Screening for copy-number alterations and loss of heterozygosity in chronic lymphocytic leukemia—A comparative study of four differently designed, high resolution microarray platforms

2008 ◽  
Vol 47 (8) ◽  
pp. 697-711 ◽  
Author(s):  
Rebeqa Gunnarsson ◽  
Johan Staaf ◽  
Mattias Jansson ◽  
Anne Marie Ottesen ◽  
Hanna Göransson ◽  
...  
Keyword(s):  
High Resolution ◽  
Chronic Lymphocytic Leukemia ◽  
Comparative Study ◽  
Loss Of Heterozygosity ◽  
Copy Number ◽  
Lymphocytic Leukemia ◽  
Copy Number Alterations

Screening for Copy Number Alterations and Loss of Heterozygosity in Chronic Lymphocytic Leukemia - A Comparative Study of Four Differently Designed, High Resolution Microarray Platforms.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2084-2084
Author(s):  
Rebeqa Gunnarsson ◽  
Johan Staaf ◽  
Mattias Jansson ◽  
Anne Marie Ottesen ◽  
Hanna Göransson ◽  
...  
Keyword(s):  
High Resolution ◽  
Chronic Lymphocytic Leukemia ◽  
Loss Of Heterozygosity ◽  
Copy Number ◽  
Lymphocytic Leukemia ◽  
Allelic Loss ◽  
Copy Number Alterations ◽  
Oligonucleotide Arrays ◽  
Snp Arrays ◽  
Trisomy 12

Abstract Screening for copy number alterations (CNA) has improved by applying genome wide microarrays, where SNP-arrays also allow analysis of loss of heterozygosity (LOH). Currently, comparisons of high resolution microarray platforms are few, thus we performed a study to evaluate the power of differently designed microarrays for copy number analysis and LOH. We here analyzed 10 diagnostic chronic lymphocytic leukemia (CLL) samples (five IGVH mutated and five IGVH unmutated) using four different high-resolution platforms: BAC-arrays (32K), oligonucleotide-arrays (185K, Agilent), and two SNP-arrays (250K, Affymetrix and 317K, Illumina). Comparison of copy number data showed that the platforms are concordant in terms of detecting large CNA, including the known recurrent alterations. Mono-allelic and bi-allelic loss of 13q14 (3 and 1 sample, respectively), mono-allelic loss of 11q (1 sample), trisomy 12 (2 samples) and mono-allelic loss of 17p (2 samples) were concordant in all platforms. These aberrations were validated with FISH, which in addition identified subclones with mono-allelic loss of 13q14 in two cases, only detected with the BAC platform, rendering a cut-off for the power of detecting subclones to approximately 25% of investigated cells. As expected, all poor prognostic aberrations were detected in patients carrying unmutated IGHV genes whereas four of five mutated samples were detected with mono-allelic loss of 13q14, as the only recurrent alteration. Furthermore, detection of small CNA were in many cases discordant between platforms. Therefore, we defined alterations identified by at least two platforms and identified 47 losses and 31 gains using this criterion. We are currently validating the presence of a number of these alterations using other techniques. Evaluation of LOH showed concordance for 86 regions between the Illumina and Affymetrix platforms. Of these regions 12 LOH coincided with CNA, leaving the remaining 74 as copy-neutral LOH. In conclusion, all platforms investigated are powerful tools for screening of CNA, however, since non-overlapping CNA were detected by individual platforms, we emphasize the importance of validating findings. Also, there is a cut-off for detecting subclones, here estimated to 25%. Genomic arrays will improve the detection of new recurrent aberrations, which may potentially refine the prognostic hierarchy established by FISH.

High Resolution Screening of Copy-Number Alterations in Chronic Lymphocytic Leukemia Using Affymetrix 250K SNP-Arrays Reveals a Higher Complexity of Genomic Alterations in Patients with Unmutated IGHV Genes

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3133-3133
Author(s):  
Rebeqa Gunnarsson ◽  
Anders Isaksson ◽  
Hanna Göransson ◽  
Larry Mansouri ◽  
Mattias Jansson ◽  
...  
Keyword(s):  
High Resolution ◽  
Copy Number ◽  
Average Length ◽  
Lymphocytic Leukemia ◽  
Copy Number Alterations ◽  
Snp Arrays ◽  
Genetic Complexity ◽  
Trisomy 12 ◽  
Allelic Deletion ◽  
Gains And Losses

Abstract Chronic lymphocytic leukemia (CLL) is a biologically heterogeneous disease where no common genetic aberration so far has been described. Although recurrent genomic aberrations of prognostic significance are well established (e.g. deletions of 11q, 13q, 17p and trisomy 12), less is known about the overall genetic complexity. Recent development of high-resolution single nucleotide polymorphism (SNP)-arrays will allow screening of genetic complexity, which includes detection of smaller copy-number alterations (CNAs) in addition to the known, recurrent aberrations. We here applied the Affymetrix GeneChip® Mapping 250K Nsp arrays and screened peripheral blood samples from 203 newly diagnosed CLL patients (≥70% tumor cells) from a population-based Scandinavian cohort. The male:female ratio was 2:1, the majority of patients was in stage A (70%) and the median age at diagnosis 61 years. Sixty percentage of cases displayed mutated IGHV genes whereas 35% showed unmutated IGHV genes. The Nexus copy-number software (Biodiscovery, Inc.) was applied and CNAs were identified in all CLL samples investigated, where deletions were more commonly detected than gains (in average, 3.5 vs. 2.2 per sample, respectively). The average length of deletions was 3.5 Mb while gains had an average length of 9.5 Mb (3.6 Mb when excluding trisomy 12). More than 50% of deletions and gains had a size ranging between 0.1–1 Mb, whereas 22% of CNAs were less than 0.1 Mb and 25% larger than 1 Mb. When investigating known, recurrent alterations, 105 samples (52%) carried del(13)(q14); 82 samples displayed a mono-allelic deletion, 15 samples a bi-allelic deletion and 8 samples carried 2 mono-allelic deletions of different sizes. The minimal overlapping region was 0.44 Mb and most of the 13q14 deletions covered the genomic region of miR-15, miR-16 and/or DLEU7. Twenty-one samples (10%) showed trisomy 12 and del(11q) was detected in 27 samples (13%), all covering the ATM gene. del(17p) was detected in 7 patients (3.4%) with one patient also carrying several gains and losses of 17q. Moreover, large alterations involving chromosome arms or entire chromosomes were recurrently observed among the samples, for instance, 3 large 8p losses and 4 large 8q gains. Interestingly, 5 samples, which all carried del(11q), displayed a gain covering whole or large parts of the p-arm of chromosome 2, a region which covers MYCN, REL and BCL11A. Furthermore, comparison of samples with different IGHV mutation status (M vs. UM) illustrated a significantly higher frequency of del(11q) (32% of UM vs. 4% of M), trisomy 12 (22% of UM vs. 5% of M) and gain of 2p (12% vs. 0% in M) in samples with unmutated IGHV genes. As expected, patients with mutated IGHV genes showed a higher frequency of del(13q) (61% of M vs. 31% of UM). In addition, large aberrations (>1Mb) were more common in UM samples which in average displayed 0.57 gains and 1.22 deletions compared to 0.26 gains and 0.67 deletions in M samples. In conclusion, with whole-genome screening using high resolution SNP-arrays, higher complexity was revealed in CLL, including a higher number of gains and losses and a higher frequency of larger aberrations in UM compared to M samples. A novel, recurrent combination of del(11q) and gain of 2p was demonstrated which deserves further investigation.

Somatic and Germline Copy Neutral Loss of Heterozygosity Are Common in Chronic Lymphocytic Leukemia

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4567-4567
Author(s):  
Megan Hanna ◽  
Bethany Tesar ◽  
Kristen E. Stevenson ◽  
Alexander R. Vartanov ◽  
Stacey M. Fernandes ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Loss Of Heterozygosity ◽  
Copy Number ◽  
Neutral Loss ◽  
Snp Array ◽  
Lymphocytic Leukemia ◽  
Alternative Mechanism ◽  
Large Set ◽  
The Impact ◽  
13Q Deletion

Abstract Abstract 4567 Chronic lymphocytic leukemia (CLL) is the most common leukemia of adults, and prognosis is still difficult to predict, although cytogenetic abnormalities identified by FISH are most helpful. Isolated reports have suggested that copy neutral loss of heterozygosity (cnLOH) can involve 13q and 17p in CLL, but the extent and the impact on clinical outcome is not well established. We therefore embarked upon characterization of cnLOH in a large set of 230 CLLs with matched normal DNA. The median age at diagnosis of CLL in this patient population was 54 (33–79). 87% of patients were Rai 0–1 at diagnosis, and 79% were chemotherapy naive at sampling. 121 of 230 patients were treated, with a median TTFT of 42 months. The median follow-up for surviving patients is 74 months. 44% of patients carried one somatic copy number abnormality (CNA) by SNP array, 20% two, 7% three, 5% four and 4% more than five. cnLOH was called by the Affymetrix Genotyping Console Software, which evaluates each SNP for copy number and then subtracts the A allele value from the B allele value within an individual sample, thereby allowing independent evaluation of tumor (somatic) and normal (germline). All calls were manually reviewed. A size cut-off of 1.0 Mb was used to determine significant cnLOH events. In total, of 230 patients, we found 26 events of somatic cnLOH (11%) and 36 events of germline cnLOH (16%), affecting 56 separate patients (24%). This frequency of cnLOH was surprisingly high and suggested that cnLOH might be an alternative mechanism affecting known loci in CLL. This was the case, as the most common events overall involved 13q in 25 patients, the X chromosome in 9 patients, chromosomes 17 and 18 in five patients each, and chromosomes 9, 11 and 12 in four patients each. Interestingly, germline events were quite common. Six patients had small regions of germline LOH with much more extensive adjacent somatic LOH, two on chr 13, one on chr 17, two on chr X and one on chr 20; these were coded as germline in the analysis. In addition, of the 25 patients with cnLOH on chromosome 13, 18 of these were in the germline and 7 were somatic. The region(s) of cnLOH were typically adjacent to a 13q deletion, and often involved the entire chromosome arm. Somatic cnLOH at 13q was associated with intermediate sized deletions including the RB gene (p=0.002). Of the 18 patients with germline cnLOH at 13q, 7 of them had no 13q deletion, while 7 had monoallelic deletion and 4 biallelic deletion. Thus 7 patients (3%) had cnLOH events at 13q, in the absence of 13q deletion, again suggesting an alternative mechanism affecting this locus. Germline cnLOH was associated with treatment prior to sampling (44% vs 17%, p<0.001), possibly due to its association with unmutated IGHV(58% vs 32%, p=0.008), and ZAP70 positivity (59% vs 36%, p=0.024). Somatic cnLOH was not associated with any patient characteristics. Neither somatic nor germline cnLOH was associated with >= 1 somatic CNA, but an association between both LOH types and >= 2 somatic CNAs was observed (p=0.053 germline and p=0.030 somatic). TTFT was reduced in patients with either germline cnLOH (61 mos vs 103, p=0.004) or somatic cnLOH (53 mos vs 107, p=0.008). Presence of two or more CNAs was also associated with short TTFT (48 mos vs 115, p<0.001). In order to assess the impact of cnLOH and CNAs on outcome independent of prior therapy, we evaluated TTFT in the 181 chemotherapy naive patients. In this subgroup, germline cnLOH was not associated with short TTFT, while somatic cnLOH (80 mos vs 125, p=0.018) and two or more somatic CNAs (80 mos vs 125, p=0.009) were. In multivariable Cox modeling including germline cnLOH, IGHV, and del 11q or 17p by FISH, the only significant predictor of TTFT was unmutated IGHV (hazard ratio (HR) 4.48, p<0.001). In multivariable Cox modeling including somatic cnLOH and the variables above, the only significant predictor of TTFT was again unmutated IGHV (HR 4.41, p<0.001). When the presence of two or more somatic CNAs was added to these models, this variable was significant along with IGHV (HR 2.04, p=0.009 in germline model; HR 1.84, p=0.033 in somatic model). We conclude that both somatic and germline cnLOH are common in CLL, affecting one quarter of patients in this dataset, and frequently involve chromosomal regions known to be important in CLL. cnLOH is associated with increased somatic CNAs and unmutated IGHV, and therefore poor prognosis. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Investigation of MDM2 Oncogene Copy Number Alterations in Cases of Chronic Lymphocytic Leukemia

2019 ◽  
Vol 36 (2) ◽  
pp. 126-127
Author(s):  
Şule Darbaş ◽  
Çiğdem Aydın ◽  
Ozan Salim ◽  
Sibel Berke Karaüzüm
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Copy Number ◽  
Lymphocytic Leukemia ◽  
Copy Number Alterations

scholarly journals Investigation of MDM2 Oncogene Copy Number Alterations in Cases of Chronic Lymphocytic Leukemia

2019 ◽  
Vol 36 (2) ◽  
pp. 126-127
Author(s):  
Şule Darbaş ◽  
Çiğdem Aydın ◽  
Ozan Salim ◽  
Sibel Berke Karaüzüm
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Copy Number ◽  
Lymphocytic Leukemia ◽  
Copy Number Alterations

scholarly journals High-resolution genomic profiling of chronic lymphocytic leukemia reveals new recurrent genomic alterations

Blood ◽  
2012 ◽  
Vol 120 (24) ◽  
pp. 4783-4794 ◽  
Author(s):  
Jennifer Edelmann ◽  
Karlheinz Holzmann ◽  
Florian Miller ◽  
Dirk Winkler ◽  
Andreas Bühler ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Copy Number ◽  
Single Nucleotide Polymorphism Array ◽  
Neutral Loss ◽  
Lymphocytic Leukemia ◽  
Copy Number Alterations ◽  
Genomic Alterations ◽  
Single Nucleotide ◽  
Truncating Mutation

Abstract To identify genomic alterations in chronic lymphocytic leukemia (CLL), we performed single-nucleotide polymorphism–array analysis using Affymetrix Version 6.0 on 353 samples from untreated patients entered in the CLL8 treatment trial. Based on paired-sample analysis (n = 144), a mean of 1.8 copy number alterations per patient were identified; approximately 60% of patients carried no copy number alterations other than those detected by fluorescence in situ hybridization analysis. Copy-neutral loss-of-heterozygosity was detected in 6% of CLL patients and was found most frequently on 13q, 17p, and 11q. Minimally deleted regions were refined on 13q14 (deleted in 61% of patients) to the DLEU1 and DLEU2 genes, on 11q22.3 (27% of patients) to ATM, on 2p16.1-2p15 (gained in 7% of patients) to a 1.9-Mb fragment containing 9 genes, and on 8q24.21 (5% of patients) to a segment 486 kb proximal to the MYC locus. 13q deletions exhibited proximal and distal breakpoint cluster regions. Among the most common novel lesions were deletions at 15q15.1 (4% of patients), with the smallest deletion (70.48 kb) found in the MGA locus. Sequence analysis of MGA in 59 samples revealed a truncating mutation in one CLL patient lacking a 15q deletion. MNT at 17p13.3, which in addition to MGA and MYC encodes for the network of MAX-interacting proteins, was also deleted recurrently.

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