Abstract BIOCHEMICAL ACTIONS OF A NOVEL CIS-TERPENONE By Lin Zhang, Ph.D. A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Virginia Commonwealth University. Virginia Commonwealth University, 2012 Director: Qibing Zhou, Ph.D Assistant Professor Department of Chemistry
Quinone methides are reactive species due to their electrophilicity, and natural quinone methide analogs have broad biological activities. Based on known actions of cytochrome P450 enzymes, we hypothesized that trans-terpenones can be metabolized in cells to form a cascade of metabolite products, which may be biologically active due to the generation of quinine and quinone methide reactive intermediates. Therefore, 3-hydroxy-cis-terpenone (HCT) and analogs were designed and synthesized in our lab to investigate this hypothesis. As a first step in testing this hypothesis, we examined effects of cis-terpenones on the viability of HepG2 cells exposed to aflatoxin B1 (AFB1). Cis-terpenones combined with AFB1 in HepG2 cells increased cell viability suggesting chemopreventive effects against AFB1. Further study revealed a mechanism for this effect: HCT and oxidized HCT (OHCT) inhibited activity of P450 1A/B, which metabolizes AFB1 to toxic metabolites, and thereby protected cells against AFB1 induced cytotoxicity. Additional studies demonstrated that HCT inhibits accumulation of tritium-labeled AFB1 in HepG2 cells. To investigate the mechanism of this effect we first determined whether specific transporters affect the accumulation of AFB1 in HepG2 cells. Effects on AFB1 accumulation by pH, selected ions, inhibitors or competitive substrates of organic cation transporters (OCT), organic anion transporters (OAT/OATP), and multi-drug-resistant efflux transporters eliminated effects of major groups of these transporters on AFB1 accumulation. The data indicated one or more unidentified proton dependent transport mechanism(s) modulate cellular accumulation of AFB1 and further demonstrated HCT inhibits accumulation of AFB1 in cells by decreasing AFB1 binding to intracellular proteins. Based on its potential chemoprotective actions, other biological actions of HCT were screened. Based on evidence that terpenes modulate immune cell production of cytokines, we examined effects of HCT on production of interleukin-6 (IL-6) and tumor necrosis factor-α (TNFα) by cultured macrophages. HCT reduced lipopolysaccharide-stimulated release of IL-6 and TNFα. MES-SA/DX5 cells that over-express the efflux transporter MDR1 were used to examine effects of OHCT on accumulation of the MDR1 substrate estrone-sulfate and to verify that MDR1 does not transport unconjugated AFB1. OHCT at a high concentration (120 µM) reduced cellular accumulation of estrone sulfate suggesting enhanced MDR1-mediated efflux
| Identifer | oai:union.ndltd.org:vcu.edu/oai:scholarscompass.vcu.edu:etd-1413 |
| Date | 01 January 2012 |
| Creators | Zhang, Lin |
| Publisher | VCU Scholars Compass |
| Source Sets | Virginia Commonwealth University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Theses and Dissertations |
| Rights | © The Author |
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