FOXO transcription factors are critical regulators of cell homeostasis and steer cell death, differentiation and longevity in mammalian cells. By combined pharmacophore-modeling-based in silico and fluorescence polarization-based screening we identified small molecules that physically interact with the DNA-binding domain (DBD) of FOXO3 and modulate the FOXO3 transcriptional program in human cells. The mode of interaction between compounds and the FOXO3-DBD was assessed via NMR spectroscopy and docking studies. We demonstrate that compounds S9 and its oxalate salt S9OX interfere with FOXO3 target promoter binding, gene transcription and modulate the physiologic program activated by FOXO3 in cancer cells. These small molecules prove the druggability of the FOXO-DBD and provide a structural basis for modulating these important homeostasis regulators in normal and malignant cells.

Department of Organic Chemistry Faculty of Science Charles University Prague Czech Republic

Department of Pediatrics 1 Medical University Innsbruck Innsbruck Austria

Department of Pediatrics 2 Medical University Innsbruck Innsbruck Austria

Department of Pharmaceutical and Medicinal Chemistry Paracelsus Medical University Salzburg Salzburg Austria

Department of Physical and Macromolecular Chemistry Faculty of Science Charles University Prague Czech Republic

Department of Structural Biology of Signaling Proteins Institute of Physiology Division BIOCEV The Czech Academy of Sciences Prague Czech Republic

Division of Molecular Pathophysiology Biocenter Medical University Innsbruck Innsbruck Austria

Independent Data Lab UG Munich Germany

Pharmaceutical Chemistry Institute of Pharmacy University of Innsbruck Innsbruck Austria

Tyrolean Cancer Research Institute Innsbruck Austria

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