Inhibitors of class I HDACs and of FLT3 combine synergistically against leukemia cells with mutant FLT3

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.

Novel hydroxamic acid derivative induces apoptosis and constrains autophagy in leukemic cells

“…HDAC3 has key survival functions in leukemic cells of various hematopoietic lineages. These include the control of cell proliferation, apoptosis, DNA replication fork stability, and genomic integrity [31,[49][50][51][52][53][54][55]. Intriguingly, a full-body knockout of HDAC3 was not lethal in mice [56].…”

“…The stronger sensitivity of such cells is consistent with the literature and multiple explanations for this have been provided for leukemic cells. These include, but are not limited to, an inhibition of the S phase-dependently induced transcription factor NF-κB [59,61,62], the induction of DNA replication stress/DNA damage [31,[49][50][51]53], inactivation of signaling through the transcription factor STAT5 [63], and a disruption of proper mitosis [64]. KH16-induced cell cycle disruptions are linked to a significant fragmentation of DNA.…”

“…We recorded cell cycle profiles by flow cytometry [30][31][32]. PI is a marker for DNA contents in different phases of the cell cycle.…”

“…/propidium iodide (PI) staining This assay was previously described by us[30][31][32]. Annexin-V binds to phosphatidylserine, which becomes exposed on the outer membrane of early and late apoptotic cells.…”

Targeting the MYC interaction network in B-cell lymphoma via histone deacetylase 6 inhibition

“…The role of HDAC6 inhibitors in cancer therapy is still a matter of debate. Recent studies showed that many cancer models are tolerant to HDAC6 inhibitor treatment (43)(44)(45).…”