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ANR funded project

JCJC : Sciences de la vie, de la santé et des écosystèmes : Physiologie, métabolisme, physiopathologie, santé publique (JCJC SVSE 1) 2010
Projet FPDAML

Role of AML1 in pathological hematopoiesis linked to the familial platelet disorder with a predisposition to acute myeloid leukaemia (FPD/AML)

The transcription factor AML1 (RUNX1) is the alpha subunit of the core binding factor (CBF) and one of the most frequent targets of chromosomal and genetic alteration in leukemias. AML1 contains a RUNT homology domain (RHD) that mediates DNA binding and heterodimerization with CBFß, and a C-terminal domain responsible for transcriptional activation. AML1 is essential for establishment of definitive hematopoiesis, has a dose dependent role in regulating the quantity of HSC and of their downstream committed progenitors, as well as a more restricted role on T cell development and platelet formation.
To explore the role of AML1 mutations in the leukemic development and thrombopenia induction, a genetic hereditary pathology: Familial platelet disorder with a predisposition to acute myeloid leukaemia (FPD/AML) represents an important model.
FPD/AML is an inherited autosomal dominant platelet disorder. The candidate gene is AML1 because different types of mutations were always found at the heterozygous state in 27 pedigrees.
The incidence of leukemia among FPD/AML affected individuals varies from 20% to 50%. Mutations implicate always the Runt domain and lead either to the absence or decrease in binding of mutant protein to DNA but:
1 - some mutations generate proteins that retain the capacity to heterodimerize with CBFß and inhibit wt AML1 transcription activity. In these cases, the mutant may behave as a dominant-negative (DN) protein.
2 – other mutations generate AML1 haploinsufficiency.
The studies performed in very few patients suggest that leukemia incidence appears to be lower in pedigrees with haploinsufficiency. The exact mechanisms by which only some genetic alterations in AML1 predispose to acute leukemia remain unknown.

Our hypothesis based on our preliminary results is that dominant negative mutations in AML1 induce a pre-leukemic state by deregulating hematopoietic stem cells and progenitors and increasing the compartment of target cells (GM-progenitors) available for secondary mutations leading to full blown leukemia whereas haplo-insufficiency does not significantly deregulate hematopoiesis and therefore does not predispose to leukemia (as suggested by mouse models). The type of mutation should thus predict the risk of AML transformation.

We proposed to perform a detailed in vitro study of hematopoiesis (multipotent and committed progenitors) in individuals belonging to five FPD/AML pedigrees with different types of mutations (informed consent obtained) and to correlate AML1 abnormalities with abnormal regulation of AML1 primary target genes. The transcriptome will be studied in patient hematopoietic progenitors and identification of AML1 target genes will be facilitated by AML1 knockdown and by identification of promoters binding AML1 (by ChIP-seq method) in control hematopoietic progenitors. This complex approach has already been started and our preliminary results (identification of NR4A3, NOV, p19 and ZBTB1) confirm that it is feasible.
The AML1 target genes and their implication in pathology will be validated in two different types of model: murine models (conditional AML1 KO, KO of candidate gene) and a human model (pluripotent stem cells that we are actually inducting from patient skin biopsies).
Contrary to the leukemic predisposition, thrombopenia, even moderate, is present in all pedigrees regardless of the mutation type suggesting a dose depending regulation of megakaryopoiesis and platelet generation. Identification of anomalies of patient megakaryopoiesis as well as identification and validation of AML1 target genes in megakaryocytes are also an important objective of these grant and should be helpful in a better understanding of molecular mechanisms regulating platelet production.

Partners

INSERM INSTITUT GUSTAVE ROUSSY

ANR grant: 367 380 euros
Beginning and duration: - 48 mois

 

ANR Programme: JCJC : Sciences de la vie, de la santé et des écosystèmes : Physiologie, métabolisme, physiopathologie, santé publique (JCJC SVSE 1) 2010

Project ID: ANR-10-JCJC-1109

Project coordinator:
Madame Hana RASLOVA (INSTITUT GUSTAVE ROUSSY)
hraslova@nulligr.fr

 

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The project coordinator is the author of this abstract and is therefore responsible for the content of the summary. The ANR disclaims all responsibility in connection with its content.