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Degree Name

PhD (Doctor of Philosophy)

Program

Biomedical Sciences

Date of Award

12-2013

Committee Chair or Co-Chairs

William L. Stone

Committee Members

Alok Agrawal, Koyamangalath Krishnan, Sharon Campbell, Victoria Palau

Abstract

Cancer cell esterases are often overexpressed and some have chiral specificities different from those of corresponding normal cells. Carboxylesterases in particular are known to be overexpressed in several cancers. Additionally, cancer cells often exhibit high levels of intrinsic oxidative stress that is required for survival and an aggressive phenotype. We hypothesized that these 2 characteristics of cancer cells could be exploited to aid in the detection and treatment of prostate cancer. We have developed a fluorogenic ester probe that is activated by carboxylesterase to help distinguish tumorigenic cells from nontumorigenic prostate cells. Ester prodrugs have the same activation mechanism and have been thought to be a promising approach in cancer therapy. Prodrugs are inactive drugs that can be selectively activated by a specific enzyme. We have developed a chiral ester prodrug strategy using native polyacrylamide gel electrophoresis (n-PAGE) and proteomic methods to compare and identify the esterase profiles of several tumorigenic and nontumorigenic prostate cell lines. Our results showed that cell lysates from LNCaP, DU 145, and PC3 prostate cancer cell lines exhibit differential esterase activity compared with non-tumorigenic RWPE-1 prostate cell lysates when incubated with α- naphthyl acetate or α-naphthyl N-acetyl-alaninate ester substrates and a diazonium salt. We have identified oxidized protein hydrolase (OPH), a serine esterase/protease that catalyzes the removal of N-acylated residues from proteins, to be differentially expressed between some tumorigenic and nontumorigenic prostate cell lines. OPH was found to have high hydrolytic activity towards the S-isomer of α-naphthyl N-acetylalaninate (S-ANAA) chiral ester. LNCaP lysates incubated with N-acetyl-alanyl-p-nitroanilide, a known OPH substrate, had twofold higher OPH activity compared with RWPE-1 lysates. We have also developed and tested novel glutathione depleting prodrugs modeled after S-ANAA that increase oxidative stress and induced apoptosis in tumorigenic prostate cells with little effect on nontumorigenic RWPE-1 cells. These results suggest that ester molecular beacon probes and ester prodrugs may be effective in identifying and treating prostate cancer tumors that overexpress esterases with little effect on normal prostate cells.

Document Type

Dissertation - Campus Only

Copyright

Copyright by the authors.

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