Novel approaches to identify intrinsic interactors of ubiquitin E3 ligases: Towards rational design of molecular glue degraders.

Targeted Protein Degradation (TPD) is a powerful strategy that utilises the body's mechanism to degrade unwanted proteins. It is based on small molecules that bring together an E3 ligase and an unwanted protein, the E3 ligase tags the unwanted protein with ubiquitin, which leads to its degradation by the proteasome. The small molecules used in TPD are classified into PROteolysis Targeted Chimeras (PROTACs) and molecular glues. PROTACs interact with both proteins, bringing them together, whereas molecular glues only have affinity for one of the proteins, or to the pre-formed protein-protein complex.
Up to this moment, molecular glues have been mainly discovered by serendipity. Recent discoveries have shown that most molecular glues stabilise some surface complementarity between the E3 ligase and the target protein. Many examples have shown that both proteins have a pre-existent interaction, which is enhanced in presence of the glue and leads to the degradation of the target protein. We hypothesize that if we were able to identify these E3 ligase-target protein pairs, we would accelerate the discovery of molecular glues.
The aim of this project is to design and develop a novel approach to identify intrinsic interactions between E3 ligases and potential target proteins, that might be otherwise missed by conventional methods. We have decided to use as a model the two most widely used E3 ligases in the TPD field: VHL and CRBN. These E3s have known intrinsic interactions that are enhanced by known molecular glue degraders, which will allow us to benchmark our method development. In order to achieve this, we will use a number of proteomic approaches to identify proteins which have an interaction with the E3 ligase. Once the interaction is validated via low throughput methods, the next step will be to design a molecular glue that stabilises and enhances the interaction. Subsequently, we will expand the approach to target novel E3 ligases.