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Indole-3-carbinol (I3C), a dietary compound found naturally in cruciferous vegetables of the Brassica genus, has been shown to cause a G1 growth arrest in many types of tumor cells, including both androgen-dependent and androgen-independent prostate cancer cell lines. While the mechanism through which I3C caused this arrest remained unclear, previous research had shown that I3C affected several components of the cell cycle, causing decreased cdk2 activity as well as increased p21waf1/cip1 (p21) protein levels in LNCaP prostate carcinoma cells. p21, a critical tumor suppressor gene, regulates cell proliferation by inhibiting the activity of cdk/cyclin complexes, thereby halting cell growth. The objective of the current study was to characterize the mechanism of p21 induction in LNCaP cells by I3C in order to determine the method through which I3C led to growth arrest. Western blotting and quantitative RT-PCR determined that I3C caused an increase in both p21 mRNA transcript and protein levels. Further studies using luciferase-reporter assays and site-directed mutagenesis showed that I3C induced p21 trascriptionally through a p53 binding site. An oligonucleotide precipitation experiment revealed that I3C increased the level of activated p53 nuclear protein able to bind its target site on the p21 promoter. Through the use of phospho-specific antibodies in western blotting experiments, I3C also demonstrated the ability to increase the levels of several phosphorylated forms of p53 that contribute to p53 protein stability and greater transactivation potential. These results conclude that one mechanism of I3C induced G1 arrest involved initially up-regulating the expression and activity of p53 through the activation of specific phosphorylation sites, thereby allowing p53 to bind the p21 promoter to induce the expression of p21. By uncovering an important mechanism of I3C action, this study strives to further the possibility of developing I3C as an anti-cancer therapeutic.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]