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Rare FLT3 deletion mutants may provide additional treatment options to patients with AML: an approach to individualized medicine

Leukemia volume 29, pages 2434–2438 (2015)Cite this article

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The receptor tyrosine kinase (RTK) FLT3 (Fms-like tyrosine kinase-3) is essential for the proliferation, differentiation and survival of hematopoietic cells.1, 2 Mutations in the gene for FLT3 have been described in acute myeloid leukemia (AML) as internal tandem duplications (ITD) localized in the juxtamembrane (JM) domain and FLT3 tyrosine kinase domain (TKD) mutations amounting to 2–30% and 7–10% of patients, respectively. In patients with AML, FLT3-ITD confers an unfavorable prognosis.3 In AML, recently discovered FLT3 mutations involve point mutations as well as short deletions in the JM domain, highlighting the importance for a deeper understanding and further studies of these aberrations.4, 5 The FLT3 JM domain (Supplementary Figure 1a) is indeed the critical regulator region of receptor autoinhibition.6 Kiyoi et al.7 had shown constitutive activation of the FLT3 receptor by deletion of a stretch of amino acids in the JM domain, generally duplicated in FLT3-ITD.

Transient transfection into HEK293T cells confirmed activation of associated signaling proteins by several deletion mutants. Even deletion of just one amino acid led to the activation of downstream signaling (Supplementary Figures 1b and c). When retrovirally expressed in the IL3-dependent pro-B cell line Ba/F3, the FLT3 Stop and delLeu (Supplementary Table 1, Id 3 and Id 2, respectively) mutants were not further studied in this work, as we focused on activating mutations in the JM domain. The delEY and the delIns mutants led to constitutive activation of STAT3, ERK1/2, SFK (Src family kinases), SHP2 and AKT (Figure 1a). Similar to FLT3-ITD, they also activated STAT5. As seen in HEK293T cells, only FLT3-ITD induced upregulated phosphorylation of Y589/Y591. These tyrosines were previously thought to be essential for STAT5 activation and transforming capacity of the RTK.9 Our data show that STAT5 was partly activated by SFKs in deletion mutant expressing cells (Figure 1b), whereas JAK2 inhibition had no effect on STAT5 phosphorylation (Supplementary Figure 2a). A similarity between FLT3 deletion mutants and the TKD mutant FLT3 D835Y (p.Asp835Tyr) is conceivable, as both do not show an increase in Y589/Y591 phosphorylation.9 Nevertheless, the missing phosphorylation of Y589/591 was associated with low STAT5 activation in D835Y expressing cells but not for FLT3 deletion mutants (Figure 1a). Immunoprecipitation experiments confirmed the phosphorylation of additional tyrosines independent from Y598/Y591 in the deletion mutants, as was already shown for FLT3-ITD (Supplementary Figure 2b).9 FLT3 ligand stimulation of the WT receptor led to the phosphorylation of ERK1/2 and AKT but not of STAT5. Interestingly, STAT3 was not activated by FLT3-ITD, suggesting that the deletion mutants and ITD induced different patterns of downstream signaling. These results were in line with those of Frohling et al.10 of STAT3 phosphorylation driven by FLT3 JM point mutations. We found that endoplasmic reticulum targeting or hypoglycosylation of FLT3 delEY mutant by brefeldin A and tunicamycin treatment, respectively, clearly diminished STAT5 and STAT3 phosphorylation, whereas there was no change with FLT3-ITD (Supplementary Figure 2c), as published.11 The same was seen for the delIns mutant only with tunicamycin treatment. All mutants exhibited decreased phosphorylation of AKT after brefeldin A and tunicamycin treatment, confirming receptor surface expression as a prerequisite for PI3K/AKT pathway activation.

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References

  1. McKenna HJ, Stocking KL, Miller RE, Brasel K, De Smedt T, Maraskovsky E et al. Mice lacking flt3 ligand have deficient hematopoiesis affecting hematopoietic progenitor cells, dendritic cells, and natural killer cells. Blood 2000; 95: 3489–3497.

    CAS  PubMed  Google Scholar 

  2. Mackarehtschian K, Hardin JD, Moore KA, Boast S, Goff SP, Lemischka IR . Targeted disruption of the flk2/flt3 gene leads to deficiencies in primitive hematopoietic progenitors. Immunity 1995; 3: 147–161.

    Article  CAS  Google Scholar 

  3. Kayser S, Schlenk RF, Londono MC, Breitenbuecher F, Wittke K, Du J et al. Insertion of FLT3 internal tandem duplication in the tyrosine kinase domain-1 is associated with resistance to chemotherapy and inferior outcome. Blood 2009; 114: 2386–2392.

    Article  CAS  Google Scholar 

  4. Reindl C, Bagrintseva K, Vempati S, Schnittger S, Ellwart JW, Wenig K et al. Point mutations in the juxtamembrane domain of FLT3 define a new class of activating mutations in AML. Blood 2006; 107: 3700–3707.

    Article  CAS  Google Scholar 

  5. Deeb KK, Smonskey MT, DeFedericis H, Deeb G, Sait SNJ, Wetzler M et al. Deletion and deletion/insertion mutations in the juxtamembrane domain of the FLT3 gene in adult acute myeloid leukemia. Leuk Res Rep 2014; 3: 86–89.

    PubMed  PubMed Central  Google Scholar 

  6. Griffith J, Black J, Faerman C, Swenson L, Wynn M, Lu F et al. The structural basis for autoinhibition of FLT3 by the juxtamembrane domain. Mol Cell 2004; 13: 169–178.

    Article  CAS  Google Scholar 

  7. Kiyoi H, Ohno R, Ueda R, Saito H, Naoe T . Mechanism of constitutive activation of FLT3 with internal tandem duplication in the juxtamembrane domain. Oncogene 2002; 21: 2555–2563.

    Article  CAS  Google Scholar 

  8. Li L, Bailey E, Greenblatt S, Huso D, Small D . Loss of the wild-type allele contributes to myeloid expansion and disease aggressiveness in FLT3/ITD knockin mice. Blood 2011; 118: 4935–4945.

    Article  CAS  Google Scholar 

  9. Rocnik JL, Okabe R, Yu J, Lee BH, Giese N, Schenkein DP et al. Roles of tyrosine 589 and 591 in STAT5 activation and transformation mediated by FLT3-ITD. Blood 2006; 108: 1339–1345.

    Article  CAS  Google Scholar 

  10. Fröhling S, Scholl C, Levine RL, Loriaux M, Boggon TJ, Bernard OA et al. Identification of driver and passenger mutations of FLT3 by high-throughput DNA sequence analysis and functional assessment of candidate alleles. Cancer Cell 2007; 12: 501–513.

    Article  Google Scholar 

  11. Choudhary C, Olsen JV, Brandts C, Cox J, Reddy PNG, Böhmer FD et al. Mislocalized activation of oncogenic RTKs switches downstream signaling outcomes. Mol Cell 2009; 36: 326–339.

    Article  CAS  Google Scholar 

  12. Rahmani M, Aust MM, Attkisson E, Williams DC, Ferreira-Gonzalez A, Grant S . Dual inhibition of Bcl-2 and Bcl-xL strikingly enhances PI3K inhibition-induced apoptosis in human myeloid leukemia cells through a GSK3- and Bim-dependent mechanism. Cancer Res 2013; 73: 1340–1351.

    Article  CAS  Google Scholar 

  13. Armstrong SA, Mabon ME, Silverman LB, Li A, Gribben JG, Fox EA . FLT3 mutations in childhood acute lymphoblastic leukemia. Blood 2004; 103: 3544–3546.

    Article  CAS  Google Scholar 

  14. Chang P, Kang M, Xiao A, Chang J, Feusner J, Buffler P et al. FLT3 mutation incidence and timing of origin in a population case series of pediatric leukemia. BMC Cancer 2010; 10: 513.

    Article  Google Scholar 

  15. Kim TW, Lee H, Kang Y, Choe MS, Ryu M, Chang HM et al. Prognostic significance of c-kit mutation in localized gastrointestinal stromal tumors. Clin Cancer Res 2004; 10: 3076–3081.

    Article  CAS  Google Scholar 

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Acknowledgements

We thank Kristina Feldberg for expert technical assistance. We acknowledge Frank D Böhmer for providing the pcDNA3-FLT3 (WT)-HA vector and M-D Hagel for initial cloning of vectors. This work was in part supported by the Novartis Foundation.

Author contribution

NC, RCP, GR, JR, PC, MS, TH, THB, SS and SK designed research. NC, GR, RCP and JR performed research. NC, GR, RCP, PC, TH, SS and SK analyzed data. NC, RCP, GR, SS and SK wrote the paper.

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Authors and Affiliations

  1. Department of Hematology, Oncology, Hemostaseology and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany

    N Chatain, J Rossa, M Schemionek, T H Brümmendorf & S Koschmieder

  2. Computational Biophysics, German Research School for Simulation Sciences and Computational Biomedicine, Institute for Advanced Simulation IAS-5 and Institute of Neuroscience and Medicine INM-9, Forschungszentrum Jülich, Jülich, Germany

    R C Perera, G Rossetti & P Carloni

  3. Jülich Supercomputing Centre, Forschungszentrum Jülich, Jülich, Germany

    G Rossetti

  4. MLL Munich Leukemia Laboratory, Munich, Germany

    T Haferlach & S Schnittger

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  1. N Chatain
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Corresponding author

Correspondence to S Koschmieder.

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Competing interests

SK has received honoraria for consultancy from Novartis, Pfizer, Brystol-Meyers Squibb, and Ariad. TH and SS have equity ownership of MLL. THB has received honoraria for consultancy from Novartis, Pfizer, Brystol-Meyers Squibb. The remaining authors declare no conflict of interest.

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Chatain, N., Perera, R., Rossetti, G. et al. Rare FLT3 deletion mutants may provide additional treatment options to patients with AML: an approach to individualized medicine. Leukemia 29, 2434–2438 (2015). https://doi.org/10.1038/leu.2015.131

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