Combination of midostaurin and ATRA exerts dose-dependent dual effects on acute myeloid leukemia cells with wild type FLT3

Abstract

Background Midostaurin combined with chemotherapy is currently used to treat newly diagnosed acute myeloid leukemia (AML) patients with FMS-like tyrosine kinase 3 (FLT3)-mutations. However, midostaurin acts as an antagonist to some chemotherapeutic agents in leukemia cell lines without FLT3 mutations. All-trans retinoic acid (ATRA) induces apoptosis when used in combination with midostaurin in FLT3-mutated AML cells. This combination has been shown to be safe in AML patients. However, the effect of this combination has not been investigated in AML without FLT3 mutations. Methods Cell proliferation was assessed by a cell counting assay. Cell death was evaluated by cell viability and Annexin-V assays. Cell differentiation was assessed by CD11b expression profiling and morphological analysis. To explore the underlying mechanisms, we studied the role of caspase3/7, Lyn, Fgr, Hck, RAF, MEK, ERK, AKT, PU.1, CCAAT/enhancer binding protein β (C/EBPβ) and C/EBPε by Western blot analysis and immunoprecipitation assays. Antitumor activity was also confirmed in mouse xenograft models established with AML cells. Results In this study, 0.1 − 0.25 μM midostaurin (mido(L)) combined with ATRA induced differentiation while 0.25 − 0.5 μM midostaurin (mido(H)) combined with ATRA triggered apoptosis in some AML cell lines without FLT3-mutations. Midostaurin combined with ATRA (mido-ATRA) also exhibited antitumor activity in mouse xenograft models established with AML cells. Mechanistically, mido(H)-ATRA-induced apoptosis was dependent on caspase-3/7. Mido(L)-ATRA inhibited Akt activation which was associated with decreased activity of Lyn/Fgr/Hck, resulted in dephosphorylation of RAF S259, activated RAF/MEK/ERK, along with upregulating the protein levels of C/EBPβ, C/EBPε and PU.1. A MEK specific inhibitor was observed to suppress mido(L)-ATRA-induced increases in the protein levels of C/EBPs and PU.1 and mido(L)-ATRA-induced differentiation. Furthermore, inhibition of Akt activity promoted mido(L)-ATRA-induced downregulation of RAF S259 phosphorylation and mido(L)-ATRA-induced differentiation. Therefore, Lyn/Fgr/Hck-associated Akt inhibition activated RAF/MEK/ERK and controlled mido(L)-ATRA-induced differentiation by upregulation of C/EBPs and PU.1. Mido(L)-ATRA also promoted assembly of the signalosome, which may facilitate RAF activation. Conclusions Midostaurin combined with ATRA exerts antitumor activity against AML with wild-type FLT3 mutations in vitro and in vivo. These findings may provide novel therapeutic strategies for some AML patients without FLT3 mutations and imply a new target of midostaurin.

In vivo imaging system (IVIS) therapeutic assessment of tyrosine kinase inhibitor-loaded gold nanocarriers for acute myeloid leukemia: a pilot study

“…The daily intraperitoneal dose for 14 consecutive days was chosen to ensure consistent exposure to the therapy, which is important to determine its therapeutic effect. This strategy was informed by existing literature data, suggesting that such a regimen could minimize toxicity while maximizing tumor inhibition (Lu et al, 2022). Moreover, we hypothesized that nanoparticles might exhibit delayed effects on tumor progression or interact differently with the tumor microenvironment over an extended period.…”

All-trans retinoic acid in hematologic disorders: not just acute promyelocytic leukemia

“…Low dose midostaurin (0.25–0.5 μM) + ATRA induced cell differentiation, whereas high dose midostaurin (0.25–0.5 μM) + ATRA led to apoptosis in FMS-like tyrosine kinase-3 (FLT3) AML cell line, and apoptosis induced by high dose midostaurin + ATRA was dependent on caspase-3/7. Low dose midostaurin + ATRA inhibited the activation of Akt, leading to dephosphorylation of RAF S259, activation of RAF/MEK/ERK, and upregulation of the protein levels of C/EBPβ, C/EBPε, and PU.1, which resulted in an increase in cell differentiation ( Lu et al, 2022 ). Nucleophosmin-1 (NPM1) mutations in AML not only dislocate NPM1 from the nucleolus but also disorganize the PML nucleosome, ATRA in combination with ATO synergistically induces proteasomal degradation of NPM1 mutations in AML cell lines or primary samples leading to differentiation and apoptosis, and downregulation of the NPM1 mutant by ATRA + ATO also enhances the therapeutic response of AML cells to doxorubicin ( El Hajj et al, 2015 ; Martelli et al, 2015 ).…”

Mitogen-Activated Protein Kinase and Nuclear Hormone Receptor Crosstalk in Cancer Immunotherapy

“…ATRA [161], together with tyrosine kinase inhibitor (midostaurin), triggered apoptosis in FLT3 wt or mutant acute myeloid leukemia cells and exerted anti-tumor efficacy in mouse xenograft models. Mechanistically, this drug combination activated caspase-3/7 and inhibited Lyn/Fgr/Hck-dependent AKT phosphorylation, resulting in RAF-MEK1/2-ERK1/2 activation and up-regulation of lineage-determining transcription factors (C/EBPβ, C/EBPε, PU.1.…”