Author's School

Graduate School of Arts & Sciences

Author's Department/Program

Chemistry

Language

English (en)

Date of Award

5-24-2010

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Chair and Committee

John-Stephen Taylor

Abstract

Tumor and cancer cells that over express heat shock protein 70: HSP70) are found to be multidrug resistant and thermo tolerant, creating a hurdle to existing therapy. Although HSP70 is recognized as an increasingly important drug target, the protein structure of this highly conserved chaperone remains challenging for direct targeting. An alternative strategy is to inhibit the transcription of HSP70. Among known small molecule inhibitors of HSP70 induction, quercetin has very low toxicity and has the advantage of being easily modified for structure-activity studies. One part of the dissertation focuses on the identification of quercetin derivatives with improved specificity and activity, and to determine the protein targets of quercetin responsible for inhibition of heat shock induction of HSP70. A library of quercetin derivatives was synthesized and screened for their ability to inhibit HSP70 induction and at the same time not enhance HSP27 phosphorylation. The derivatives that inhibit HSP70 induction were also found to be inhibitors of both Ck2 kinase and CaMKII kinase that are known to activate heat shock transcription factor 1 that leads to HSP70 induction. A biotinylated quercetin affinity agent was also developed that was able to pull down the CK2 kinase target in vitro and several other proteins in vivo under UVA irradiation. In collaboration with mass spectrometry center, these unknown protein targets were identified by proteomic studies, and found to be previously identified chemotherapeutic targets. Another goal of this dissertation was to develop polyamide-based gene inhibitors of heat shock induction that interfere with the binding of the heat shock transcription factor. A series of polyamides that targeted the heat shock elements were designed and synthesized and demonstrated to bind the target DNA by DNase I footprinting. These polyamides were evaluated by gel shift assay for their ability to block binding of the heat shock factor and the most effective polyamide was shown to decrease HSP70 expression in Jurkat cells by western blot assay.

DOI

https://doi.org/10.7936/K7KK98TV

Comments

Permanent URL: http://dx.doi.org/10.7936/K7KK98TV

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