Abstract 2989: Loss of methylation in a regulatory region of BCL2 in MDS/AML.

Abstract

Abstract Background: Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic disorders characterized by ineffective hematopoiesis and increased risk of progression to acute myeloid leukemia (AML). A hallmark of MDS is increased apoptosis, while AML undergoes decreased apoptosis and increased proliferation. BCL2, an anti-apoptotic protein, is overexpressed in high grade MDS and AML and may confer poor prognosis and increased chemotherapy resistance. In this study we examined the DNA methylation status of BCL2 in MDS and AML. Methods: We conducted DNA methylation profiling on genomic DNA extracted from bone marrows of 22 refractory anemia with ringed sideroblasts (RARS), 8 refractory anemia with excess blasts (RAEB), 21 de novo AML, 21 therapy related AML, and 7 normal volunteers. We used both Illumina 27K BeadChip and 450K BeadChip methylation arrays to assess CpG methylation. Standard student T-test was used to screen differentially methylated MDS and AML CpG sites compared to normal samples. We validated the microarray data by combined bisulfite restriction analysis (COBRA) and bisulfite sequencing. Results: We identified a region in the BCL2 gene body that was among the most differentially methylated in MDS and AML versus normal bone marrow. Within that amplicon, BCL2 in both RARS, a low grade MDS, and RAEB, a high grade MDS, were hypermethylated compared to normal bone marrow (43% vs 18%, 48% vs 18%, respectively, p<0.05). BCL2 in de novo AML and therapy related AML were hypomethylated compared to normal bone marrow (12% vs 35%, 13% vs 35%, respectively, p<0.05). We validated these results by COBRA and bisulfite sequencing. This region coincided with H3K27ac and H3K4m1 enhancer type marks in ENCODE data in normal cells. Conclusions: Our study identifies a region of BCL2 that is hypermethylated in both low and high risk MDS while hypomethylated in AML compared to normal bone marrows. We hypothesize aberrant epigenetic modification of BCL2 plays a key role in apoptosis and transformation from MDS to AML, and our data support the idea of combining demethylating drugs with BCL2 antagonists. We are currently investigating whether BCL2 protein expression correlates with methylation, and plan to map the histone marks with CHIP-seq to correlate chromatin accessibility and DNA methylation in BCL2. Citation Format: Sangmin Lee, Naomi Galili, Stephen Nimer, Mark D. Minden, Vundavalli V. Murty, Suresh Jhanwar, Azra Raza, Benjamin Tycko. Loss of methylation in a regulatory region of BCL2 in MDS/AML. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2989. doi:10.1158/1538-7445.AM2013-2989