The development of novel molecular technologies for screening and understanding membrane interactions of proteolysis targeting chimeras (PROTACs)

Lead Research Organisation: Imperial College London
Department Name: Dept of Chemistry

Abstract

Small molecule-mediated inhibition of protein function is the mechanism of action of the vast majority of clinical agents, however a key limitation of this approach is that pharmacologically relevant protein inhibition is often only achieved upon more than 90 percent target binding, requiring high dosing which can lead to adverse effects. There is subsequently a well-established demand for new and more efficient therapeutic strategies to directly inhibit the function of target proteins. Proteolysis targeting chimeras (PROTACs) are bifunctional molecules that exploit the ubiquitin-proteasome pathway for target protein degradation. PROTACs comprise of a ligand for a protein target connected via a linker to a ligand for an E3 ubiquitin ligase. Upon entering the cell, the PROTAC binds to both the target protein and an E3 ligase, which ubiquitinates the target protein and induces degradation by proteasome.

PROTAC development is now reaching the stage where their use as novel therapeutics is being seriously considered and explored. As such, focus has shifted towards the pharmacokinetics of newly developed PROTACs with an aim to produce effective, bioavailable compounds that might be suitable candidates for clinical trials. One of the key considerations in this process is the ability of the PROTAC to diffuse across biological membranes and reach their protein counterparts.

A mechanistic understanding of how PROTACs are able to cross membranes (including the relative roles of active and passive diffusion) and how the efficacy of this translocation process correlates with the structure of the PROTAC (biofunctional moieties and linkage motifs) and are vital to their successful development.

Through physical science innovation this project will seek to study how molecular interactions between membranes and PROTACs correlate with established values of PCtotal and PCaqueous for PROTACs using atomistic computational and biomembrane experimental approaches. This will include the development of novel droplet interface bilayer patch clamp approaches and Amber parameters for GSK ligands. During these studies the student will study permeation into (SS NMR, SAXS and AMBER Simulations) and translocation of PROTACs across lipid bilayers (DIB/patch clamp/Rheo DIB/microscopy assays and AMBER Simulations) with reference to membrane rigidity, spontaneous curvature, membrane charg, membrane fluidity, membrane asymmetry, membrane permeability and headgroup type . These studies will enable the student to decouple which global biophysical properties of membrane systems account for differences in observed PROTAC behaviour and the mode of PROTAC-membrane interaction.

The next phase of the project will establish the molecular basis of the PROTAC-membrane interactions based upon the emerging hypothesis from the above results. The student will aim to predict differences in degree of interaction with biological membranes of a series of (systematically) modified PROTACs and correlate this set against in-house GSK measurements. By varying the chemical fragments present in the PROTACs under study and monitoring their behaviour at the membrane they will evaluate which (and nature of) of the sites on PROTACs are key for downstream activity. This will allow us to develop a predictive model which can be tested both in-vitro and in-silico using model membrane systems

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
EP/R512540/1 01/10/2017 30/09/2021
1929275 Studentship EP/R512540/1 30/09/2017 30/09/2021 Robert Strutt