Array CGH As a Complementary Tool in the Diagnosis of Myelodysplastic Syndromes

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3827-3827
Author(s):  
María Abáigar ◽  
Eva Lumbreras ◽  
Irene Rodríguez ◽  
Javier Sánchez-del-Real ◽  
María Díez-Campelo ◽  
...  
Keyword(s):  
Risk Stratification ◽  
Myelodysplastic Syndromes ◽  
Copy Number ◽  
Normal Karyotype ◽  
Chromosome 8 ◽  
Comparative Genomic ◽  
Whole Genome ◽  
Acgh Analysis ◽  
Conventional Cytogenetics ◽  
Copy Number Changes

Abstract Abstract 3827 Background: Myelodysplastic syndromes (MDS) are a heterogeneous group of hematological disorders in which diagnosis, risk stratification, and treatment selection are based on morphological and cytogenetic studies in bone marrow (BM) samples. MDS are characterized by several recurrent chromosomal abnormalities, most of them unbalanced, with a widely variable prognosis. The assessment of these genomic defects is essential for a correct risk stratification of these patients. However, conventional cytogenetic (CC) techniques are not sufficient for the study of all MDS patients, because of the high proportion of normal karyotypes (40–50%) and unsuccessful cytogenetics (10%) (defined as the absence of mitosis). Array-based comparative genomic hybridization (aCGH) technology allows the screening of copy number changes among the whole genome in one single experiment and offers a higher resolution than conventional cytogenetics. Aims: To assess the potential application of aCGH in the clinical diagnosis of MDS as complementary tool to conventional cytogenetics. Patients and Methods: The study cohort comprises a total of 263 patients: MDS (203) and MDS/MPN (60) patients that have been previously studied by CC and FISH. Among the whole series, 33 (12.5%) patients had no successful cytogenetic results due to the absence of mitosis. In the remaining 230 patients with evaluable metaphases, 42 (16%) had an aberrant, while 188 (71.5%) presented a normal karyotype. Within this last group, 141 had ≥20 good-quality metaphases evaluated, 37 had 10–20 metaphases studied, and 10 patients had ≤10 successful metaphases. Copy number changes were analysed in all patients included in the study using NimbleGen Human CGH 12×135K Whole-Genome Tiling Array (Roche NimbleGen). Sex-matched human commercial DNA samples were used as reference. Data were analysed using the segMNT algorithm in NimbleScanv2.6 Software. Subsequently all genomic abnormalities found by aCGH analysis were confirmed by FISH. Results: Using aCGH methodology, copy number changes (greater than 600 bp) were detected in 54 patients of the global series: 4.3% of the normal karyotype patients, 88.1% of cases with abnormal cytogenetics, and 27.3% of patients with unsuccessful cytogenetics. Overall a high correlation (94.3%) between the cytogenetic changes observed by CC and CGH arrays was observed. Thus aCGH analysis revealed the same genomic abnormalities showed by CC in 88.1% of patients. In the remaining 11.9% genomic results were discordant between aCGH and CC, because of the presence of balanced translocations, not assessable by aCGH, and clonal cell populations below 30%. Furthermore, additional genomic abnormalities (n=36) not detected by CC were found by aCGH. The most frequent aberrations were losses affecting chromosomes 5 (33%), 7/7q (17%), 20q (14%), and Y (14%), as well as gains involving chromosome 8 (14%). Interestingly, other abnormalities, mainly losses, were found in chromosomes 4, 12, and 17. Focusing on the 188 patients with normal karyotype by CC, the aCGH profiling results were concordant with cytogenetics in 98% of those patients with ≥20 metaphases studied and in 92% of those with 10–20 metaphases. However, only 80% of those patients with ≤10 successful metaphases and no changes by CC displayed no copy number changes by aCGH. The most frequent abnormality found by aCGH among these normal karyotype cases was the presence of 5q deletion (2%), while other chromosomes affected were 7, 8, 11, 12 and 20. All these abnormalities were confirmed by FISH. Regarding the patients with unsuccessful cytogenetics, 72.7% of cases displayed a normal aCGH profile, while 27.3% showed at least one genomic imbalance The most frequent genomic aberrations were losses in 4q (6%), 5q (12%) and 7q (9%), and gain of chromosome 8 (6%). In addition, three of these cases showed a complex karyotype, showing more than 5 abnormalities. Conclusion: The use of aCGH karyotyping in the diagnosis of MDS could be used as a complementary technique to conventional karyotyping in the evaluation of MDS patients. Mainly in patients with unsuccessful cytogenetics and those with normal karyotype and <20 good-quality metaphases evaluated. Disclosures: Hernández: Celgene: Research Funding.

High-resolution whole genome tiling path array CGH analysis of CD34+ cells from patients with low-risk myelodysplastic syndromes reveals cryptic copy number alterations and predicts overall and leukemia-free survival

Blood ◽  
2008 ◽  
Vol 112 (8) ◽  
pp. 3412-3424 ◽  
Author(s):  
Daniel T. Starczynowski ◽  
Suzanne Vercauteren ◽  
Adele Telenius ◽  
Sandy Sung ◽  
Kaoru Tohyama ◽  
...  
Keyword(s):  
Myelodysplastic Syndromes ◽  
Copy Number ◽  
Prognostic Significance ◽  
Normal Karyotype ◽  
Low Risk ◽  
International Prognostic Scoring System ◽  
Comparative Genomic ◽  
Whole Genome ◽  
Copy Number Alterations ◽  
Cd34 Cells

Abstract Myelodysplastic syndromes (MDSs) pose an important diagnostic and treatment challenge because of the genetic heterogeneity and poorly understood biology of the disease. To investigate initiating genomic alterations and the potential prognostic significance of cryptic genomic changes in low-risk MDS, we performed whole genome tiling path array comparative genomic hybridization (aCGH) on CD34+ cells from 44 patients with an International Prognostic Scoring System score less than or equal to 1.0. Clonal copy number differences were detected in cells from 36 of 44 patients. In contrast, cells from only 16 of the 44 patients displayed karyotypic abnormalities. Although most patients had normal karyotype, aCGH identified 21 recurring copy number alterations. Examples of frequent cryptic alterations included gains at 11q24.2-qter, 17q11.2, and 17q12 and losses at 2q33.1-q33.2, 5q13.1-q13.2, and 10q21.3. Maintenance of genomic integrity defined as less than 3 Mb total disruption of the genome correlated with better overall survival (P = .002) and was less frequently associated with transformation to acute myeloid leukemia (P = .033). This study suggests a potential role for the use of aCGH in the clinical workup of MDS patients.

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.

Whole Genome Association Study in Leukemia: Frequent Loss of Heterozygosity (LOH) and Single Nucleotide Polymorphism (SNP) in Acute Myeloid Leukemia with a Normal Karyotype.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2838-2838
Author(s):  
Young Y. Lee ◽  
Joowon Park ◽  
Sung-Soo Yoon ◽  
Kwang-Sung Ahn ◽  
Jung H. Choi ◽  
...  
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Copy Number ◽  
Myeloid Leukemia ◽  
Normal Karyotype ◽  
Whole Genome ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Genome Wide ◽  
Copy Number Changes ◽  
Acute Myeloid

Abstract Acute myeloid leukemia (AML) is a heterogeneous disease with various chromosomal aberrations. The karyotype at diagnosis is generally recognized as the single most valuable prognostic factor. However, using conventional cytogenetic techniques, karyotype abnormalities are detected in only half of all AML cases and the other half are commonly described as normal-karyotype AML. Patients with normal-karyotype AML are classified as an intermediate risk group with a 5-year overall survival of between 35% and 45%, but clinical outcome may vary greatly. Also, classical cytogenetic methods are limited in resolution and dependent on highly skilled labor. Therefore, the appropriate choice of additional treatment in patients who attained first remission (chemotherapy versus autologous transplantation versus allogeneic transplantation) is unclear for these patients. Thus, additional markers with prognostic significance are needed to identify clinically relevant subgroups among AML patients with a normal karyotype. Recently, human genome-wide studies gain popularity to identify the genetic basis of complex disorders such as AML. Altered transcript levels in AML genomes are often related to copy number changes, and genome-wide detection of allelic imbalance in AML cells by polymorphic genetic markers has become an important technique to identify genetic events involved in the progression of AML. By using high density single-nucleotide polymorphism (SNP) microarrays designed to genotype more than 300K SNPs in the human genome DNA, the resolution of the whole genome scanning technique has increased considerably and allowed accurate and reproducible determination of copy number changes in whole genome of AML. It became possible to distinguish between LOH regions with underlying homozygous deletions and those with copy-neutral events. In the present study, we performed genetic changes in untreated AML with normal cytogenetics with infinium 300K SNP chip. SNP-based mapping array data and fluorescence in situ hybridization (FISH) copy number data correlated well. The most frequently identified alterations are located at 3p, 6q, 8p, 13q, 21q and 22p. LOH is found in these large regions and also in smaller regions throughout the genome with a median size of 1 Mb. Alterations was correlated with response to chemotherapy. Twenty six candidate genes showed significant evidence of linkage in the presence of disequilibrium, and ten of these were expressed in AML patients who failed to attain remission. Three other genes showing statistical evidence were not expressed. Many of the genes reported here have not been previously reported in relation to progression of AML. We show that this panel of markers adds important prognostic information for this largest subgroup in AML.

High-Resolution CGH Analysis of CD34+ Cells from Lower-Risk Myelodysplastic Patients Reveals Cryptic Copy Number Alterations and Predicts Overall and Leukemia-Free Survival.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2431-2431
Author(s):  
Daniel T. Starczynowski ◽  
Suzanne Vercauteren ◽  
Adele Telenius ◽  
Sandy Sung ◽  
Kaoru Tohyama ◽  
...  
Keyword(s):  
Risk Stratification ◽  
Copy Number ◽  
Lower Risk ◽  
Array Cgh ◽  
Comparative Genomic ◽  
Whole Genome ◽  
Copy Number Alterations ◽  
Ancillary Test ◽  
Cd34 Cells ◽  
Marrow Cells

Abstract The myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematologic malignancies characterized by peripheral cytopenias, a hypercellular marrow with ineffective hematopoiesis and a propensity to progress to acute myeloid leukemia (AML). MDS is thought to arise from a primitive (CD34+) hematopoietic cell that has acquired genetic and/or epigenetic abnormalities. Risk stratification according to the International Prognostic Scoring System (IPSS) defines patient categories that correlate with survival and the likelihood of transformation to AML. Greater than 50% of individuals diagnosed with MDS are in the lower-risk groups. The importance of cytogenetics in risk stratification has been verified in several studies, but is of limited value in patients with lower-risk subtypes because approximately 50% of these patients do not have karyotypic abnormalities detectable using standard techniques. To further our biological understanding of lower-risk subtypes of MDS, and to identify potential MDS-initiating alterations in the genome, we looked for alterations in DNA extracts from purified CD34+ marrow cells from 44 MDS lower-risk patients using a submegabase bacterial artificial chromosome (BAC) array to perform whole genome comparative genomic hybridization (CGH) analyses. These studies identified numerous cryptic structural DNA alterations that were not detectable by standard cytogenetic analysis and were also not found in 15 age-matched normal controls. Although most patients tested had a normal karyotype, 23 recurring, novel copy number alterations of a median size of 0.6 megabases were identified. These included gains at 11q24.2-qter, 17q11.2 and 17q12, and losses at 2q33.1-q33.2 and 14q12. Comparison of changes in CD34+ marrow cells with DNA from CD3+ cells isolated from the same patients showed that a recurring duplication at band 17q12 was exclusive to the CD34+ cells in 2 of 3 patients. Validation of this copy number gain at chr 17q12 by FISH confirmed duplication of the locus. In addition, whole genome array CGH analysis of CD34+ marrow cells from karyotypically normal (n = 25) and abnormal (n = 15) lower risk MDS patients revealed extensive genome alterations (involving &gt;3 Mb) correlated with poorer overall survival, and was more frequently associated with transformation to AML as compared to IPSS stratification alone. Our studies suggest that array CGH may be useful as an ancillary test to better stratify lower-risk subtypes of MDS and, at the same time lead to the identification of early mutations that contribute to disease initiation.

Utility of Oligonucleotide Array Comparative Genomic Hybridization to Identify Cryptic Copy Number Alterations in Myelodysplastic Syndromes

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5076-5076 ◽  
Author(s):  
Wendy K Chung ◽  
James Weisberger ◽  
Yi Sun ◽  
Pauline Brenholz ◽  
Stephanie Warren ◽  
...  
Keyword(s):  
Comparative Genomic Hybridization ◽  
Myelodysplastic Syndromes ◽  
Copy Number ◽  
Array Comparative Genomic Hybridization ◽  
Chromosome Analysis ◽  
Comparative Genomic ◽  
Copy Number Alterations ◽  
Genomic Hybridization ◽  
Conventional Cytogenetics ◽  
Gains And Losses

Abstract Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematological neoplasms characterized by peripheral cytopenias due to ineffective hematopoiesis and significant cytologic atypia in one or more of the myeloid lineages. There is a variable risk of progression to acute myeloid leukemia, which is dependent on the blast count and certain recurrent cytogenetic abnormalities. Cytogenetic characterization is important in both diagnosis and prognosis, but can be of limited value in lower-risk MDS subtypes because of the low frequency of karyotypic abnormalities detected by chromosome analysis. Recently, array comparative genomic hybridization (aCGH) has been shown to be an important technique in detecting gross and cryptic copy number changes in leukemias. However, there is little data on aCGH application in MDS. We used a custom design 105,000 probe oligonucleotide aCGH platform to analyze 44 bone marrows, in which a diagnosis of MDS was made to determine the clinical utility of oligonucleotide aCGH. All bone marrows were analyzed by morphology, flow cytometry, cytogenetics, and fluorescent in situ hybridization (FISH). Gender matched normal controls were used to assess copy number alterations in MDS samples. We classified significant gains and losses as a minimum of 10% deviation from the baseline of at least 10 contiguous oligonucleotide probes spanning at least 1 Mb and as not within a previously defined region of copy number polymorphism in the Database of Genomic Variants. Array CGH was able to detect all cytogenetically identified imbalances present in more than 1 cell, but as expected did not detect any of the balanced translocations or inversions. Of the 24 cases that were cytogenetically normal, 9 (38%) had anomalies detected by aCGH. In 5 (25%) of the 20 cases with at least one cytogenetic abnormality detected previously by chromosome analysis, additional imbalances were identified by aCGH. Recurrent copy number losses detected by aCGH were observed in 13/44 (29.5%) of cases and included 5q21 in 5 cases, chromosome 7 in 2 cases, 12p13 in 3 cases, 13q21-31 in 3 cases, and 20q12-13 in 2 cases. Oligonucleotide aCGH was able to provide extremely high resolution of the boundaries of the gains and losses due to the high oligonucleotide probe density. In 14/44 cases (31.8%), oligonucleotide aCGH provided novel information not provided by conventional cytogenetics and FISH and can be used to precisely define recurrent gains and losses. Our data demonstrates the clinical applicability of oligonucleotide aCGH in the diagnosis of MDS in addition to conventional cytogenetics to detect new cryptic abnormalities which may provide novel targets for therapy.

scholarly journals Whole-genome sequencing from the New Zealand Saccharomyces cerevisiae population reveals the genomic impacts of novel microbial range expansion

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Peter Higgins ◽  
Cooper A Grace ◽  
Soon A Lee ◽  
Matthew R Goddard
Keyword(s):  
Saccharomyces Cerevisiae ◽  
New Zealand ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Range Expansion ◽  
Copy Number ◽  
Small Scale ◽  
Whole Genome ◽  
Copy Number Changes ◽  
The Impact

Abstract Saccharomyces cerevisiae is extensively utilized for commercial fermentation, and is also an important biological model; however, its ecology has only recently begun to be understood. Through the use of whole-genome sequencing, the species has been characterized into a number of distinct subpopulations, defined by geographical ranges and industrial uses. Here, the whole-genome sequences of 104 New Zealand (NZ) S. cerevisiae strains, including 52 novel genomes, are analyzed alongside 450 published sequences derived from various global locations. The impact of S. cerevisiae novel range expansion into NZ was investigated and these analyses reveal the positioning of NZ strains as a subgroup to the predominantly European/wine clade. A number of genomic differences with the European group correlate with range expansion into NZ, including 18 highly enriched single-nucleotide polymorphism (SNPs) and novel Ty1/2 insertions. While it is not possible to categorically determine if any genetic differences are due to stochastic process or the operations of natural selection, we suggest that the observation of NZ-specific copy number increases of four sugar transporter genes in the HXT family may reasonably represent an adaptation in the NZ S. cerevisiae subpopulation, and this correlates with the observations of copy number changes during adaptation in small-scale experimental evolution studies.

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