Transformation by Oncogenic Mutants and Ligand-Dependent Activation of FLT3 Wild-type Requires the Tyrosine Residues 589 and 591

Vempati, Sridhar; Reindl, Carola; Wolf, Ulla; Kern, Ruth; Petropoulos, Konstantin; Naidu, Vegi M.; Buske, Christian; Hiddemann, Wolfgang; Kohl, Tobias; Spiekermann, Karsten

doi:10.1158/1078-0432.ccr-07-1873

Cited by 23 publications

(24 citation statements)

References 46 publications

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“…This results in only a fraction of phosphorylated JM Flt3 AL mutant relative to Flt3 ITD to undergo the same processing delay as Flt3 ITD. This is supported by the report of Tyr589/591 also being required for ligand-independent proliferation of cells expressing Flt3 AL (Vempati et al, 2008). The non-phosphorylated JM Flt3 AL fraction is efficiently processed and traffics to the plasma membrane, and would transmit downstream signals similarly to those of Flt3 WT.…”

Section: A Working Model Of Differential Signalingsupporting

confidence: 52%

Differential signaling of Flt3 activating mutations in acute myeloid leukemia: a working model

Chan

2011

Protein Cell

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Receptor tyrosine kinases couple a wide variety of extracellular cues to cellular responses. The class III subfamily comprises the platelet-derived growth factor receptor, c-Kit, Flt3 and c-Fms, all of which relay cell proliferation signals upon ligand binding. Accordingly, mutations in these proteins that confer ligand-independent activation are found in a subset of cancers. These mutations cluster in the juxtamembrane (JM) and catalytic tyrosine kinase domain (TKD) regions. In the case of acute myeloid leukemia (AML), the juxtamembrane (named ITD for internal tandem duplication) and TKD Flt3 mutants differ in their spectra of clinical outcomes. Although the mechanism of aberrant activation has been largely elucidated by biochemical and structural analyses of mutant kinases, the differences in disease presentation cannot be attributed to a change in substrate specificity or signaling strength of the catalytic domain. This review discusses the latest literature and presents a working model of differential Flt3 signaling based on mis-localized juxtamembrane autophosphorylation, to account for the disease variation. This will have bearing on therapeutic approaches in a complex disease such as AML, for which no efficacious drug yet exists.

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Section: A Working Model Of Differential Signalingsupporting

confidence: 52%

Differential signaling of Flt3 activating mutations in acute myeloid leukemia: a working model

Chan

2011

Protein Cell

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“…Y589 and Y591 have been identified as Src association sites [26] and involved in the activation of STAT5 [34]. Moreover, these two residues were shown to be crucial for the transforming potential of FLT3-ITD and D835Y as well as for the ligand-dependent activation of wild-type FLT3 [42]. Y599 was demonstrated to be involved in binding of the protein tyrosine phosphatase SHP2 [26].…”

Section: Discussionmentioning

confidence: 99%

Oncogenic Flt3 receptors display different specificity and kinetics of autophosphorylation

Razumovskaya¹,

Masson²,

Khan

et al. 2009

Experimental Hematology

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Objective. Fms-like tyrosine kinase-3 (FLT3), a growth factor receptor normally expressed in hematopoietic progenitor cells, has been shown to have an important role in the development of acute myeloid leukemia c due to activating mutations. FLT3 mutations are found in approximately one third of AML patients and correlate with a poor prognosis, thus making the FLT3 receptor a potential therapeutic target. The aim of the investigation is to analyze the kinetics and specificity of FLT3 autophosphorylation in wild-type FLT3 as well as in the oncogenic FLT3 mutants.Methods. We have used Ba/F3 cells stably expressing either wild-type, ITD or D835Y mutants of FLT3 in order to compare the site selectivity of tyrosine phosphorylation sites. By the use of a panel of phosphospecific antibodies directed against potential tyrosine phosphorylation sites in FLT3, we identified several novel phosphorylation sites in FLT3 and studied the kinetics and specificity of ligand-induced phosphorylation in living cells.Results. Eight phosphorylated tyrosines (pY589, pY591, pY599, pY726, pY768, pY793, pY842 and pY955) were investigated and shown to be differentially phosphorylated in the wild-type versus the mutated receptors. Furthermore, we show that tyrosines 726, 793 and 842 are novel phosphorylation sites of FLT3 in intact cells. Conclusion.In this study, we have looked at the site-specific phosphorylation in the wild-type FLT3 in comparison to the mutants found in AML. We observed not only quantitative changes but more importantly, qualitative differences in the phosphorylation patterns of the wild-type and the mutated FLT3 receptors, which might contribute to the understanding of the mechanisms by which FLT3 contributes to AML in patients with mutations in FLT3.

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“…This result agrees with the assumption that SRC binding to FLT3-ITD is essential for robust STAT5 activation and with the results of previous studies demonstrating that tyrosine residues 589/591 in FLT3-ITD are essential for strong STAT5 activation and for the induction of myeloproliferative disease in mice. 28,35,36 SRC SH2-mediated binding to tyrosine 589/591 requires phosphorylation of these sites. To compare the level of phosphorylation at tyrosine 589/591 in FLT3-ITD, FLT3-TKD, and FLT3-WT after ligand stimulation, we used site-specific anti-pY Abs ( Figure 2C).…”

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confidence: 99%

“…36 Vempati et al showed that the tyrosines 589 and 591 are also important for growth of Ba/F3 cells expressing FLT3-WT stimulated with FL or FLT3-TKD. 28 Previous studies have compared the signaling of FLT3-ITD and FLT3-TKD and demonstrated that the localization of FLT3-ITD and FLT3-TKD significantly alters the transforming capability and activation of downstream signaling molecules. In the case of FLT3-ITD, retention in the endoplasmic reticulum was suggested One representative of at least 3 independent experiments is shown.…”

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confidence: 99%

“…In contrast, several other groups proposed the involvement of various signaling molecules, including LYN, in the process of STAT5 activation by FLT3-ITD in murine and human cells. 28,47,48 Up to now, there have been no investigations of the signaling requirements of FLT3-ITD compared with FLT3-TKD in leading to differential STAT5 activation.In the present study, we have shown that FLT3-ITD but not FLT3-TKD strongly binds in vitro to the SH2 domain of SRC. No binding of FLT3-ITD to the SH2 domains of other SFK family members such as LCK, HCK, or FYN could be detected.…”

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confidence: 99%

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SRC is a signaling mediator in FLT3-ITD– but not in FLT3-TKD–positive AML

Leischner

Albers

Grundler

et al. 2012

Blood

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Mutations of Fms-like tyrosine kinase 3 (FLT3) are among the most frequently detected molecular abnormalities in AML patients. Internal tandem duplications (ITDs) are found in approximately 25% and point mutations within the second tyrosine kinase domain (TKD) in approximately 7% of AML patients. Patients carrying the FLT3-ITD but not the FLT3-TKD mutation have a significantly worse prognosis. Therefore, both FLT3 mutations seem to exert different biologic functions. FLT3-ITD but not FLT3-TKD has beenshown to induce robust activation of the STAT5 signaling pathway. In the present study, we investigated the mechanisms leading to differential STAT5 activation and show that FLT3-ITD but not FLT3-TKD uses SRC to activate STAT5. Coimmunoprecipitation and pull-down experiments revealed an exclusive interaction between SRC but not other Src family kinases and FLT3-ITD, which is mediated by the SRC SH2 domain. We identified tyrosines 589 and 591 of FLT3-ITD to be essential for SRC binding and subsequent STAT5 activation. Using sitespecific Abs, we found that both residues were significantly more strongly phosphorylated in FLT3-ITD compared with FLT3-TKD. SRC inhibition and knockdown blocked STAT5 activation and proliferation induced by FLT3-ITD but not by FLT3-TKD. We conclude that SRC might be a therapeutic target in FLT3-ITD ؉ AML. IntroductionThe Fms-like tyrosine kinase-3 (FLT3) receptor tyrosine kinase is expressed on blast cells in most patients with acute myeloid leukemia (AML). 1 Activating mutations of FLT3 have been detected in approximately 30% of AML patients. 2,3 Two distinct groups of FLT3 mutations have been described: (1) internal tandem duplications (FLT3-ITDs), which are most common in the juxtamembrane or the beta1-sheet of the tyrosine kinase domain-1 coding sequence in approximately 20%-27% of patients with AML; and (2) point mutations within the second tyrosine kinase domain (FLT3-TKDs) in approximately 7% of AML patients. 2,4,5 Both types of mutations activate the FLT3 receptor constitutively, leading to activation of downstream signaling proteins and resulting in factor-independent proliferation of murine lymphoid and myeloid cells. [6][7][8][9][10] Studies have identified FLT3-ITD as a prognostic factor because patients harboring this type of mutation have elevated peripheral blood and BM blast counts and an increased chance of relapse and inferior overall survival. 2,11,12 For FLT3-TKD mutations, a clear association with an inferior outcome could not be demonstrated in all studies and the relevance of FLT3-TKD for clinical prognosis is being investigated further. 13 Because FLT3 mutations can be found in approximately 30% of all AML patients and FLT3-ITD is associated with an inferior clinical prognosis, FLT3 is considered as an attractive therapeutic target. 14 Therefore, several tyrosine kinase inhibitors (TKIs) targeting FLT3 have been developed and tested in AML patients. 15,16 These studies showed that a single application of the inhibitor led to a short-term clinical response in some pat...

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Transformation by Oncogenic Mutants and Ligand-Dependent Activation of FLT3 Wild-type Requires the Tyrosine Residues 589 and 591

Cited by 23 publications

References 46 publications

Differential signaling of Flt3 activating mutations in acute myeloid leukemia: a working model

Differential signaling of Flt3 activating mutations in acute myeloid leukemia: a working model

Oncogenic Flt3 receptors display different specificity and kinetics of autophosphorylation

SRC is a signaling mediator in FLT3-ITD– but not in FLT3-TKD–positive AML

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