Defense Date

2007

Document Type

Dissertation

Degree Name

Doctor of Philosophy

Department

Chemistry

First Advisor

Dr. Qibing Zhou

Second Advisor

Dr. Suzanne Ruder

Abstract

DESIGN, SYNTHESIS, AND EVALUATION OF DITERPENONES AS POTENT CHEMOPREVENTIVE AGENTS FOR AFB1 INDUCED CARCINOGENISIS IN HUMAN LIVER CELLS By Miguel A. Zuniga, Ph.D. A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Virginia Commonwealth University. Major Director: Qibing Zhou, Ph.D Assistant Professor Department of Chemistry Terpene quinone methides (TPQMs) have been isolated from a variety of plants and show broad activities against bacteria, fungi, and cancerous cell lines. The biological activity has been attributed to the reactive electrophilic QM moiety, this structural feature has long been recognized as an intermediate in organic synthesis and in certain biosynthetic processes. It has been shown that quinone methide structures play a key role in the chemistry of several classes of antibiotic drugs and antitumor compounds such as mitomycin C and anthracyclines. The goal of this study was to understand the basis of QM bioactivity so that terpene catechols as analogs of natural TPQMs precursors can be designed as effective chemopreventive agents.In order to investigate the oxidation pathway of terpene QM precursors, a homoconjugated diterpene catechol was synthesized. A review of the literature revealed that Cu2+ -induced oxidation of simple catechols proceeds through a two-step one electron transfer process, and o-quinone is the sole oxidation product. In contrast, our studies showed direct p-QM formation from a diterpene catechol and no o-quinone oxidation products were observed. Furthermore, the Cu2+-induced oxidation pathway of our homoconjugated diterpene catechol revealed multiple p-QM formations under aqueous conditions. The implications of these findings suggest that terpene QM precursors can cause extensive DNA damage through in situ generated hydroxyl radicals or by DNA alkylations with p-QMs. To elucidate the Cu2+-induced DNA damage mechanism, a series of catechol analogues of natural terpene QM precursors were designed to investigate potential effects of stereochemistry, substitutiional, and functional groups on nucleobase alkylation and production of reactive oxygen species. The results of these tests suggested that production of ROS was the dominant mechanism for the observed DNA damage in the Cu2+-induced oxidation regardless of stereo and structural differences of catechols or subsequent oxidation products as QM or quinone. From the DNA damage study we found that the presence of NADH significantly enhanced the extent of DNA damage by oxidation of these catechols. More specifically, in the case of alkene catechols, our results showed that DNA damage was independent of the concentration of catechols, thus providing ample evidence for production of ROS through the redox cycle of catechols/quinones. Additional support for the formation of hydroxyl radical and futile redox cycling was clearly demonstrated by comparison of the fragmentation pattern with that of a Fenton reaction. The identify of the ROS was also shown to be in the form of a Cu(I)OOH complex by radical scavenging and metal chelation experiments. Cis-terpenones were first shown to have chemoprotective activity by Dr. Zhou and colleagues. In collaboration with their efforts to identify the mode of action of cis-terpenones, another project to achieve an isotope labeled cis-terpenone was undertaken. The isotope study was employed to obtain and experimentally demonstrate the feasibility of incorporating a radioactive label in cis-terpenone for the future studies of cis-terpenone metabolism. An analysis of the deuterium labeled cis-terpenone from the isotope exchange reaction showed that the isotope was being incorporated into multiple positions through scattering processes. This non-radioactive isotope study made it possible to optimize the conditions prior to using a radioactive tritium label, which will be a requisite for future metabolic studies.As an extension of this work, a structure activity relationship (SARs) study was undertaken with a focus on improving the physiochemical properties and chemical stability of cis-terpenones. The primary purpose of this study was to attempt to explain the reason for the observed protective effects of cis-terpenones against AFB1. Considerable efforts were made to introduce an unsaturated double bond in the structure of cis-terpenone by an intramolecular Pd (II) catalyzed Heck reaction. Unfortunately, this method was unsuccessful and which was attributed to the disconnection of our starting material during the formation of an enolate intermediate. A second model study to generate desired coumarin and ditepene related structures was investigated with a Diels Alder [4+2] cycloaddition reaction. After numerous attempts, we found that the successfull synthesis of these compounds was highly dependent on the temperature, solvent, and the use of stabilizers in the reaction. Finally, the targeted diterpene analogs were screened for protective effects against AFB1 by the MTT cell viability assay. However, these preliminary data showed that additional structural features and key modifications are still required to better correlate the structure with the mechanism of chemoprotection.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

June 2008

Included in

Chemistry Commons

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