Abstract
FMS-like tyrosine kinase 3 (FLT3) is a transmembrane protein expressed on normal hematopoietic stem and progenitor cells (HSC) and retained on malignant blasts in acute myeloid leukemia (AML). We engineered CD8+ and CD4+ T-cells expressing a FLT3-specific chimeric antigen receptor (CAR) and demonstrate they confer potent reactivity against AML cell lines and primary AML blasts that express either wild-type FLT3 or FLT3 with internal tandem duplication (FLT3-ITD). We also show that treatment with the FLT3-inhibitor crenolanib leads to increased surface expression of FLT3 specifically on FLT3-ITD+ AML cells and consecutively, enhanced recognition by FLT3-CAR T-cells in vitro and in vivo. As anticipated, we found that FLT3-CAR T-cells recognize normal HSCs in vitro and in vivo, and disrupt normal hematopoiesis in colony-formation assays, suggesting that adoptive therapy with FLT3-CAR T-cells will require subsequent CAR T-cell depletion and allogeneic HSC transplantation to reconstitute the hematopoietic system. Collectively, our data establish FLT3 as a novel CAR target in AML with particular relevance in high-risk FLT3-ITD+ AML. Further, our data provide the first proof-of-concept that CAR T-cell immunotherapy and small molecule inhibition can be used synergistically, as exemplified by our data showing superior antileukemia efficacy of FLT3-CAR T-cells in combination with crenolanib.
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Acknowledgements
We thank Silke Frenz and Elke Spirk for their expertise in performing the mouse experiments. H.J. was supported by a grant from the German Excellence Initiative awarded to the Graduate School of Life Sciences (GSLS), University of Würzburg. I.G.G. was supported by a grant from Fundación Alfonso Martin Escudero, Spain. M.H. is a member of the Young Scholar Program (Junges Kolleg) and Extraordinary Member of the Bavarian Academy of Sciences (Bayerische Akademie der Wissenschaften). The research was supported by the Deutsche Forschungsgemeinschaft via SFB/TR 221‚ modulation of graft-versus-host and graft-versus leukemia immune responses after allogeneic stem cell transplantation.This work was also supported by German Cancer Aid (Max Eder Program Award 110313 [M.H.].
Author Contributions
HJ designed and performed the experiments, analyzed the data and wrote the manuscript. IG-C, TN, ST and JR designed and performed the experiments, and analyzed the data. WH, JBM and JS analyzed the data. HB provided biologic material and analyzed the data. MH and HE designed experiments, analyzed the data, wrote the manuscript and supervised the project.
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MH and HJ are co-inventors on a patent related to the use of FLT3-CAR T-cells to treat AML filed by the University of Würzburg, Würzburg, Germany. MH is co-inventor on patents related to CAR-technologies filed by the Fred Hutchinson Cancer Research Center, Seattle, WA and the University of Würzburg, Würzburg, Germany. The remaining authors declare that they have no conflict of interest.
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Jetani, H., Garcia-Cadenas, I., Nerreter, T. et al. CAR T-cells targeting FLT3 have potent activity against FLT3−ITD+ AML and act synergistically with the FLT3-inhibitor crenolanib. Leukemia 32, 1168–1179 (2018). https://doi.org/10.1038/s41375-018-0009-0
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DOI: https://doi.org/10.1038/s41375-018-0009-0
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