Abstract 612: Fisetin, a dietary flavonoid and novel mTOR inhibitor for treatment and prevention of prostate cancer

The development and progression of prostate cancer involves deregulation of several signaling pathways including the mammalian target of rapamycin (mTOR). The mTOR kinase forms an important component of PTEN/PI3K/Akt signaling pathway. Mutations in the tumor suppressor gene PTEN are observed at high frequency during the development of prostate cancer and often accompanied by activation of AKT and mTOR pathways. We initiated studies to identify polyphenols from dietary sources that could interact with the mTOR receptor and in silico studies showed that fisetin physically interacts with mTOR. Fisetin is a tetrahydroxyflavone, found in strawberry, apple, persimmon, kiwi, onion and cucumber where it serves as a coloring agent. Blind docking of fisetin to the mTOR target was performed with Autodock4 using the NMR structure 2NPU from the Protein Data Bank. Results placed fisetin in two clustered sites located between and on either side of the 4 helices. The binding energies were in the -7 to -8 Kcal/mol range for the binding constant. The binding on one site included hydrogen bonding to a glutamate by two hydroxyl groups. The second site was mostly hydrophobic, with the ring of fisetin stacking on rings from the peptide. We used a unique family of human prostate epithelial cell lines, derived from RWPE-1 by exposure to N-methyl-N-nitrosourea (MNU) to study the effect of fisetin on mTOR signaling. The MNU cell lines, in order of increasing malignancy and phenotype are; WPE1-NA22, WPE1-NB14, WPE1-NB11, and WPE1-NB26 and mimic multiple steps in the process of prostate carcinogenesis. We observed that transformed cells exhibited increase in the activity and phosphorylation of mTOR, PI3K, Akt, Erk and p-S6 that was associated with gradual loss of PTEN and decrease in the levels of p-AMPK, a negative regulator of mTOR. We treated cells with fisetin (0-80 μM) for 24-72 hours and observed that transformed cells exhibited greater sensitivity to fisetin induced cell death. As cells transform from a normal phenotype to a malignant phenotype they acquire higher mTOR expression and become more susceptible to fisetin induced cell death. In presence of fisetin, cells expressing higher mTOR levels formed less number of colonies that were also smaller in size. Fisetin treatment resulted in inhibition of PI3K, p-Akt and p-Erk. Treatment of cells with fisetin, downregulated components of mTOR including Raptor, Rictor, PRAS40 and GμL leading to loss of mTOR complexes 1/2. Fisetin also activated the mTOR repressor TSC2 through inhibition of Akt, activation of AMPK and PTEN. It is noteworthy that mTOR and Erk/MAPK signaling pathways function cooperatively to promote development of advanced prostate cancer. Our observations suggest that fisetin, through its ability to inhibit mTOR signaling, could be developed as a novel non-toxic agent for slowing the progression of the disease and as an adjuvant for the management of advanced prostate cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 612. doi:1538-7445.AM2012-612