A fluorescence polarization based high throughput screening assay to discover small molecule inhibitors of coiled coil tetramerization - ICS84 The 84th Annual Meeting of the Israel Chemical Society

Mai Shamir ¹ Hadar Amartely ² Mario Lebendiker ² Haim Barr ³ Shai Izraeli ⁴ Assaf Friedler ¹

¹Institute of Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
²Wolfson Center for Applied Structural Biology, The Hebrew University of Jerusalem, Jerusalem, Israel
³The Nancy and Stephen Grand National Center for Personalized Medicine, Weizmann Institute of Science, Rehovot, Israel
⁴Rina Zaizov Pediatric Hematology Oncology Division, Schneider Children’s Medical Center, Petach Tikva, Israel

Protein-protein interactions (PPI) represent an emerging and highly challenging class of therapeutic targets. PPI are central to all biological processes and are often deregulated in diseases. Despite the importance of PPI in biology, they are extremely difficult to target therapeutically due to their flexibility, large size, and the complex topology of PPI interfaces. These features, together with poor compatibility of PPI interfaces with small molecules available in libraries used for screening, limited the applicability of classical drug screening approaches for identification of PPI inhibitors.

STIL is a 1288 residues, 150kDa, cytosolic protein that plays an important role in mitosis by regulating centrosomal duplication. STIL is over-expressed in numerous types of cancer, and its expression is associated with metastasis. STIL is a promising new potential target for cancer therapy. The Coiled Coil domain (CCD) of STIL (residues 718-749) is the major determinant that mediates STIL oligomerization. Since STIL oligomerization is essential for its activity its inhibition could serve as an anti-cancer strategy. We developed a fluorescence polarization (FP) based high throughput screen (HTS) to discover small-molecules that inhibit the CCD oligomerization. Screening of over 94,000 different compounds and IC₅₀ measurements of the discovered Hits resulted in 10 final Hits with IC₅₀<75µM. The direct influence of the Hits on the oligomerization of the CCD was validated using anion exchange chromatography. Our results open the way for specific targeting of coiled coil domains of proteins, which interact in a similar hydrophobic mechanism to STIL.