FOXM1 contributes to treatment failure in acute myeloid leukemia
Irum Khan, … , Olga Frankfurt, Andrei L. Gartel
Irum Khan, … , Olga Frankfurt, Andrei L. Gartel
Published August 9, 2018
Citation Information: JCI Insight. 2018;3(15):e121583. https://doi.org/10.1172/jci.insight.121583.
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Research Article Hematology Oncology

FOXM1 contributes to treatment failure in acute myeloid leukemia

Abstract

Acute myeloid leukemia (AML) patients with NPM1 mutations demonstrate a superior response to standard chemotherapy treatment. Our previous work has shown that these favorable outcomes are linked to the cytoplasmic relocalization and inactivation of FOXM1 driven by mutated NPM1. Here, we went on to confirm the important role of FOXM1 in increased chemoresistance in AML. A multiinstitution retrospective study was conducted to link FOXM1 expression to clinical outcomes in AML. We establish nuclear FOXM1 as an independent clinical predictor of chemotherapeutic resistance in intermediate-risk AML in a multivariate analysis incorporating standard clinicopathologic risk factors. Using colony assays, we show a dramatic decrease in colony size and numbers in AML cell lines with knockdown of FOXM1, suggesting an important role for FOXM1 in the clonogenic activity of AML cells. In order to further prove a potential role for FOXM1 in AML chemoresistance, we induced an FLT3-ITD–driven myeloid neoplasm in a FOXM1-overexpressing transgenic mouse model and demonstrated significantly higher residual disease after standard chemotherapy. This suggests that constitutive overexpression of FOXM1 in this model induces chemoresistance. Finally, we performed proof-of-principle experiments using a currently approved proteasome inhibitor, ixazomib, to target FOXM1 and demonstrated a therapeutic response in AML patient samples and animal models of AML that correlates with the suppression of FOXM1 and its transcriptional targets. Addition of low doses of ixazomib increases sensitization of AML cells to chemotherapy backbone drugs cytarabine and the hypomethylator 5-azacitidine. Our results underscore the importance of FOXM1 in AML progression and treatment, and they suggest that targeting it may have therapeutic benefit in combination with standard AML therapies.

Authors

Irum Khan, Marianna Halasi, Anand Patel, Rachael Schultz, Nandini Kalakota, Yi-Hua Chen, Nathan Aardsma, Li Liu, John D. Crispino, Nadim Mahmud, Olga Frankfurt, Andrei L. Gartel

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Figure 4

Ixazomib inhibits FOXM1 and induces cell death in primary AML cells.

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Ixazomib inhibits FOXM1 and induces cell death in primary AML cells. (A)...
(A) AML mononuclear cells show significant downregulation of FOXM1 mRNA expression, as well as its downstream targets (AurkB, Cdc25B, and Plk1) following 24-hour treatment in liquid culture, as assessed by quantitative PCR. Data are expressed as the mean ± SEM (n = 11–13 patients); P < 0.05 by 1-way ANOVA followed by Tukey’s multiple comparison post test. (B) A representative Western blot from a newly diagnosed patient with complex karyotype AML (80% circulating blasts) shows significant FOXM1 protein inhibition after ixazomib treatment ex vivo with induction of cell death by caspase-3 cleavage. (C) Cytospins prepared from the mononuclear cells isolated from patients with relapsed AML show high nuclear expression of FOXM1 in blast cells and downregulation after 24-hour incubation with ixazomib (100× magnification). (D) Plot shows quantification of nuclear FOXM1 in AML cytospin slides by the Aperio microscope as a percentage compared with DMSO-treated cells; mean ± SEM of 3 different patient samples. P < 0.0001 by 1-way ANOVA followed by Tukey’s multiple comparison post test. (E) Treatment with 75 nM of ixazomib for 24 hours is associated with markedly increased apoptosis, as shown by a representative histogram of annexin V–PE staining. (F) Plot shows fold increase in cell death by flow cytometry after annexin V–PE staining compared with control cells; mean ± SEM of 5 patient samples . P < 0.05 by 1-way ANOVA followed by Tukey’s multiple comparison post test. (G) CFU assay confirms inhibition of colony-forming activity in peripheral blood mononuclear cells from AML patients following treatment with ixazomib. Colonies were imaged with the EVOS XL Core Imaging System using the 4× objective. (H) Plot shows quantification as the percentage of colony growth compared with control cells ± SEM of 4 independent triplicate experiments; P < 0.0001 by unpaired 2-tailed t test.