Abstract
Acute myeloid leukemia (AML) with mutations in the FMS-like tyrosine kinase (FLT3) is a clinically unresolved problem. AML cells frequently have a dysregulated expression and activity of epigenetic modulators of the histone deacetylase (HDAC) family. Therefore, we tested whether a combined inhibition of mutant FLT3 and class I HDACs is effective against AML cells. Low nanomolar doses of the FLT3 inhibitor (FLT3i) AC220 and an inhibition of class I HDACs with nanomolar concentrations of FK228 or micromolar doses of the HDAC3 specific agent RGFP966 synergistically induce apoptosis of AML cells that carry hyperactive FLT3 with an internal tandem duplication (FLT3-ITD). This does not occur in leukemic cells with wild-type FLT3 and without FLT3, suggesting a preferential toxicity of this combination against cells with mutant FLT3. Moreover, nanomolar doses of the new FLT3i marbotinib combine favorably with FK228 against leukemic cells with FLT3-ITD. The combinatorial treatments potentiated their suppressive effects on the tyrosine phosphorylation and stability of FLT3-ITD and its downstream signaling to the kinases ERK1/ERK2 and the inducible transcription factor STAT5. The beneficial pro-apoptotic effects of FLT3i and HDACi against leukemic cells with mutant FLT3 are associated with dose- and drug-dependent alterations of cell cycle distribution and DNA damage. This is linked to a modulation of the tumor-suppressive transcription factor p53 and its target cyclin-dependent kinase inhibitor p21. While HDACi induce p21, AC220 suppresses the expression of p53 and p21. Furthermore, we show that both FLT3-ITD and class I HDAC activity promote the expression of the checkpoint kinases CHK1 and WEE1, thymidylate synthase, and the DNA repair protein RAD51 in leukemic cells. A genetic depletion of HDAC3 attenuates the expression of such proteins. Thus, class I HDACs and hyperactive FLT3 appear to be valid targets in AML cells with mutant FLT3.
Highlights
The membrane-bound FMS-like tyrosine kinase-3 (FLT3) is activated by the FLT3 ligand and contributes to the development of hematopoietic cells
We show that both FLT3-ITD and class I histone deacetylase (HDAC) activity promote the expression of the checkpoint kinases CHK1 and WEE1, thymidylate synthase, and the DNA repair protein RAD51 in leukemic cells
Cells that were treated with synergistically active combinations of AC220 plus FK228, accumulated higher levels of cleaved caspase-3 than cells that were treated with the single drugs (Fig. 1C)
Summary
The membrane-bound FMS-like tyrosine kinase-3 (FLT3) is activated by the FLT3 ligand and contributes to the development of hematopoietic cells. Extended author information available on the last page of the article juxtamembrane domain, and an intracellular tyrosine kinase domain (Majothi et al 2020; Marensi et al 2021; Scholl et al 2020). Internal tandem duplications in FLT3 (FLT3-ITD) occur in 20–35% of patients with de novo AML and tyrosine kinase domain (FLT3-TKD) mutations occur in 5–10% of patients with de novo AML (Heidel et al 2006; Majothi et al 2020; Marensi et al 2021; Scholl et al 2020)
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