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- FitzGerald, Edward A., 1991-
(författare)
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Fragment-based drug discovery : Novel methods and strategies for identifying and evolving fragment leads
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The need for new drugs became ever more apparent in the year 2020 when the world was faced with a viral pandemic. How drugs are discovered and their relevance to society became part of daily discussions in workplaces and homes throughout the world. Consequently, efficient strategies for preclinical drug discovery are clearly needed. The aim of this thesis has been to contribute to the drug discovery process by developing novel methods for fragment-based drug discovery (FBDD), a rapidly developing approach where success relies on access to sensitive and informative analytical methods as well as chemical compounds with suitable properties. This process is fundamentally dependent on the interplay between scientists and engineers across biology, chemistry and physics. This project is characterized by the development and implementation of novel biophysical methods over a series of studies, which are subdivided into: 1. Development of biosensor assays and approaches for challenging targets, 2. Discovery of fragments targeting dynamic proteins using biosensors, and 3. Reconstruction of ligands using fragment-based strategies.A selection of diverse targets was used as challenging prototypes for the target agnostic methodologies described herein. The targets in focus were: acetylcholine-binding protein (AChBP), a soluble homologue of ligand gated ion channels, and two complex multi-domain epigenetic enzymes lysine specific demethylase 1 (LSD1) and SET and MYND domain-containing protein 3 (SMYD3). Expression, purification, engineering of protein variants, and biochemical characterization were required before robust screening strategies could be established.Three types of biosensors, based on different time-resolved and very sensitive detection principles (SPR, SHG, GCI), were used to identify and characterize the kinetics of the interactions of novel fragments for the proteins. For SPR, a variety of multiplexed assays were designed for the screening of fragments against difficult targets. Notably, it led to the identification of an allosteric ligand and site in SMYD3, which was subsequently characterized kinetically and structurally using X-ray crystallography, and further evolved using computational approaches.An innovative SHG assay for the specific detection of ligands inducing conformational changes was developed and used for fragment screening against AChBP. It revealed that fragments with a potential to serve as functional regulators of ligand gated ion channels can be identified using this technique. The combined application of the novel biophysical and computational approaches enabled the identification of useful starting points for drug discovery projects.
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| 2. |
- Talibov, Vladimir O., 1991-, et al.
(författare)
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Biophysical approach reveals a novel allosteric ligand binding site of SMYD3 histone methyltransferase
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- SET-and MYND-domain containing protein 3 (SMYD3) is a lysine methyltransferase that plays a role in epigenetic regulation. The protein was shown to have cancerogenic activities and is considered to be a perspective drug target. Here, we propose a Surface Plasmon Resonance-based (SPR) biophysical platform to aid SMYD3 drug discovery. The SPR screening assay was validated with a small subset of drug-like compounds, and resulted in an assay hit. The hit compound-SMYD3 complex structure was solved, and a new allosteric ligand binding site of the protein was revealed. The interaction was found localized within the previously reported SMYD3-heat shock protein 90 (HSP90) recognition site, thereby rendering the hit compound as a perspective candidate for a development of a protein-protein interface inhibitor.
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