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Leukemogenic potency of the novel FLT3-N676K mutant

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Abstract

The novel FMS-like tyrosine kinase 3 (FLT3)-N676K point mutation within the FLT3 kinase domain-1 was recently identified in 6 % of de novo acute myeloid leukemia (AML) patients with inv(16). Because FLT3-N676K was encountered almost exclusively in inv(16) AML, we investigated the transforming potential of FLT3-N676K, the cooperation between FLT3-N676K and core binding factor ß-smooth muscle myosin heavy chain (CBFß-SMMHC) (encoded by the inv(16) chimeric gene CBFB-MYH11) in inducing acute leukemia, and tested the sensitivity of FLT3-N676K-positive leukemic cells to FLT3 inhibitors. Retroviral expression of FLT3-N676K in myeloid 32D cells induced AML in syngeneic C3H/HeJ mice (n = 11/13, median latency 58 days), with a transforming activity similar to FLT3-internal tandem duplication (ITD) (n = 8/8), FLT3-TKD D835Y (n = 8/9), and FLT3-ITD-N676K (n = 9/9) mutations. Three out of 14 (21.4 %) C57BL/6J mice transplanted with FLT3-N676K-transduced primary hematopoietic progenitor cells developed acute leukemia (latency of 68, 77, and 273 days), while no hematological malignancy was observed in the control groups including FLT3-ITD. Moreover, co-expression of FLT3-N676K/CBFß-SMMHC did not promote acute leukemia in three independent experiments (n = 16). In comparison with FLT3-ITD, FLT3-N676K induced much higher activation of FLT3 and tended to trigger stronger phosphorylation of MAPK and AKT. Importantly, leukemic cells carrying the FLT3-N676K mutant in the absence of an ITD mutation were highly sensitive to FLT3 inhibitors AC220 and crenolanib, and crenolanib even retained activity against the AC220-resistant FLT3-ITD-N676K mutant. Taken together, the FLT3-N676K mutant is potent to transform murine hematopoietic stem/progenitor cells in vivo. This is the first report of acute leukemia induced by an activating FLT3 mutation in C57BL/6J mice. Moreover, further experiments investigating molecular mechanisms for leukemogenesis induced by FLT3-N676K mutation and clinical evaluation of FLT3 inhibitors in FLT3-N676K-positive AML seem warranted.

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Acknowledgments

This work was supported by the Deutsche José Carreras Leukämie-Stiftung (grant: 13/22) and the Deutsche Forschungsgemeinschaft (grants: Li 1608/2-1, FI 405/5-1 to FH and TF, SCHE 550/6-1, ED 34/4-1, H.W. & J. Hector-Stiftung to MS and ME). KH and ZP were supported by the China Scholarship Council (2011638024 and 201406100008). GB was supported by the Cluster of Excellence REBIRTH. We thank Axel Schambach (Hannover Medical School, Hannover, Germany) and Christopher A. Klug (Birmingham) for providing pSRS11 SF iGFP pre and human CBFB-MYH11 cDNA, respectively, and Mwe Mwe Chao for the critical reading of this paper and Rena-Mareike Struß for the technical assistance. We are very grateful to Cell Sorting Core Unit, Iris Dallmann, and Karin Battmer (all Hannover Medical School) for their support.

Author’s contribution

KH performed the research; collected, analyzed, and interpreted the data; and wrote the manuscript. MY and ZP performed the research and interpreted the data. FHH, TF, MS, GB, KW, NvN, and AG performed the research, interpreted the data, contributed the reagents, and provided the administrative support. ZL conceived the concept, designed the experiments, analyzed and interpreted the data, and wrote the manuscript. All authors agreed on the final version of the manuscript.

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Authors and Affiliations

  1. Department of Hematology, Hemostasis, Oncology, and Stem Cell Transplantation, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany

    Kezhi Huang, Min Yang, Zengkai Pan, Michaela Scherr, Matthias Eder, Arnold Ganser & Zhixiong Li

  2. Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany

    Kezhi Huang, Min Yang & Zhixiong Li

  3. Department of Hematology and Oncology, Medical Center, Otto-von-Guericke University Magdeburg, Magdeburg, Germany

    Florian H. Heidel & Thomas Fischer

  4. Internal Medicine II, Hematology and Oncology, University Hospital Jena, Jena, Germany

    Florian H. Heidel

  5. Institute of Pathology, Hannover Medical School, Hannover, Germany

    Guntram Büsche

  6. Department of Molecular Hematopoiesis, Hannover Medical School, Hannover, Germany

    Karl Welte

  7. Institute of Human Genetics, Hannover Medical School, Hannover, Germany

    Nils von Neuhoff

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  1. Kezhi Huang
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Huang, K., Yang, M., Pan, Z. et al. Leukemogenic potency of the novel FLT3-N676K mutant. Ann Hematol 95, 783–791 (2016). https://doi.org/10.1007/s00277-016-2616-z

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