Activating mutations of FLT3 are among the most frequent alterations in AML. Two distinct types of mutations have been described: Duplication of the juxtamembraneous domain (ITD) and point mutations of the tyrosine kinase domain (TKD). Although both lead to constitutive FLT3 signaling, FLT3-ITD signals through the STAT5 pathway whereas FLT3-TKD and FLT3 WT activate STAT5 to a much lesser degree. In a murine bone marrow transplantation model, FLT3-ITD leads to a myeloproliferative neoplasm; FLT3-TKD induces a lymphoid disease with significantly longer latency. Interaction of SRC with FLT3-ITD was recently shown to be necessary for STAT5 activation.
Fetal liver cells from constitutive STAT5 knockout mice were infected with FLT3 oncogenes and cultured on methylcellulose. Conditional STAT5 knockout mice were analyzed for phenotype and latency of hematologic disorders after transplantation with FLT3-ITD or FLT3-TKD-infected bone marrow. Expression of STAT5 target genes was determined using Realtime-PCR. Phospho-STAT5 levels were quantified by intracellular staining and flow cytometry.
Depletion of STAT5 leads to a dramatic decrease in the ability of fetal liver cells to form colonies in Methylcellulose when infected with FLT3-ITD, but not when infected with FLT3-TKD. In-vivo studies using a murine conditional knockout model proved that STAT5 is necessary for the induction of a myeloproliferative disease by FLT3-ITD, while FLT3-TKD is not dependent on STAT5. Mice with deletion of the STAT5 allele displayed expansion of mainly lymphoid cells with an increased latency similar to the FLT3-TKD disease. In additional studies differential activation of STAT5 in myeloid and lymphoid progenitors was found. The STAT5 target gene Oncostatin M is exclusively expressed in myeloid progenitor cells infected with FLT3-ITD and in mice with intact STAT5 that were transplanted with FLT3-ITD. Furthermore, elevated SRC expression levels could be detected in proliferating myeloid progenitor cells compared to lymphoid progenitors.
STAT5 is critical for the induction of a myeloproliferative disease by FLT3-ITD in a murine transplantation model. Interaction of SRC and FLT3-ITD leads to the activation of STAT5 in myeloid progenitors, but not in lymphoid progenitors expressing low levels of SRC. Oncostatin M upregulation is likely to be a downstream process contributing to the phenotype and latency of the disease.
No relevant conflicts of interest to declare.
Asterisk with author names denotes non-ASH members.