Abstract 1897: Isothiocyanates and their metabolites in the plasma of mice fed broccoli sprouts isothiocyanate extract, broccoli sprouts, sulforaphane or erucin: A potential role in bladder cancer prevention

Background: Epidemiologic evidence suggests that cruciferous vegetables, particularly broccoli, may reduce bladder cancer risk. Our objectives were to characterize glucosinolates (GLUs) and their metabolites, isothiocyanates (ITCs), found in broccoli and broccoli sprouts, define bioactivity in bladder cancer cell lines in vitro, and to determine absorption and bioavailability in mice. Methods: Broccoli and broccoli sprouts GLUs and ITCs were characterized by HPLC-MS-MS and cyclocondensation assay. 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 (NBC) and a panel of human bladder cancer cell lines (BCC), 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. We defined absorption in control female mice and those fed diet 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; for two weeks (n=16/group). Plasma mercapturic acid pathway metabolites of sulforaphane and erucin were extracted and then quantified by UPLC-MS/MS. Results: We observed significant dose-dependent decreases in cell viability of bladder cancer cell lines by broccoli or broccoli sprouts extracts as well as by pure ITCs. Broccoli sprout ITCs and SFN (IC50 = 2.52 µM) followed by ECN (IC50 = 2.24 µM) were the most potent inhibitors. Normal bladder urothelial cells were less sensitive than cancer cells. SFN and ECN resulted in a dose-dependent induction of apoptosis and modulation of the cell cycle in BCC. Metabolites of sulforaphane and erucin were absent in control mice and present in plasma of all treated groups, with dietary and pure phytochemical treatments, resulting in similar plasma metabolite concentrations (micromolar range). N-acetyl cysteine conjugates were found at highest concentrations, followed by cysteinyl and glutathione conjugates and small amounts of detectable cysteinyl glycine conjugates and free SFN. Evidence of inter-conversion of sulforaphane and erucin was seen. Conclusion: Broccoli extracts and pure ITCs inhibit cell viability, induce apoptosis and cell cycle arrest in non-invasive and invasive human bladder cancer cell lines. In vivo, dietary and gavaged ITCs are readily absorbed and metabolized to various conjugates. The distribution to bladder cancer xenografts will also be discussed. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1897.