Hematological Phenotypes in GATA2 Deficiency Syndrome Arise from Secondary Injuries and Maladaptation to Proliferation
Authors: Fernandez Orth Juncal , Koyunlar Cansu , Weiss Julia Miriam , Gioacchino Emanuele , De Looper Hans , Andrieux Geoffroy , Ter Borg Mariëtte , Zink Joke , Gonzalez-Menendez Irene , Hoogenboezem Remco , Yigit Baris , Gussinklo Kirsten , Mulet-Lazaro Roger , Raaijmakers Marc , Menendez Irene Gonzalez , Wantzen Charlotte , Pfeiffer Sophie , Molnar Christian , Bindels Eric , Bohler Sheila , Sanders Mathijs Arnoud , Quintanilla-Fend Leticia , Wlodarski Marcin , Boerries Melanie , Touw Ivo Paul , Niemeyer Charlotte Marie , Erlacher Miriam , De Pater Emma 
Published: December 2024

Introduction:

GATA2 is a crucial transcription factor for hematopoietic stem and progenitor cell (HSPC) development. Germline mutations in GATA2 lead to monocytopenia, neutropenia, and B and NK cell lymphopenia, as well as an increased risk of progressing to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Despite the high risk for myeloid neoplasia in early life, the underlying mechanisms remain unclear. In a bed to benchside approach, we characterized two mouse models and a cell line reflecting a broad spectrum of GATA2 haploinsufficiency-related phenotypes and investigated the molecular mechanisms involved.

Methods:

We examined the hematopoietic systems of GATA2 haploinsufficient mice (Gata2+/- and vav-Cre;Gata2fl/+ models) during aging and in functional assays. We performed serial transplantations with BM cells and low-dose (2x104) sorted LSK cells into lethally irradiated recipients. Mice were monitored for up to one year and tissue was analyzed by flow cytometry, histopathology, RNAseq, WES and array CGH. Additionally, we tested K562 cell lines with one GATA2 allele for genomic instability.

Results:

Gata2+/- HSPC analysis showed reduced numbers of phenotypic HSCs in embryonic fetal liver, adult, and aged mouse BM compared to WT mice, but no other hematological abnormalities could be found. In contrast, adult vav-Cre;Gata2fl/+ mice had normal HSC numbers initially, but at 15 months, HSC numbers significantly decreased, accompanied by mild cytopenia. Beside that, the BM composition of 60-week-old vav-Cre;Gata2fl/+ mice was similar to that of Gata2fl/+ controls with only mild dysplasia. In order to trigger an MDS/AML like phenotype, we applied cellular stress to Gata2 haploinsufficient HSPCs by performing serial transplantations of aged Gata2+/- BM cells into lethally irradiated mice. These transplantations resulted in monocytopenia and B lymphopenia, with a block in the transitional B cell compartment linked to pro-B cell senescence. We further increased the pressure by transplanting low numbers of vav-Cre;Gata2fl/+ or Gata2fl/+ sorted LSK cells. Under this condition, ten out of 28 recipients (35%) of vav-Cre;Gata2fl/+ mice died between 83 and 200 days (mean 142 days) post-transplantation. Analysis of ten deceased vav-Cre;Gata2fl/+ recipients revealed an aplastic BM in six of them, and T cell leukemia with thymic and splenic involvement in three animals. Importantly, all three mice had few leukemic cells in the otherwise empty BM. In one animal, the BM was normocellular with dysplastic megakaryo- and erythropoiesis, and left-shifted myelopoiesis, being B and T cells increased in numbers. Whether this MDS-like histopathological picture was preceded by BM failure is unclear.

In order to identify the molecular events behind leukemogenesis induced by Gata2 haploinsufficiency, we performed WES and array-CGH. We identified mutations in oncogenes and tumor suppressors, as well as chromosomal aberrations, including trisomy 15 (analogous to human trisomy 8), Pten deletions, Notch1 and Uba1 mutations. In three of the mice that developed BM failure without progression to a leukemic stage, no abnormalities were detected in the BM. Recipients of vav-Cre;Gata2fl/+ cells reconstituted their hematopoietic system before developing BMF, although reconstitution was delayed. Transcriptional profiles of Gata2+/- HSCs showed deregulated proliferation and DNA damage responses throughout development. K562 cells with GATA2 haploinsufficiency exhibited increased cytokinesis defects compared to WT cells, suggesting a direct effect on chromosomal segregation and a potential mechanism for leukemic transformation in Gata2 deficiency syndrome.

Discussion:

The aim of this study was to identify the cellular and molecular events leading to MDS/AML in the GATA2 syndrome. Using several in vitro and in vivo models, we could show that leukemia occurs secondary to BM failure in a GATA haploinsufficient system. Our data further shows that dysregulation of both proliferation and DNA damage response in addition to chromosomal instability and aging are the major drivers of malignant transformation.