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Designed nanomolar small-molecule inhibitors of Ena/VASP EVH1 interaction impair invasion and extravasation of breast cancer cells

Authors

  • M. Barone
  • M. Müller
  • S. Chiha
  • J. Ren
  • D. Albat
  • A. Soicke
  • S. Dohmen
  • M. Klein
  • J. Bruns
  • M. van Dinther
  • R. Opitz
  • P. Lindemann
  • M. Beerbaum
  • K. Motzny
  • Y. Roske
  • P. Schmieder
  • R. Volkmer
  • M. Nazaré
  • U. Heinemann
  • H. Oschkinat
  • P. Ten Dijke
  • H.G. Schmalz
  • R. Kühne

Journal

  • Proceedings of the National Academy of Sciences of the United States of America

Citation

  • Proc Natl Acad Sci U S A 117 (47): 29684-29690

Abstract

  • Battling metastasis through inhibition of cell motility is considered a promising approach to support cancer therapies. In this context, Ena/VASP-depending signaling pathways, in particular interactions with their EVH1 domains, are promising targets for pharmaceutical intervention. However, protein-protein interactions involving proline-rich segments are notoriously difficult to address by small molecules. Hence, structure-based design efforts in combination with the chemical synthesis of additional molecular entities are required. Building on a previously developed nonpeptidic micromolar inhibitor, we determined 22 crystal structures of ENAH EVH1 in complex with inhibitors and rationally extended our library of conformationally defined proline-derived modules (ProMs) to succeed in developing a nanomolar inhibitor (K(d) = 120 nM, MW = 734 Da). In contrast to the previous inhibitor, the optimized compounds reduced extravasation of invasive breast cancer cells in a zebrafish model. This study represents an example of successful, structure-guided development of low molecular weight inhibitors specifically and selectively addressing a proline-rich sequence-recognizing domain that is characterized by a shallow epitope lacking defined binding pockets. The evolved high-affinity inhibitor may now serve as a tool in validating the basic therapeutic concept, i.e., the suppression of cancer metastasis by inhibiting a crucial protein-protein interaction involved in actin filament processing and cell migration. SIGNIFICANCE: Protein–protein interactions mediated by proline-rich motifs are involved in regulation of many important signaling cascades. These motifs belong to the most abundant recognition motifs in the eukaryotic genome and preferentially adopt a left-handed polyproline helix II, a secondary structure element that has been notoriously difficult to mimic with small molecules. Here, we present a structure-guided design effort yielding a toolkit of chemical entities that enables rational construction of selective small molecule inhibitors for these protein domains. We succeeded in developing an inhibitor for the Ena/VASP protein family that is active in vivo and reduces extravasation of invasive breast cancer cells in a zebrafish model.


DOI

doi:10.1073/pnas.2007213117