Abstract 3704: Diindolylmethane induces ER stress mediated autophagy in ovarian cancer cells

Abstract

Abstract Endoplasmic reticulum (ER) is responsible for biosynthesis, folding assembly and modification of various soluble and membrane proteins. ER stress is the imbalance in protein folding resulting in the accumulation of misfolded proteins in ER lumen. In our study, we observed that Diindolylmethane (DIM), a compound present in cruciferous vegetables induces ER stress in ovarian cancer cells. Treatment of SKOV-3, OVCAR-3 and TOV-21G ovarian cancer cells with varying concentrations of DIM for 24hours resulted in the upregulation of Bip, a molecular chaperone. Bip or Grp78 binds to IRE1α, ATF6 and PERK under normal condition, however, during ER stress, Bip dissociates and releases all the three proteins. DIM treatment also significantly increased the expression of IRE1α, ER oxidase (ERO1) and CHOP. CHOP translocate to nucleus and initiate apoptotic stimuli in cells undergoing ER stress. Moreover, DIM induced ER stress also lead to the activation of eIF2α by phosphorylation. Activation of eIF2α inhibits protein translation to avoid further protein accumulation in ER. The release of calcium by DIM treatment in ovarian cancer cells was evaluated by Accuri C6 flowcytometer. The advantage with Accuri C6 is that calcium levels can be monitored without interruption. DIM treatment increased the cytosolic calcium levels in SKOV3 cells. Increase in cytosolic calcium is known to cause ER stress. In addition, our flowcytometry results indicate that DIM induces autophagy in all the ovarian cancer cells in a concentration dependent manner. Exposure of SKOV-3, OVCAR-3 and TOV-21G cells to DIM upregulated the expression of LC3B, a potent indicator of autophagy. DIM induced autophagy was blocked by bafilomycin, an autophagy inhibitor. Treatment of cells with mithramycin (ER stress inhibitor) blocked ER stress proteins activated by DIM. Interestingly, we also observed that blocking DIM-induced ER stress by mithramycin blocked autophagy in ovarian cancer cells. Taken together these results suggest that induction of autophagy in ovarian cancer cells was associated with ER stress caused by DIM. Further mechanistic studies are in progress. [Supported in part by R01 grants CA106953 and CA129038 (to S.K.S) awarded by the National Cancer Institute]. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3704. doi:10.1158/1538-7445.AM2011-3704