Exploiting ELIOT for Scaffold-Repurposing Opportunities: TRIM33 a Possible Novel E3 Ligase to Expand the Toolbox for PROTAC Design.

The field of targeted protein degradation, through the control of the ubiquitin-proteasome system (UPS), is progressing considerably; to exploit this new therapeutic modality, the proteolysis targeting chimera (PROTAC) technology was born. The opportunity to use PROTACs engaging of new E3 ligases that can hijack and control the UPS system could greatly extend the applicability of degrading molecules. To this end, here we show a potential application of the ELIOT (E3 LIgase pocketOme navigaTor) platform, previously published by this group, for a scaffold-repurposing strategy to identify new ligands for a novel E3 ligase, such as TRIM33. Starting from ELIOT, a case study of the cross-relationship using GRID Molecular Interaction Field (MIF) similarities between TRIM24 and TRIM33 binding sites was selected. Based on the assumption that similar pockets could bind similar ligands and considering that TRIM24 has 12 known co-crystalised ligands, we applied a scaffold-repurposing strategy for the identification of TRIM33 ligands exploiting the scaffold of TRIM24 ligands. We performed a deeper computational analysis to identify pocket similarities and differences, followed by docking and water analysis; selected ligands were synthesised and subsequently tested against TRIM33 via HTRF binding assay, and we obtained the first-ever X-ray crystallographic complexes of TRIM33α with three of the selected compounds.