Abstract 3411: Targeting of EWS-FLI1 with small molecule YK-4-279 reduces xenograft growth by disruption of disordered protein-protein interactions

Abstract

Abstract Introduction: EWS-FLI1 oncoprotein is expressed as a result of a tumor-specific translocation that occurs in patients with Ewing's Sarcoma. Development of strategies to therapeutically target EWS-FLI1 revealed its significant intrinsically disordered regions. The intrinsically disordered regions enhance protein-protein interactions while presenting an opportunity for small molecule targeting. We hypothesized that a small molecule could bind to EWS-FLI1 and prevent a key protein-protein interaction with RNA Helicase A (RHA) leading to decreased cell growth. Methods: Small molecule library screening was performed using surface plasmon resonance. Chemical optimization followed by fluorescence polarization enhanced and confirmed compound binding to EWS-FLI1, respectively. Testing of compounds included co-immunoprecipitation, cell growth, caspase-3, and xenografts. Results: A library of 3000 compounds (NCI, DTP) was screened for EWS-FLI1 binding. NSC635437 had an RUactual: RUtheorhetic of 0.9 and was chosen for further evaluation based upon potential for chemical derivatization. One optimized compound, YK-4-279, significantly reduced the protein-protein interaction between EWS-FLI1 and GST-RHA(647-1075). SPR measurement of the KD was 9.48 microM for the affinity of YK-4-279 with EWS-FLI1. YK-4-279 has an IC50 of 1 microM for a series of ESFT cell lines, while cell lines that lack EWS-FLI1 are 10-30 fold more resistant to the compound. YK-4-279 reduced the growth of Ewing's Sarcoma xenografts while not affecting the growth of PC3 prostate tumors. PONDR analysis of EWS-FLI1 and RHA indicate that their interaction is occurring in an ordered-disordered transition region of RHA. Conclusions: The lead compound derivative YK-4-279 demonstrates that the EWS-FLI1/RHA interaction can be blocked with a detrimental effect on Ewing's Sarcoma cells both in vitro and in vivo. These findings both validate the interaction of EWS-FLI1 with RHA as an oncogenic target and provide a lead compound for therapeutic development. Additional studies are underway to evaluate the contributions of the disordered regions to small molecule binding and developing strategies of optimization. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3411.