PROTAC & Molecular Glue Tools for Drug Discovery

Tools Accelerating Targeted Protein Degradation Research

PROTACs (Proteolysis-targeting Chimeras)

PROTAC are hetero-bifunctional small molecules that contain two functional ligands connected via a linker; one ligand binds to a target protein and the other ligand binds to an E3 ligase (Figure 1).

Figure 1: PROTAC structure. The target protein of interest (POI) and the E3 ligase are linked via a linker. This leads to the POI ubiquitination.

PROTACs artificially hijack the components of the UPS (Ubiquitin Proteasome System) to degrade a target protein. By bringing the target protein and the E3 ligase into proximity, the POI is ubiquitinated and degraded via the proteasome (Figure 2).

The use of PROTACs for drug discovery approaches became a major therapeutic strategy for previously “undruggable” proteins in oncology, immunology, and neurodegeneration - opening a new therapeutic avenue across multiple disease areas by allowing the elimination of proteins that cannot be inhibited by classical small molecules and a rapid and reversible control of protein levels.

Figure 2: PROTACs mechanism of action. PROTACs bind the E3 ligase and the protein of interest (T) leading to its ubiquitination and degradation via the proteasome.

However, this mechanism remains complex and the PROTAC discovery platform faces several challenges with an unmet need for functional assays that can study the effects of PROTACs on the ubiquitination of targets and their degradation. LifeSensors has developed high-throughput methods to directly monitor PROTAC mediated ubiquitylation and degradation of target proteins, including functional ubiquitination and degradation assays in cells. PROTAC-mediated ubiquitination in vitro permits rapid screening of compound libraries and allows simplification of the medicinal chemistry approach to rationally design potent molecules.

Molecular Glue

Molecular glues are small molecules that stabilize interactions between a target protein and an E3 ligase (Figure 3). 

Figure 3: Molecular glue structure.

They have the same function as PROTACs and are divided in two classes:

1. Glues that bind to an E3 ligase to change its conformation so that it binds and ubiquitinates a neo-substrate
2. Glues that bind to a target protein, change its conformation such that the structure is altered, exposing lysines to be ubiquitinated by neighboring ligases. Depending upon the type of ligase employed for ubiquitination, if lysine 48 chains are built on the target protein, the proteasome will recognize the target and degrade it. If lysine 63 chains are built on the target protein, the protein may be trafficked to another compartment with a loss of function. However, lysine 63 chains most likely promote autophagy, and the protein will be degraded by lysosomal degradation mechanisms.

LifeSensors has a variety of tools to discover and tease out the mechanism of molecular glue mediated ubiquitination and degradation.

DUBTACs (Deubiquitinating targeting chimeras) 

DUBs are a large group of proteases that remove ubiquitin from proteins, rescuing ubiquitinated proteins from proteasome attack (Figure 4). They antagonize the E3 ligase activity, regulate protein trafficking and are implicated in epigenetic chromatin remodeling.

Figure 4: DUBs mechanism of action. 

DUBTACs consists of three components: (1) a DUB recruiter, (2) a target protein binder and (3) a linker connecting both entities. DUBTACs recruit DUBs to a target protein and remove ubiquitin chains, resulting in stabilization of target proteins, restoring protein levels, function, and their rescue from degradation via the proteasome.

Figure 5: DUBTACs mechanism of action

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  Article content created by Tebubio using courtesy materials provided by LifeSensors.