Annals of Hematology

, Volume 91, Issue 3, pp 331–344

3,4-Diarylmaleimides—a novel class of kinase inhibitors—effectively induce apoptosis in FLT3-ITD-dependent cells

  • Florian H. Heidel
  • Thomas S. Mack
  • Elena Razumovskaya
  • Marie-Christine Blum
  • Daniel B. Lipka
  • Anne Ballaschk
  • Jan-Peter Kramb
  • Stanislav Plutizki
  • Lars Rönnstrand
  • Gerd Dannhardt
  • Thomas Fischer
Original Article

DOI: 10.1007/s00277-011-1311-3

Cite this article as:
Heidel, F.H., Mack, T.S., Razumovskaya, E. et al. Ann Hematol (2012) 91: 331. doi:10.1007/s00277-011-1311-3

Abstract

FLT3 kinase has become an attractive drug target in AML with up to 30% of cases harboring internal-tandem-duplication (ITD) mutations. For these, conferring a worse prognosis and decreased overall survival, several FLT3 tyrosine kinase inhibitors (TKIs) are currently being tested in clinical trials. However, when using these drugs as monotherapy, the problem of short duration of remissions and high incidence of TKI resistance has emerged. Here, we investigated two members of a novel class of tyrosine kinase inhibitors, 3,4-diarylmaleimides, for their efficacy on mutated FLT3 kinase. These compounds inhibit FLT3 kinase in an ATP-competitive manner and effectively inhibit phosphorylation of downstream targets. 3,4-Diarylmaleimides (DHF125 and 150) induce apoptosis in FLT3-ITD-dependent cells lines and patient blasts at low micromolar concentrations. They are retained in the cytoplasm of exposed cells for more than 24 h and synergize with chemotherapy and midostaurin. Both 3,4-diarylmaleimides show inhbition of FLT3-ITD-related kinase autophosphorylation at distinct tyrosine residues when compared to midostaurin. In conclusion, this novel group of compounds shows differential inhibition patterns with regard to FLT3 kinase and displays a promising profile for further clinical development. Currently, experiments evaluating toxicity in murine models and unraveling the exact binding mechanism are under way to facilitate a potential clinical application.

Keywords

AMLFLT3Tyrosine kinase inhibitorTyrosine phosphorylation

Supplementary material

277_2011_1311_Fig7_ESM.jpg (32 kb)
Fig. S1

Synergy of DHF compounds with either daunorubicin or cytarabine as investigated in apoptosis assays. Co-incubation of cells with DHF125 or DHF150 and daunorubicin (left panel) or cytarabine (right panel) demonstrated increased efficacy with regard to apoptosis induction. Synergy could be calculated using concentrations above the IC10 for daunorubicin and all effective doses applied for cytarabine. (JPEG 32 kb)

277_2011_1311_MOESM1_ESM.tiff (1.1 mb)
High-resolution image (TIFF 1,092 kb)
277_2011_1311_Fig8_ESM.jpg (49 kb)
Fig. S2

3,4-Diarylmaleimide compounds induce apoptosis in FLT3-ITD harboring AML blasts at micromolar concentrations while leaving normal progenitor cells largely unaffected. diarylmaleimide compounds show induction of apoptosis in primary AML blasts. Primary cells were incubated for 72 h with different inhibitor concentrations. Apoptosis was determined as the amount of cells subG1 exceeding the baseline apoptosis rate of AML cells in culture (baseline apoptosis is graphed as 0%). As indicated for every single patient sample DHF125 (a, left panel) led to increase of basal apoptosis by 10–17% while DHF150 (a, right panel) elevated the rate of apoptotic cells by 17–30%. Using bone marrow cells from healthy donors, colony formation was analyzed upon increasing doses of diarylmaleimide inhibitors—in methylcellulose supplemented with cytokines—for 10 days. Impairment of colony formation was not detectable up to a dose of 5 μM of either compound. Incubation of these cells with 10 μM of either inhibitor led to almost complete loss of colony formation, suggesting toxicity on healthy hematopoietic stem and progenitor cells. (JPEG 49 kb)

277_2011_1311_MOESM2_ESM.eps (100 kb)
High-resolution image (EPS 99 kb)

Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Florian H. Heidel
    • 1
  • Thomas S. Mack
    • 1
  • Elena Razumovskaya
    • 2
  • Marie-Christine Blum
    • 1
  • Daniel B. Lipka
    • 1
  • Anne Ballaschk
    • 1
  • Jan-Peter Kramb
    • 3
  • Stanislav Plutizki
    • 3
  • Lars Rönnstrand
    • 2
  • Gerd Dannhardt
    • 3
  • Thomas Fischer
    • 1
  1. 1.Department of Hematology and Oncology, Medical CenterOtto-von-Guericke UniversityMagdeburgGermany
  2. 2.Experimental Clinical Chemistry, Wallenberg Laboratory, Department of Laboratory MedicineSkåne University Hospital, Lund UniversityMalmöSweden
  3. 3.Department of PharmacyJohannes-Gutenberg-UniversityMainzGermany