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BROCCOLI ISOTHIOCYANATES AS CHEMOPREVENTIVE AGENTS AND EPIGENETIC MODULATORS OF BLADDER CANCER

Abbaoui, Besma
http://rave.ohiolink.edu/etdc/view?acc_num=osu1312835540

Abstract Details

2011, Doctor of Philosophy, Ohio State University, Integrated Biomedical Science Graduate Program.
Bladder cancer is the fifth most common neoplasm and one of the most expensive cancers to treat and monitor. Epidemiologic evidence suggests that cruciferous vegetables, particularly broccoli, may reduce bladder cancer risk. These vegetables contain high concentrations of glucosinolates (GLUs) that are enzymatically hydrolyzed to bioactive isothiocyanates (ITCs) by myrosinase. The most abundant ITC produced in broccoli, sulforaphane (SFN) is under investigation for its potential chemopreventive activity. Our objectives were to characterize GLUs and ITCs in broccoli and broccoli sprouts, determine their absorption and bioavailability in mice, define their bioactivity in bladder cancer in vitro and in vivo and define their potential mechanisms of action. Broccoli and broccoli sprouts GLUs and ITCs were characterized by HPLC-MS-MS. In vitro effects of non-hydrolyzed (GLUs) and hydrolyzed broccoli and broccoli sprouts (ITCs), and pure ITCs sulforaphane (SFN), erucin (ECN), iberin and allyl ITC, were studied on normal bladder urothelial cells and a panel of human bladder cancer cell lines, representing the spectrum of bladder cancer biology (RT4: non-invasive; J82 and UMUC3: invasive). Cell viability (MTS and SRB assays), apoptosis (caspase-3/7 activity and PARP cleavage) and cell cycle analysis (flow cytometry) were performed. For in vivo studies, female athymic nude mice were subcutaneously injected with UMUC3 cells and fed diets containing 4% freeze-dried broccoli sprouts, or 2% freeze-dried broccoli sprouts extract; or gavaged daily with 295 µmol/kg sulforaphane or 295 µmol/kg erucin (n=12/group). Tumor growth rate was monitored and final tumor weights, cell proliferation (Ki67) and apoptosis (PARP) were compared. Bioavailability of isothiocyanates was determined by extracting plasma, bladder and tumor metabolites and quantifying them by UPLC-MS/MS. The potential of SFN and ECN to act as histone deacetylase inhibitors (HDACI) was assessed by HDAC activity assay, HAT assay, biomarkers of HDACI: AcH3, p21 and TS (western blotting), and by LCMS. We found broccoli sprouts have higher levels of glucosinolates and isothiocyanates than broccoli. Broccoli sprout ITCs and SFN followed by ECN were the most potent inhibitors of bladder cancer cell viability and normal bladder urothelial cells were least sensitive to this inhibition. SFN and ECN resulted in dose-dependent induction of apoptosis and G2/M cell cycle accumulation. In vivo, a significant decrease in final tumor weight (36-60% reduction), decreased tumor cell proliferation (50%) and induction of apoptosis (2-fold) was observed in broccoli sprout and pure ITC gavage treatment groups when compared to control. Metabolites of SFN and ECN were present in plasma, tumor and bladder tissue of all treated groups (pico-micromolar range). Evidence of inter-conversion of SFN and ECN was observed. HDAC and HAT activity were significantly inhibited by both SFN and ECN treatment, AcH3 and p21 were increased and LCMS revealed decrease in phosphorylated histone H1, a plausible novel biomarker of bladder cancer progression. Our studies show that broccoli sprout isothiocyanates are abundant, bioavailable and have significant bioactivity in inhibiting bladder cancer in vitro and in vivo in association with epigenetic modulation. This work supports further preclinical and clinical studies to determine the potential chemopreventive and therapeutic effects of these compounds in bladder cancer.
Steven Clinton, MD, PhD (Advisor)
Tatiana Oberyszyn, PhD (Committee Member)
Steven Schwartz, PhD (Committee Member)
Roger Briesewitz, PhD (Committee Member)
Biomedical Research
Broccoli; Bladder Cancer; Isothiocyanates; Proteomics; Metabolism; Transitional Cell Carcinoma

Recommended Citations

Citations

  • Abbaoui, B. (2011). BROCCOLI ISOTHIOCYANATES AS CHEMOPREVENTIVE AGENTS AND EPIGENETIC MODULATORS OF BLADDER CANCER [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1312835540

    APA Style (7th edition)

  • Abbaoui, Besma. BROCCOLI ISOTHIOCYANATES AS CHEMOPREVENTIVE AGENTS AND EPIGENETIC MODULATORS OF BLADDER CANCER. 2011. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1312835540.

    MLA Style (8th edition)

  • Abbaoui, Besma. "BROCCOLI ISOTHIOCYANATES AS CHEMOPREVENTIVE AGENTS AND EPIGENETIC MODULATORS OF BLADDER CANCER." Doctoral dissertation, Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1312835540

    Chicago Manual of Style (17th edition)

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© 2011, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.