size from less than 1 Mbp to entire chromosomes, were identified using 1- to 2-Mbp and 2- to 4-Mbp resolution BAC arrays. Copy number changes of 9 minimal regions significantly correlated with overall survival, of which 6 were 10 Mbp or smaller. On multivariate analysis, loss of chromosomes 2 (2.4-4.1 Mbp) and 16 (33.8-35.6 Mbp) were found to be prognostic indicators of poor survival, independent of clinical features routinely used to predict outcome. Loss of chromosome 1 (78.2-79.1 Mbp) was predictive of good outcome. For a subset of 55 specimens classified according to cell-of-origin expression signature subtype, gain of chromosome 12 (45.4-53.8 Mbp) was found to be significantly associated with the germinal center B-cell-like DLBCL subtype. Overall, array-CGH identified relatively small genomic regions associated with outcome, which, along with follow-up expression studies, may reveal target genes important in DLBCL clinical behavior. (Blood. 2006;107:2477-2485)

scholarly journals Mixed Gastric Carcinomas Show Similar Chromosomal Aberrations in Both their Diffuse and Glandular Components

2006 ◽  
Vol 28 (5-6) ◽  
pp. 283-294
Author(s):  
Beatriz Carvalho ◽  
Tineke E. Buffart ◽  
Rui M. Reis ◽  
Thomas Mons ◽  
Cátia Moutinho ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Chromosomal Aberrations ◽  
Copy Number ◽  
Array Cgh ◽  
Comparative Genomic Hybridisation ◽  
Comparative Genomic ◽  
Dna Copy Number ◽  
Gastric Carcinomas ◽  
Phenotypic Divergence ◽  
Copy Number Changes

Gastric cancer is one of the most frequent malignancies in the world. Nonetheless, the knowledge of the molecular events involved in the development of gastric carcinoma is far from complete. One of the hallmarks of gastric cancer is chromosomal instability resulting in abnormal DNA copy number changes throughout the genome. Mixed gastric carcinomas constitute a rare histological entity, containing the two main histological phenotypes (diffuse and intestinal). Very little is known about the underlying mechanisms of phenotypic divergence in these mixed tumours. To the best of our knowledge only E-Cadherin mutations were implicated so far in the divergence of these tumours and nothing is known about the involvement of chromosome copy number changes in the two divergent histological components. In this study, we compared the DNA copy number changes, in the two different components (diffuse and intestinal) of mixed gastric carcinomas by microarray – comparative genomic hybridisation (array CGH). The analysis of 12 mixed gastric carcinomas showed no significant differences in array CGH profiles between the diffuse and intestinal components of mixed carcinomas. This supports the idea that the phenotypic divergence within mixed gastric carcinomas is not caused by DNA chromosomal aberrations.

scholarly journals Amplification at 9p in Cervical Carcinoma by Comparative Genomic Hybridization

2001 ◽  
Vol 22 (3) ◽  
pp. 159-163 ◽  
Author(s):  
Kowan J. Jee ◽  
Young Tak Kim ◽  
Kyu Rae Kim ◽  
Yan Aalto ◽  
Sakari Knuutila
Keyword(s):  
Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Comparative Genomic Hybridization ◽  
Copy Number ◽  
Comparative Genomic ◽  
Malignant Cells ◽  
Genomic Hybridization ◽  
Carcinoma Of Cervix ◽  
Chromosome Arm ◽  
Copy Number Changes

DNA copy number changes were studied by comparative genomic hybridization on 10 tumor specimens of squamous cell carcinoma of cervix obtained from Korean patients. DNA was extracted from paraffin‐embedded sections after removal of non‐malignant cells by microdissection technique. Copy number changes were found in 8/10 tumors. The most frequent changes were chromosome 19 gains (n=6) and losses on chromosomes 4 (n=4), 5 (n=3), and 3p (n=3). A novel finding was amplification in chromosome arm 9p21‐pter in 2 cases. Gains in 1, 3q, 5p, 6p, 8q, 16p, 17, and 20q and losses at 2q, 6q, 8p, 9q, 10p, 11, 13, 16q, and 18q were observed in at least one of the cases.

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