Role of ASXL1 mutations in myeloid malignancies Addition of Sex Combs-like 1 (ASXL1) is an Enhancer of Trithorax and Polycomb (ETP) protein as ASXL1 loss in Drosophila causes homeotic transformations and alterations in Trithorax and Polycomb- mediated gene expression. Recently, recurrent mutations in ASXL1 were discovered in patients with myeloid malignancies, including myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPN), and acute myeloid leukemia (AML). Moreover, we have identified from high-throughput sequencing studies of patients with MDS, MPN, and de novo AML that mutations in ASXL1 are associated with significantly poorer overall survival in uniformly treated patient cohorts. This data suggests that mutations in ASXL1 represent novel markers of adverse outcome in myeloid malignancies. We therefore have performed several studies to characterize effects of ASXL1 loss and overexpression of wild-type and mutant ASXL1 on hematopoiesis. To study loss of ASXL1 function, we performed gene expression analyses following shRNA knockdown (KD) of ASXL1 in the ASXL1 wild-type leukemia cell lines, UKE1 and SET2 as well as in CD34+ hematopoietic stem/progenitors. In unbiased analyses we observed significant and reproducible increases in HOXA9, HOXA5, and MLL expression with ASXL1 loss following KD of ASXL1. ChIP (chromatin immunoprecipitation) of HOXA9 and MLL promoters revealed enrichment of the transcriptional activating mark H3K4me3 and decreases in H3K9me3, a repressive mark, at these promoters. ChIP-Seq studies in UKE1 cells with ASXL1 KD similarly reveal a striking decrease in the H3K27me3 repressive mark with ASXL1 loss compared with control cells. Moreover, ChIP using an anti-ASXL1 antibody reveals that ASXL1 itself is present at the promoter of MLL and HOXA5. Our data suggests that ASXL1 mutations (a) promote aberrant hematopoiesis by dysregulated expression of Hox genes and (b) affect target gene expression directly, and by altering the chromatin state of promoters. We propose a series of studies with the following specific aims:
Specific Aim #1 : Identify target genes of ASXL1 loss and alterations in chromatin state due to ASXL1 loss in normal and leukemic hematopoietic cells using in vitro assays.
Specific Aim #2 : Determine the in vivo effects of ASXL1 loss on hematopoiesis by targeted disruption of ASXL1.
Specific Aim #3 : Determine the role of ASXL1 mutations in the pathogenesis of MPNs and AML through studies in patient samples and in in vivo models in cooperation with known MPN and AML disease alleles.

Public Health Relevance

Mutations in the gene ASXL1 have recently been identified in patients with a variety of myeloid blood cancers. Through careful analysis of this gene in independent sets of patients uniformly treated for these disorders, we have identified that ASXL1 mutations are associated with adverse outcome in leukemia patients. In this project we describe a series of studies aimed at improving our understanding of how ASXL1 mutations contribute to leukemia and to poor outcome in patients with these malignancies. The written critiques and criteria scores of individual reviewers are provided in essentially unedited form in the Critique section below. Please note that these critiques and criteria scores were prepared prior to the meeting and may not have been revised subsequent to any discussions at the review meeting. The Resume and Summary of Discussion section above summarizes the final opinions of the committee.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Clinical Investigator Award (CIA) (K08)
Project #
Application #
Study Section
Subcommittee G - Education (NCI)
Program Officer
Perkins, Susan N
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Sloan-Kettering Institute for Cancer Research
New York
United States
Zip Code
Inoue, D; Kitaura, J; Matsui, H et al. (2015) SETBP1 mutations drive leukemic transformation in ASXL1-mutated MDS. Leukemia 29:847-57
Chung, Young Rock; Kim, Eunhee; Abdel-Wahab, Omar (2014) Femoral bone marrow aspiration in live mice. J Vis Exp :
Micol, Jean-Baptiste; Abdel-Wahab, Omar (2014) Collaborating constitutive and somatic genetic events in myeloid malignancies: ASXL1 mutations in patients with germline GATA2 mutations. Haematologica 99:201-3
Lee, Stanley Chun-Wei; Abdel-Wahab, Omar (2014) The mutational landscape of paroxysmal nocturnal hemoglobinuria revealed: new insights into clonal dominance. J Clin Invest 124:4227-30
Abdel-Wahab, Omar; Klimek, Virginia M; Gaskell, Alisa A et al. (2014) Efficacy of intermittent combined RAF and MEK inhibition in a patient with concurrent BRAF- and NRAS-mutant malignancies. Cancer Discov 4:538-45
Chung, Stephen S; Kim, Eunhee; Park, Jae H et al. (2014) Hematopoietic stem cell origin of BRAFV600E mutations in hairy cell leukemia. Sci Transl Med 6:238ra71
Micol, Jean-Baptiste; Duployez, Nicolas; Boissel, Nicolas et al. (2014) Frequent ASXL2 mutations in acute myeloid leukemia patients with t(8;21)/RUNX1-RUNX1T1 chromosomal translocations. Blood 124:1445-9
Abdel-Wahab, Omar; Levine, Ross L (2013) Mutations in epigenetic modifiers in the pathogenesis and therapy of acute myeloid leukemia. Blood 121:3563-72
Beguelin, Wendy; Popovic, Relja; Teater, Matt et al. (2013) EZH2 is required for germinal center formation and somatic EZH2 mutations promote lymphoid transformation. Cancer Cell 23:677-92
Quintas-Cardama, Alfonso; Abdel-Wahab, Omar; Manshouri, Taghi et al. (2013) Molecular analysis of patients with polycythemia vera or essential thrombocythemia receiving pegylated interferon *-2a. Blood 122:893-901

Showing the most recent 10 out of 12 publications