Abstract
Internal tandem duplications (ITD) of the FMS-like tyrosine kinase 3 (FLT3) predict poor prognosis in acute myeloid leukemia (AML) and often co-exist with inactivating DNMT3A mutations. In vitro studies implicated Grb2-associated binder 2 (GAB2) as FLT3-ITD effector. Utilizing a Flt3-ITD knock-in, Dnmt3a haploinsufficient mouse model, we demonstrate that Gab2 is essential for the development of Flt3-ITD driven AML in vivo, as Gab2 deficient mice displayed prolonged survival, presented with attenuated liver and spleen pathology and reduced blast counts. Furthermore, leukemic bone marrow from Gab2 deficient mice exhibited reduced colony-forming unit capacity and increased FLT3 inhibitor sensitivity. Using transcriptomics, we identify the genes encoding for Axl and the Ret co-receptor Gfra2 as targets of the Flt3-ITD/Gab2/Stat5 axis. We propose a pathomechanism in which Gab2 increases signaling of these receptors by inducing their expression and by serving as downstream effector. Thereby, Gab2 promotes AML aggressiveness and drug resistance as it incorporates these receptor tyrosine kinases into the Flt3-ITD signaling network. Consequently, our data identify GAB2 as a promising biomarker and therapeutic target in human AML.
Highlights
25% of all acute myeloid leukemia (AML) carry an internal tandem duplication (ITD) in the juxtamembrane domain of the receptor tyrosine kinase (RTK) FMS-like tyrosine kinase 3 (FLT3), leading to constitutive PI3K/AKT, ERK and STAT5 signaling [1,2,3]
We crossed Flt3-ITD knock-in mice [17] with animals harboring floxed DNA methyltransferase 3a (Dnmt3a) (Dnmt3af/f) alleles [21] and transgenic Mx-Cre mice [22] to obtain Dnmt3af/+/Flt3ITD/ITD as well as Dnmt3af/+/Flt3ITD/ITD/Mx-Cre mice (Fig. 1A; crossing scheme in Supplementary Fig. 1A). The latter lose their floxed Dnmt3a allele upon polyinosinicpolycytidylic acid induced Cre expression generating Flt3ITD/ITD/Dnmt3a−/+ animals, which will from hereon be referred to as AML mice, while Dnmt3af/+/Flt3ITD/ITD mice that do not express Cre and retain their floxed Dnmt3a allele will be designated as myeloproliferative neoplasms (MPNs) mice
This genetically engineered mouse model (GEMM) closely recapitulates the human disease, as FLT3-ITD and DNMT3A mutations frequently co-exist in cytogenetically normal AML and forecast dismal prognosis [34]
Summary
25% of all AML carry an internal tandem duplication (ITD) in the juxtamembrane domain of the receptor tyrosine kinase (RTK) FLT3, leading to constitutive PI3K/AKT, ERK and STAT5 signaling [1,2,3]. The docking protein GAB2 amplifies signals downstream of cytokine and growth factor receptors, including FLT3-ITD [4]. GAB2 is recruited to activated receptors via the adaptor GRB2 and is subsequently phosphorylated at multiple tyrosine residues [5]. These modifications serve as binding sites for the tyrosine phosphatase SHP2 and the p85 subunit of PI3K, resulting in the activation of ERK and AKT/mTOR axes, respectively [5,6,7]. Given its contribution to three oncogenic pathways, it is anticipated that GAB2 emerges as an important player in solid tumors and leukemia [9]
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