Identification of acquired copy number alterations and uniparental disomies in cytogenetically normal acute myeloid leukemia using high-resolution single-nucleotide polymorphism analysis

Abstract Abstract 2929 Background: Azacitidine (AZA) has changed the outcome of patients (pts) with myelodysplastic syndromes (MDS) or acute myeloid leukemia with multi-lineage dysplasia (AML-MLD) unfit for intensive chemotherapy. AZA is a hypomethylating agent providing about 50% of responses in MDS and AML with low blast count (Fenaux et al., Lancet Oncol 2009, JCO 2010). To date, no consensus genetic predictor of response has been reported. Methods: In MDS (including RAEB-t) and AML-MLD (>30% blasts) patients treated by AZA in 5 centers, we performed genomewide single nucleotide polymorphism (SNP) analysis using SNP 6.0 arrays (Affymetrix, High Wycombe, U.K.) on bone marrow (BM) samples. Patients having received ≥ 1 cycle of AZA and who had bone marrow evaluation after ≥ 4 cycles, or who died or progressed before completion of 4 cycles were considered evaluable (the last 2 groups were considered as treatment failures). Responses were scored according to IWG 2006 criteria for MDS and to Cheson et al. (JCO 2003) for AML. DNA were prepared for hybridization according to the manufacturers' recommendations. Affymetrix CEL files for each sample were analyzed using the Genotyping Console software (v3.0.2). Genotyping was performed using Birdseed V2 algorithm. Unpaired Copy Number and LOH analysis was performed with Regional GC correction. Copy number and UPD were also analyzed using the Copy Number Analyzer for GeneChip (CNAG version 3.3.0.1) algorithm (http://www.genome.umin.jp/CNAGtop2.html). Results: The study population included 92 pts: F/M: 41/51; median age 72 (range 35–88). Diagnosis at AZA onset was MDS in 54 (RAEB-1 n=6, RAEB-2 n=37, RAEB-t=11, IPSS int-1 in 7, int-2 in 15, high in 32, undetermined in 2) and AML-MLD in 38 pts. Cytogenetic according to IPSS was favorable in 33, intermediate in 19, unfavorable in 28, unknown in 11. Median number of cycles was 6 (range 1–41). All pts received the approved (75 mg/m2 for 7 days every 4 weeks) or a reduced AZA schedule (75 mg/m2 for 5 days every 4 weeks). Median overall survival (OS) of our cohort was 22 months. DNA samples from 52 patients were available for SNP analysis. There were no significant differences between the SNP and no SNP subgroups in term of median age, sex ratio, disease status at AZA onset, cytogenetic according to IPSS, median number of cycles, responses and OS. We listed aneuploidies (CNA) and uniparental disomies (UPD) detected by SNP analysis in the samples, and focused on 18 chromosomal bands (1p13.2, 2q34, 3p14.2, 3q26, 4q24, 5q33.1, 5q35, 6p21.3, 7q36.1, 9p21.3, 11p13, 11q23.3, 13q12.2, 15q21, 15q26.2, 20q11.21, 21q22.1) containing genes implicated in MDS or AML. Preliminary results show correlations between some CNA/UPD and response to AZA and OS, such as 1/a UPD at 9p21 (6% of patients) associated to a better hematologic improvement (X2, p=0.037), 2/an amplification at 20q11.21 (8% of patients) correlated with a poorer response (X2, p=0.048) and a trend toward poorer OS (Log-Rank, p=0.091), and 3/deletion and UPD at 15q21 (6% of patients) strongly correlated to poorer OS (Log-Rank, p=0.001). Several additional CNA and UPD of potential interest are presently under investigation. Conclusion: SNP analysis using SNP 6.0 is a powerful tool to decipher genome complexity in BM samples of MDS and AML patients treated with AZA. Our data suggest that this approach could allow characterizing profiles of responder versus non responder pts. Our study must be extended to a larger cohort and relevant anomalies must be confirmed by more sensitive techniques such as high-scale sequencing. Disclosures: Cluzeau: Celgene: Consultancy. Raynaud:Celgene: Consultancy.

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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 ◽

10.1182/blood.v110.11.2838.2838 ◽

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.

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