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

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 >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.