Abstract A60: Analysis of gene copy number changes in chronic lymphocytic leukemia

Abstract

Abstract Chronic lymphocytic leukemia (CLL) is the most frequent type of leukemia in the elderly and is accompanied by the accumulation of malignant monoclonal B-cells in the bone marrow, peripheral blood or lymph nodes which harbor various genetic aberrations. Genomic instability is an important characteristic of leukemia cells and plays a role both in its development as well as in the response to therapy. Somatic copy number changes in distinct genes are associated with the development of specific leukemias. Copy number variations are important diagnostic tools and help in classifying the disease and guiding the therapy. Current strategies for the detection of chromosomal aberrations in CLL are laborious and time consuming. Using multiplex ligation dependent probe amplification (MLPA) copy number changes at multiple loci can be detected simultaneously in a single simple PCR reaction. In this study, we investigated the copy number alterations in several key genes in CLL by MLPA. DNA was isolated from the venous blood samples from 92 patients with CLL and 10 healthy controls. Copy number changes of 20 genes were analyzed at 29 different sites by MLPA. DNA samples were hybridized with the sequence specific probes for each gene and the hybridized probe pairs were ligated. PCR reaction was performed to amplify the ligated probes. PCR products were separated by capillary electrophoresis using the ABI Prism 310 genetic analyzer. The peaks were evaluated for the genomic gains and losses after normalization of each peak. Reference genes were used for normalization of each gene signal. The data from the patients were compared with control samples from healthy individuals. Various genetic aberrations were detected in the patients with CLL. The most frequently observed genetic aberrations were losses in the chromosomal regions 13q14.3 and 13q14.2 and involved the DLEU7 (49 %), DLEU2 (41 %), KCNRG (35.9 %) and RB1 (14 %) genes. Copy number losses in the MYCN, ESR1, EIF3S3, CDKN2A, CDKN2B, ATM, CHFR and TP53 genes were lower with a frequency ranging between 1-10 %. The most frequently amplified gene was the MYC gene (30.4 %), followed by the LRMP (17.4 %), ESR1 (15 %) and CHFR (14 %) genes. Gains in the MYCN, IGF2R, PARK2, CCND2, CDK4, IFNG and LDLR genes were observed with a lower frequency. Our data indicate that copy number changes in several important genes are frequent in the pathogenesis of CLL. Simultaneous analysis and characterization of these genetic alterations can be performed using MLPA and the test can provide valuable clinical information when employed as a first-line screening approach for CLL patients. Citation Format: Nejat Dalay, Mustafa Isin, Guven Cetin, Melih Aktan. Analysis of gene copy number changes in chronic lymphocytic leukemia. [abstract]. In: Proceedings of the AACR Special Conference on Chromatin and Epigenetics in Cancer; Jun 19-22, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2013;73(13 Suppl):Abstract nr A60.