Abstract 3786: Activation of KLF4 by 3,3′-Diindolylmethane contributes to the cell differentiation and grow inhibition of pancreatic cancer cells

Abstract

Abstract 3,3′-Diindolylmethane (DIM), the major bioactive metabolite of the phytochemical indole-3-carbinol present in cruciferous vegetables, is a promising cancer preventative and anti-tumor agent for various tumor types, including pancreatic cancer. However the underlying molecular mechanisms by which DIM exerts its antitumor activity remain poorly understood. Our previous studies indicate that KLF4 has a tumor suppressive function in pancreatic cancer by up-regulation of p27 expression and induction of cell cycle arrest, which has also been suggested to be the mechanistic action of DIM in pancreatic cancer cells. However, whether KLF4 is involved in DIM's antitumor activity and if so, what are the underlying molecular mechanisms in pancreatic cancer have not been defined. In the present study, we found that DIM treatment dose dependently induced the expressions of KLF4 and its downstream target gene p27 and p21 in several, and cytokeratin 19 in Panc-28 and PANC-1 pancreatic cancer cells. Consistently, induction of KLF4 expression correlated with significant inhibition of cell cycle progression and cell growth as determined by Flow cytometry and MTT assay, while knockdown or conditional knockout of KLF4 expression significantly reduced the sensitivity of pancreatic cancer cells to DIM treatment determined by IC50 assay. These results indicate that KLF4 plays a critical role in mediating the antitumor activity of DIM in pancreatic cancer cells. To further investigate the molecular mechanism underlying DIM's induction of KLF4 expression, KLF4 promoter deletion mutation analysis identified a DIM responsible region (from nucleotide −780 to −346) in the KLF4 promoter, within which a candidate Egr-1 cis-acting element was found by computer-based motive analysis. Functionally, significant induction of Egr-1 protein expression was observed as early as 1 hr after DIM treatment in time-course experiments, which was prior to the induced expression of KLF4 in FG cells. Consistently, forced or knocking down Egr-1 expression increased or reduced the response of BxPC3 cells to DIM treatment, which was correlated with increased or decreased KLF4 expression. In addition, chromatin immunoprecipitation assay showed that Egr-1 protein physically interacted with and DIM treatment increased the binding of Egr-1 to the DIM responsive region of KLF4 promoter in FG cells in vivo. Collectively, our results suggest that Egr-1/KLF4 signaling pathways play an important role in DIM's antitumor activity in pancreatic cancer cells. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. 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 3786.