-Author name in bold denotes the presenting author
-Asterisk * with author name denotes a Non-ASH member
Clinically Relevant Abstract denotes an abstract that is clinically relevant.

PhD Trainee denotes that this is a recommended PHD Trainee Session.

Ticketed Session denotes that this is a ticketed session.

89 Dysregulation of DNA Methylation Involves in Progression of Myeloid Leukemia in Down Syndrome

Acute Myeloid Leukemia: Biology, Cytogenetics and Molecular Markers in Diagnosis and Prognosis
Program: Oral and Poster Abstracts
Type: Oral
Session: 617. Acute Myeloid Leukemia: Biology, Cytogenetics and Molecular Markers in Diagnosis and Prognosis: Omics-based Approaches for the Identification of Novel Genetic Modalities in AML
Saturday, December 5, 2015: 1:00 PM
Tangerine 3 (WF3-4), Level 2 (Orange County Convention Center)

Satoshi Saida, MD, PhD1,2*, Masahiro Nakamura, MD, PhD3*, Tsutomu Toki, PhD4, Yoko Nishinaka-Arai, Ph.D.5*, Kiminori Terui, MD, PhD4*, Kenichi Yoshida, MD, Ph.D.6*, Seishi Ogawa, MD, Ph.D.6, Tatsutoshi Nakahata, MD, PhD2,5, Toshio Heike, MD, PhD1,2*, Ken-ichiro Watanabe, MD, PhD1,2,7*, Akira Watanabe, PhD2,3* and Etsuro Ito, MD, PhD2,4

1Department of Pediatrics, Graduate School of Medicine Kyoto University, Kyoto, Japan
2AMED-CREST, Japan Agency for Medical Research and Development, 1-7-1 Otemachi, Chiyoda-ku, Tokyo 100-0004, Tokyo, Japan
3Department of Life Science Frontiers, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
4Department of Pediatrics, Hirosaki University Graduate School of Medicine, Hirosaki, Japan
5Clinical Application Department, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
6Department of Pathology and Tumor Biology, Graduate School of Medicine, Kyoto University, Kyoto, Japan
7Division of Hematology and Oncology, Shizuoka Children's Hospital, Shizuoka, Japan

Transient abnormal myelopoiesis (TAM) is a clonal pre-leukemic disorder in neonates of Down syndrome. Although self-limiting in a majority of cases, TAM may evolve as non-self-limiting acute megakaryoblastic leukemia (DS-AMKL) after spontaneous remission. As such, the study of TAM/DS-AMKL can provide insight into the mechanisms of leukemic evolution. Our recent study has shown that DS-AMKL from TAM is shaped by acquisition of new mutations and clonal selection, and genomic mutations in epigenetic regulators frequently preceded other driver mutations. In this study, we address biological significance of the mutations in a progression from TAM to DS-AMKL using a newly established xenograft model of malignant conversion of the leukemia.

We first analyzed global DNA methylation profile of clinical samples including 21 TAM and 10 DS-AMKL, using Infiniun 450K DNA methylation array (Illumina). Although averaged DNA methylation levels of both TAM and DS-AMKL were approximately the same, we observed two distinct clusters delineating TAM and DS-AMKL samples by both hierarchical clustering and principle component analysis (PCA). We identified differentially methylated cytosines on the promoters (TAM/AMKL-DMCpGs) between the two groups, and found that most of TAM/AMKL-DMCpGs were hyper-methylated in DS-AMKL samples compared to TAM, indicating that hyper-methylation of the subset of CpGs is a hallmark of DS-AMKL.

To follow sequential alterations of DNA methylation profile, we established a xenograft model of the leukemia, which mimicked the progression of TAM to DS-AMKL. We transplanted TAM blast cells (1x106 cells) derived from the patient, who developed DS-AMKL later, into NOD/SCID/γc null (NOG) mice through tail vein. One series of the xenografts exhibited engraftment in the mouse and could passage by serial engraftment in vivo, whereas the other exhibited engraftment, but could not maintain in the mice by serial transplantation. The global DNA methylation profile of engrafted cells from the first generation mice was closed to TAM patients, whereas those from the forth, fifth and sixth generations approached to DS-AMKL patients. Moreover, hyper-methylation of TAM/AMKL-DMCpGs was also observed in engrafted TAM cells with higher numbers of passages, demonstrated that the xenograft model mimicked dysregulation of DNA methylation in disease progression of TAM. We further investigated the impact of dysregulation of DNA methylation on gene expression. As expected, RNA-seq and hierarchal clustering analyses of genes regulated by TAM/AMKL-DMCpGs displayed distinct two clusters, one with TAM samples and the first generation engraftments and another with DS-AMKL samples and 4th, 5th and 6thgeneration engraftments.

We propose that epigenetic dysregulation is a crucial step for malignant conversion of TAM into DS-AMKL.

Disclosures: No relevant conflicts of interest to declare.

*signifies non-member of ASH