Small-molecule induced degradation of fascin as a strategy for metastasis prevention

Memarzadeh, Sarah (2022) Small-molecule induced degradation of fascin as a strategy for metastasis prevention. PhD thesis, University of Glasgow.

Abstract

The risk of being diagnosed with cancer during our lifetime is estimated to be around 50%, and – although survival rates have vastly improved over the last few decades – cancer remains the second leading cause of death worldwide. Metastasis, the ability of cancer cells to spread and colonise throughout the body, is the major contributor to cancer mortality. Identification of suitable targets for the development of anti-metastatic therapeutics is challenging due to the complex nature of the metastatic process. Cancer cell motility and formation of invasive actin structures are vital for a multitude of steps in this metastatic cascade. The actin-bundling protein fascin has been found to be crucial for the formation of those invasive membrane protrusions and its expression correlates with aggressive metastatic cancers, however, fascin is not found in healthy epithelial tissue. Despite representing an attractive target, disruption of protein–protein interactions of structural proteins with traditional small-molecule drugs remains a difficult task in drug discovery. Therefore, this project aims at overcoming this challenge by recruiting the cell’s natural protein degradation mechanism to the scaffold protein in order to induce its targeted degradation by means of proteolysis-targeting chimeras (PROTACs). Selective degradation of the pro-metastatic target could impede assembly of invasive membrane protrusions, reduce invasiveness of cancer cells and thereby prevent formation of metastases.

Chapter 1 discusses the process of metastasis and the role of cancer cell motility in the metastatic cascade. It goes on to describe actin-based cell membrane protrusions implicated with cell migration and invasion, and fascin’s involvement in the formation of invasive actin structures and cancer cell metastasis. Drug discovery efforts to develop a small molecule capable of inhibiting the actin-bundling activity of fascin are mentioned. The second part of Chapter 1 details the mechanisms behind cellular protein degradation and how it can be exploited therapeutically using PROTACs. Considerations to be taken into account when designing bi-functional degrader molecules are explained, and evaluation of physicochemical properties of PROTACs are discussed. The last part of Chapter 1 briefly summarises the project’s aims and hypotheses.

Chapter 2 describes the development of a HaloTag-based model system used to prove the concept for targeted protein degradation of fascin. The first part of Chapter 2 reports the modular design and synthesis of active compounds and mono-functional controls. The second part of Chapter 2 details biological assays used to determine compound activity as well as experiments assessing the effect on cell function and phenotype, emphasising the importance of using cell lines suitable for evaluating proposed outcomes.

Chapter 3 details the design and synthesis of three series of fascin-targeting PROTACs based on different fascin ligands, as well as evaluation of physicochemical properties of the degrader molecules. The second part of Chapter 3 describes biological assays carried out to test activity of the compounds in different cell lines and their binding to fascin. It highlights difficulties encountered with cell permeability and attempted strategies to overcome these challenges.

Chapter 4 concludes on the findings of the PhD project and proposes future work to be carried out.

Chapter 5 describes experimental protocols for the biological and chemical components of this project.

The appendix contains compound characterisation data as well as supplementary experimental data for biological experiments.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Additional Information: Supported by funding from Cancer Research UK.
• Colleges/Schools: College of Science and Engineering > School of Chemistry
• Supervisor's Name: France, Dr. David
• Date of Award: 2022
• Depositing User: Theses Team
• Unique ID: glathesis:2022-82828
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 27 Apr 2022 09:31
• Last Modified: 05 Oct 2022 13:55
• Thesis DOI: 10.5525/gla.thesis.82828
• URI: https://theses.gla.ac.uk/id/eprint/82828