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Pediatric Acute Myeloid Leukemia (AML) – Section 14

Myeloid Pre-Leukemia and Leukemia of

David Cruz Hernandez1,2, Paresh Vyas1,2 1 MRC MHU, BRC Hematology Theme, Oxford Biomedical Research Centre, Oxford Centre for Hematology, WIMM, Radcliffe Department of Medicine, University of Oxford, Oxford OX3 9DU, UK; 2 Department of Hematology, Oxford University Hospitals NHS Trust, Oxford OX3 7LE, UK

Take-home messages:  Maturation arrest is a hallmark of myeloid leukemia and remains largely unexplored at the molecular level.  ML-DS is an excellent model to conduct mechanistic studies due to its genetic simplicity to explore maturation arrest and mutation cooperativity.

Introduction Current state of the art A hallmark of blood cancers is the gradual acquisition of genetic Trisomy 21 alters HSC homeostasis or epigenetic changes that progressively disrupts normal hemato- The initiating genetic alteration in the multi-step model of poiesis and eventually leads to full transformation. Therefore, ML-DS is an extra copy of 21 (T21) in fetal most leukaemias are preceded by a pre-leukemic phase. For hematopoietic stem cells. T21 causes expansion of the immuno- example, Myeloid Leukemia of Down Syndrome (ML-DS) is an phenotypically defined fetal-liver HSC compartment with skewed acute leukemia with megakaryoblastic and erythroid features that cellular differentiation toward the erythroid/megakaryocytic clonally evolves from a preleukemic condition referred as lineage, and conversely sever impairment∗ of B-lymphopoiesis Transient Abnormal Myelopoiesis or TAM. The pathogenesis and reduced granulocyte-monocyte output. 2 A cell-autonomous of ML-DS is well characterized and consists of 3 key temporally 1 imbalance due to T21 might be responsible for perturbation separated stages. First, partial or complete trisomy 21 (T21) of fetal hematopoiesis. In the few cases where ML-DS develops alters hematopoietic stem cell (HSC) homeostasis in fetal liver. fi from partial T21 suggest that the leukemic∗ risk is con ned to an Second, somatic mutations acquired during fetal life in the key 8.5Mb region on 3 where key hematopoietic transcription factor GATA1 cooperate with T21 and result in the lie, such as RUNX1, ETS2, ERG, DYRK1A. Unfortunately, T pre-leukemic myeloproliferative disorder known as ransient it is challenging to identify differences between Abnormal Myelopoiesis, TAM. Third, a single allele mutation in aneuploid and disomic cells mainly due to modest differences that genes encoding the cohesin complex, epigenetic regulators or can be masked by patient variability. The molecular mechanism in the JAK family kinases transforms the TAM clone to full-blown by which T21 alters HSC homeostasis and predisposes to ML-DS leukemia. Due to its genetic simplicity, ML-DS and its precursor remains largely unexplored. clonal disease TAM, will server as a disease model to define the molecular mechanism leading to maturation arrest (Fig. 1). Aberrant transcription factor function due to GATA1s The second step on the evolution of ML-DS requires mutations in the X-chromosome encoded erythro-megakaryocyte transcription factor GATA1. In normal GATA1-expressing cells, 2 GATA1 isoforms are detected; a full-length 414 amino-acid (aa) This work was supported by Lady Tata Memorial Trust (DCH). PV is supported by (GATA1fl) and an N-terminal truncated 331-aa protein, GATA1s. Bloodwise Specialist Programme Grant 13001 and by the NIHR Oxford Somatic GATA1 mutations identified in TAM and ML-DS fl Biomedical Centre Research Fund. abrogate∗ GATA1 production but leave GATA1s∗ expression The authors in fact have no conflict of interest to disclose. intact. 4 Experimental evidence from murine modes 5,6,7 as well as Copyright © 2020 the Author(s). Published by Wolters Kluwer Health, Inc. on from patient-derived induced pluripotent cells,8 demonstrate that behalf of the European Hematology Association. This is an open access article exclusive expression of GATA1s promotes megakaryopoiesis at the distributed under the terms of the Creative Commons Attribution-Non expense of erythropoiesis. At the molecular level, Gata1s fails to Commercial-No Derivatives License 4.0 (CCBY-NC-ND), where it is permissible to repress Gata2 and Runx1 transcription despite both Gata1 and download and share the work provided it is properly cited. The work cannot be 6 changed in any way or used commercially without permission from the journal. Gata1s having similar chromatin occupancy at both loci. GATA2 HemaSphere (2020) 4:S2 and RUNX1 expression are required for hematopoietic stem and Received: 2 April 2020 / Accepted: 6 April 2020 progenitor cells (HSPCs) to balance self-renewal and differentiation

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Hernandez and Vyas Myeloid Leukemia of Down Syndrome

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Figure 1. Myeloid Leukaemia of Down Syndrome (ML-DS) as multistep disease model of leukemogenesis. Trisomy 21 is the initial genetic change in ML-DS leukemogenesis. It leads to expansion of hematopoietic stem cells and the erythroid/megakaryocytic compartment but impairment of lymphopoiesis. It is expected that, close to 30% of Down syndrome neonates acquire mutations in the GATA1 during fetal life. These mutations abrogate expression of the full-length GATA1 isoform but lave expression of the short GATA1 isoform intact inducing a myeloproliferative disorder known as TAM. 10% of TAM neonates develop ML-DS before the age of 4 after acquiring additional secondary mutations.

but their expression is normally down-regulated during terminal required to produce the high cohesin mutation frequency observed – differentiation.9 11 Moreover, RUNX1 has been shown to play a in ML-DS. fundamental role during lineage fate decisions at the erythroid/ megakaryocytic branching point by repressing KLF1 transcription, Future perspectives an erythroid specific transcription factor and promoting a megakaryocytic, FLI1-driven, gene expression program.11 Howev- Transformation of human preleukemia to leukemia is not er, since similar GATA1 mutations are not leukemogenic in the entirely understood. ML-DS and its precursor disease TAM are absence of trisomy 21, the key constitutional abnormality present in genetically simple models where to study the transforming genetic all patients with Down syndrome, the interaction between trisomy and epigenetic changes to define maturation arrest at the 21 and GATA1s is clearly of importance. It is still not clear if molecular level. Despite our understanding of role of GATA1 GATA1s inability to repress at least two key transcription factors, and the mutation landscape in ML-DS, there are many questions GATA2 and RUNX1, cooperates with trisomy 21 to induce cellular left unexplored. Why do GATA1 mutant progenitor cells have a maturation delay. remarkable proliferative advantage during T21 fetal life? Why are N’ terminally truncated GATA1 mutations virtually absent from Altered chromatin accessibility and activated signaling adult leukemia? Does the function of GATA1 changes according in ML-DS to ontogeny? Why there is a higher frequency of mutations in the cohesin complex in ML-DS than in adult acute myeloid leukemia? Finally, transformation of TAM to ML-DS is most likely due to a Nonetheless, with the mutational landscape of TAM and ML-DS single allele mutation in genes encoding the cohesin complex (47%), defined as well as understanding of ML-DS pathogenesis new JAK MPL KIT the family kinases, , (48%) or epigenetic∗ regulators concept will emerge if mechanistic studies are performed on such as KANSL1, EZH2, and SUZ12 (36%). 12 This mutation oncogenic cooperativity. A full understanding of oncogenic frequency in ML-DS samples illustrates that transformation occurs cooperation is necessary to prevent progression of pre-leukemia by cooperation between aberrant transcription factor function due to transformed leukemia. to exclusive GATA1s expression and activated signaling and/or deregulation of epigenetic process. With the advent of gene editing References technologies such as CRISPR/Cas9, it becomes possible to functionally annotate the genetic variants during transformation 1. Garnett C, Cruz Hernandez D, Vyas P. GATA1 and cooperating mutations in myeloid leukaemia of Down syndrome. IUBMB life. of TAM to ML-DS. A murine model that closely resembles Gata1s – driven pre-leukemia served as a platform on which to interrogate 2020;72:119 130. ∗2. Roy A, Cowan G, Mead AJ, et al. Perturbation of fetal liver predicted∗ loss-of-function variants required to induce frank 12 hematopoietic stem and progenitor cell development by trisomy 21. leukemia. In this murine model of frank leukemia, a similar Proc Natl Acad Sci. 2012;109:17579–17584. frequency of variants inducing activated signaling (32%) were Roy A et al, shows that an extra copy of chromosome 21 perturbs stem cell observed. There was also an over representation of mutations in homeostasis in fetal hematopoietic cells by enhancing megakaryocyte/ epigenetic regulators (40%). Surprisingly, there was a reduced erythroid output and reducing B-cell lineage output. mutational frequency in cohesin complex (5%), a four-protein ring- ∗3. Korbel JO, Tirosh-Wagner T, Urban AE, et al. The genetic shaped structure necessary to bring distant regulatory units in close architecture of Down syndrome phenotypes revealed by high- proximity to promoters through DNA-loop formation. This tells us resolution analysis of human segmental trisomies. Proc Natl Acad that cohesin mutations might not be easily studied in murine tissue Sci U S A. 2009;106:12031–12036. because of the difference in chromosome architecture between Korbel JO et al, shows that the increase risk for myeloid leukemia in Down species. Moreover, it suggests that trisomy 21 background might be syndrome individuals is confined to 8.35 megabases in chromosome 21.

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Hernandez and Vyas Myeloid Leukemia of Down Syndrome

∗4. Wechsler J, Greene M, McDevitt MA, et al. Acquired mutations in 8. Byrska-Bishop M, VanDorn D, Campbell AE, et al. Pluripotent stem GATA1 in the megakaryoblastic leukemia of Down syndrome. Nat cells reveal erythroid-specific activities of the GATA1 N-terminus. J Genet. 2002;32:148–152. Clin Invest. 2015;125:993–1005. The genetic alteration that promote acute megakaryoblastic leukemia in 9. Briegel K, Lim KC, Plank C, et al. Ectopic expression of a Down syndrome remained elusive until Wechsler J et al, demonstrated that conditional GATA-2/estrogen receptor chimera arrests erythroid leukemic cells ML-DS contain mutation in the GATA1 locus that lead to differentiation in a hormone-dependent manner. Genes Dev. loss of the GATA1full-length isoform. 1993;7:1097–1109. ∗5. Li Z, Godinho FJ, Klusmann J-H, et al. Developmental stage- 10. Ikonomi P, Rivera CE, Riordan M, et al. Overexpression of GATA-2 selective effect of somatically mutated leukemogenic transcription inhibits erythroid and promotes megakaryocyte differentiation. Exp factor GATA1. Nat Genet. 2005;37:613–619. Hematol. 2000;28:1423–1431. Zhe Li et al, developed the first mouse model of exclusive expression of the 11. Kuvardina ON, Herglotz J, Kolodziej S, et al. RUNX1 represses the short isoform of Gata1. This model demonstrates and enhanced erythroid gene expression program during megakaryocytic differen- proliferation of the megakaryocyte lineage and conversely the erythroid tiation. Blood. 2015;125:3570–3579. lineage is markedly diminished. ∗12. Labuhn M, Perkins K, Matzk S, et al. Mechanisms of 6. Ling T, Birger Y, Stankiewicz MJ, et al. Chromatin occupancy and progression of myeloid preleukemia to transformed myeloid epigenetic analysis reveal new insights into the function of the leukemia in children with down syndrome. Cancer Cell. GATA1N terminus in erythropoiesis. Blood. 2019;134:1619–1631. 2019;36:340–1340. 7. Shimizu R, Takahashi S, Ohneda K, et al. In vivo requirements for Labuhn et al provide the most comprehensive functional interrogation of GATA-1 functional domains during primitive and definitive the genetic variants of pre-leukemia and leukemia in ML-DS. erythropoiesis. EMBO J. 2001;20:5250–5260.

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