2. Academic Validation
  3. Biophysical and pharmacokinetic characterization of a small-molecule inhibitor of RUNX1/ETO tetramerization with anti-leukemic effects

Biophysical and pharmacokinetic characterization of a small-molecule inhibitor of RUNX1/ETO tetramerization with anti-leukemic effects

  • Sci Rep. 2022 Aug 19;12(1):14158. doi: 10.1038/s41598-022-17913-6.
Mohanraj Gopalswamy 1,  Tobias Kroeger 1,  David Bickel 1,  Benedikt Frieg 2,  Shahina Akter 1,  Stephan Schott-Verdugo 1 2 3,  Aldino Viegas 4 5,  Thomas Pauly 2 4,  Manuela Mayer 6,  Julia Przibilla 6,  Jens Reiners 7,  Luitgard Nagel-Steger 2 4,  Sander H J Smits 7,  Georg Groth 8,  Manuel Etzkorn 4 9,  Holger Gohlke 10 11 12
Affiliations

Affiliations

  • 1 Institute for Pharmaceutical and Medicinal Chemistry, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany.
  • 2 Institute of Biological Information Processing (IBI-7: Structural Biochemistry), Forschungszentrum Jülich GmbH, Wilhelm-Johnen-Str, 52425, Jülich, Germany.
  • 3 John von Neumann Institute for Computing (NIC), Jülich Supercomputing Centre (JSC), Institute of Bio- and Geosciences (IBG-4: Bioinformatics), Forschungszentrum Jülich GmbH, Wilhelm-Johnen-Str, 52425, Jülich, Germany.
  • 4 Institute for Physical Biology, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany.
  • 5 UCIBIO, Chemistry Department, Faculty of Sciences and Technology, NOVA University of Lisbon, 2829-516, Caparica, Portugal.
  • 6 Pharmacelsus GmbH, Science Park 2, 66123, Saarbrücken, Germany.
  • 7 Institute of Biochemistry and Center for Structural Studies, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany.
  • 8 Institute of Biochemical Plant Physiology, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany.
  • 9 Jülich Center for Structural Biology (JuStruct), Forschungszentrum Jülich GmbH, Wilhelm-Johnen-Str, 52425, Jülich, Germany.
  • 10 Institute for Pharmaceutical and Medicinal Chemistry, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany. gohlke@uni-duesseldorf.de.
  • 11 Institute of Biological Information Processing (IBI-7: Structural Biochemistry), Forschungszentrum Jülich GmbH, Wilhelm-Johnen-Str, 52425, Jülich, Germany. gohlke@uni-duesseldorf.de.
  • 12 John von Neumann Institute for Computing (NIC), Jülich Supercomputing Centre (JSC), Institute of Bio- and Geosciences (IBG-4: Bioinformatics), Forschungszentrum Jülich GmbH, Wilhelm-Johnen-Str, 52425, Jülich, Germany. gohlke@uni-duesseldorf.de.
PMID: 35986043 DOI: 10.1038/s41598-022-17913-6
Abstract

Acute myeloid leukemia (AML) is a malignant disease of immature myeloid cells and the most prevalent acute leukemia among adults. The oncogenic homo-tetrameric fusion protein RUNX1/ETO results from the chromosomal translocation t(8;21) and is found in AML patients. The nervy homology region 2 (NHR2) domain of ETO mediates tetramerization; this oligomerization is essential for oncogenic activity. Previously, we identified the first-in-class small-molecule inhibitor of NHR2 tetramer formation, 7.44, which was shown to specifically interfere with NHR2, restore gene expression down-regulated by RUNX1/ETO, inhibit the proliferation of RUNX1/ETO-depending SKNO-1 cells, and reduce the RUNX1/ETO-related tumor growth in a mouse model. However, no biophysical and structural characterization of 7.44 binding to the NHR2 domain has been reported. Likewise, the compound has not been characterized as to physicochemical, pharmacokinetic, and toxicological properties. Here, we characterize the interaction between the NHR2 domain of RUNX1/ETO and 7.44 by biophysical assays and show that 7.44 interferes with NHR2 tetramer stability and leads to an increase in the dimer population of NHR2. The affinity of 7.44 with respect to binding to NHR2 is Klig = 3.75 ± 1.22 µM. By NMR spectroscopy combined with molecular dynamics simulations, we show that 7.44 binds with both heteroaromatic moieties to NHR2 and interacts with or leads to conformational changes in the N-termini of the NHR2 tetramer. Finally, we demonstrate that 7.44 has favorable physicochemical, pharmacokinetic, and toxicological properties. Together with biochemical, cellular, and in vivo assessments, the results reveal 7.44 as a lead for further optimization towards targeted therapy of t(8;21) AML.

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