Development of WWP2 ubiquitin ligase inhibitors using biophysical, structural and synthetic approaches.

Hughes, Gregory (2019) Development of WWP2 ubiquitin ligase inhibitors using biophysical, structural and synthetic approaches. Doctoral thesis, University of East Anglia.

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Abstract

The ubiquitin- proteasome pathway (UPS) involves the targeting of proteins for their degradation by the 26S proteasome. Ubiquitin is covalently attached to targeted proteins via a three step enzymatic cascade. The three enzymes involved in this process are known as E1 activating, E2 conjugating and E3 ligases. The key to substrate selectively is within the E3 ligases. E3 ligases can be divided into three domain containing groups: HECT, RING and RING between RING (RBR). Of particular interest is the Nedd4 group of HECT ligases. The Nedd4 group all share similar domain make up where they have substrate recruiting WW domains and a catalytic HECT domain, responsible for transferring ubiquitin onto the substrates. Themis-expressionofamemberofthisgroup,WWP2 ,has been linked to tumour growth in various cancers. Targeting the UPS for cancer therapeutic use has already been shown to be achievable with the proteaseome inhibitor Bortezomib. Targeting the more specific WWP2 ubiquitin ligase could lead to less side effects and greater efficacy. Research into developing inhibitors for WWP2 could lead to the discovery of novel therapeutics. The aim of this thesis is to design, synthesise and evaluate novel WWP2 ubiquitin ligase inhibitors. Different methodologies are explored in order to reveal WWP2 small molecule binding events. A thermal shift screen is employed to search for binders, and orthogonal methods for validation are tested. Using these methods, novel WWP2 ligands are discovered with inhibition activity. Crystallography is used to structural elucidate a novel binding site for WWP2 ligands. Data acquired from ligand NMR techniques is used to validate this binding site. The combined X-ray crystallography and ligand NMR data is used to inform organic
synthesis of analogues for a particular inhibitor. These analogues are tested for inhibition against WWP2 and Nedd4 autoubiquitination. IC50 values generated from this analogue series further validate the novel binding site and also offer higher levels of activity of below 1 µM.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Science > School of Chemistry
Depositing User: Katherine Whittaker
Date Deposited: 14 Feb 2020 11:04
Last Modified: 14 Feb 2020 11:04
URI: https://ueaeprints.uea.ac.uk/id/eprint/74200
DOI:

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