Date of Award

1-1-2011

Document Type

Campus Access Thesis

Department

College of Pharmacy

Sub-Department

Pharmaceutical Science

First Advisor

Michael D Wyatt

Second Advisor

Campbell McInnes

Abstract

Polo like kinase 1 (PLK1) is a member of a conserved family of serine/threonine kinases that plays an important role in the regulation of mitotic progression. PLK1 is over-expressed in a number of tumor types, thus making it a good therapeutic target. Because other PLK family members are putative tumor suppressors, it is imperative that a PLK1 inhibitor be exclusive for PLK1. By targeting the non-catalytic polo box domain (PBD) of PLK1, which modulates sub-cellular localization and mitotic functions of PLK1, we can obtain selective PLK1 inhibition, which can lead to mitotic arrest and inhibit tumor cell proliferation. The structure-guided discovery strategy called REPLACE uses structure activity relationships of peptide inhibitors to generate a pharmaceutically acceptable lead molecule by replacing individual amino acid residues with non-peptide fragments. REPLACE was used to identify fragment mimetics for determinants from the endogenous phosphopeptide sequence, LLCSpTPNGL (CDC25c), known to specifically bind to the PBD of PLK1. Using this approach, FLIPs (Fragment Ligated Inhibitory Peptides) were generated and tested in a fluorescence polarization (FP) assay to determine binding potential for the PBD. Small molecule fragment alternatives for the N-terminal tripeptide (LLC) were identified through this approach and shown to possess increased binding relative to the truncated peptide. To further understand the essential binding sites and specificity determinants needed by the endogenous peptide, peptide analogs were synthesized and tested in the FP assay, so their ability to bind to PBD could be compared to that of the parent peptide. As part of the development process, peptides and FLIPs have been examined for anti-proliferative activity in cancer cells and to determine PLK1 inhibitory activity. By targeting the PBD of PLK1 using REPLACE, we believe we can develop a non-ATP competitive inhibitor that is not only PLK1 specific but is less toxic to normal cells and safer than current chemotherapeutics on the market.

Rights

© 2011, Kara Michelle Estes

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