Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.715244
Title: Combining library screening approaches, and modifying peptides with helix constraints, to generate novel antagonists of oncogenic Activator Protein-1
Author: Baxter, Daniel
Awarding Body: University of Bath
Current Institution: University of Bath
Date of Award: 2016
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Abstract:
Activator Protein-1 (AP-1) is an oncogenic transcription factor that is dysregulated in numerous human cancers, making it an attractive therapeutic target. AP-1 forms via interaction of cJun and cFos proteins, which intertwine to generate a ‘coiled coil’ (CC) structure. Thus, the cJun/cFos α-helical CC domains responsible for dimerisation are appealing targets for inhibiting AP-1 formation and activity. Helical peptide antagonists that sequester cJun can be derived from the cFos CC domain by selection of more optimal amino acids for increased binding affinity. Peptides can then be downsized and modified to improve therapeutic potential. Two approaches aimed to identify novel short peptides against cJun. The first was to covalently cyclise amino acid side chains in existing cFos-derived peptide “FosW”, with the aim of constraining FosW into a stable helix to allow downsizing without significant loss of binding structure and affinity. Using circular dichroism spectroscopy and isothermal titration calorimetry, a series of helix constrained peptides were characterised, from which a peptide was identified that retained 88 % of FosW binding affinity whilst being 22 % shorter, and which entered breast cancer cells in vitro, with preliminary data suggesting potential ability to inhibit AP-1 in cellulo. The second approach was to combine two existing high-throughput peptide selection systems, with the aim of benefitting from overlap in their strengths and weaknesses. Combination of in vitro CIS display and in cellulo Protein-fragment Complementation Assay successfully isolated a high affinity peptide from a hugely diverse library, and future refinements to further exploit this approach, particularly for short peptide selection, were formulated. Thus, molecules and techniques derived here may expedite the future development of therapies for cancers featuring AP-1 dysregulation.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.715244  DOI: Not available
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