Previous studies including those from our laboratory have shown that cruciferous vegetable constituent phenethyl isothiocyanate (PEITC) suppresses growth of human prostate cancer cells in culture and in vivo but the mechanism of its anticancer effect is not fully understood. We now demonstrate, for the first time, that PEITC treatment causes Atg5-dependent autophagic and apoptotic cell death in prostate cancer cells regardless of their androgen-responsiveness or p53 status. Exposure of PC-3 (androgen-independent, p53 null) and LNCaP cells (androgen-responsive, wild type p53), but not a normal human prostate epithelial cell line (PrEC), to PEITC resulted in several specific features characteristic of autophagy including appearance of membranous vacuoles (revealed by transmission electron microscopy), formation of acidic vesicular organelles (revealed by acridine orange staining), and processing and recruitment of microtubule associated protein 1 light chain 3 (LC3) to autophagosomes (revealed by immunoblotting and immunohistochemistry). Autophagy inhibitors 3-methyl adenine and chloroquine conferred partial but significant protection against PEITC-mediated growth inhibition (trypan blue dye exclusion assay) and apoptosis (cytoplasmic histone-associated DNA fragmentation and cleavage of procaspae-3). These results indicated that PEITC-induced autophagy and apoptosis were interrelated. The PEITC treatment suppressed activating phosphorylations of Akt and mTOR, which are implicated in regulation of autophagy by different stimuli. However, the PEITC-mediated processing and recruitment of LC3 to autophagosomes was only marginally reduced by ectopic expression of constitutively active Akt or mTOR regulator Rheb. On the other hand, knockdown of Atg5 protein significantly protected against PEITC-mediated autophagy as well as apoptotic DNA fragmentation and cleavage of procaspase-3. Xenograft model using PC-3 cells and C. elegans expressing lgg-1:GFP fusion protein provided in vivo evidence for PEITC-induced autophagy. In conclusion, the present study indicates that Atg5 plays a critical role in regulation of PEITC-mediated autophagy and apoptosis in human prostate cancer cells. This study was supported by the US PHS grant CA101753, awarded by the National Cancer Institute.
Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 4787.
100th AACR Annual Meeting-- Apr 18-22, 2009; Denver, CO