Phenethyl isothiocyanate mediates autophagy via the AMPK/mTOR signal pathway in human glioblastoma multiforme cells.

Phenethyl isothiocyanate (PEITC), isolated from cruciferous vegetables, exhibits anticancer activity against various human cancer cells. Our previous studies demonstrated that PEITC suppresses the growth of glioblastoma multiforme (GBM) 8401 cells in vitro and in vivo. In this study, we investigated the effects of PEITC on autophagy and its underlying molecular mechanisms in GBM 8401 cells. PEITC altered the morphology of GBM 8401 cells and triggered acidic vesicular organelle and autophagosome formation, as confirmed by acridine orange (AO) and monodansylcadaverine (MDC) staining, respectively. PEITC-treated cells transfected with green fluorescent protein (GFP)-microtubule-associated protein 1A/1B-light chain 3 (LC3) exhibited punctate autophagosomes, and autophagosome formation was further confirmed via transmission electron microscopy (TEM). Treatment with chloroquine (CQ) inhibited autophagy, resulting in a decrease in the number of autophagosome puncta. CQ also enhanced PEITC-induced cytotoxicity in GBM 8401 cells. Furthermore, the expression of autophagy-associated proteins including GβL, mTOR, mTORSer2481, Raptor, Rictor, and, Beclin 1 was decreased, whereas the expression of AMPKαThr172, AMPKα1Ser485/AMPKα2Ser491, PI3K class III, VPS34, autophagy protein 5 (Atg5), Atg7, Atg12-Atg5, Atg16L1, Atg3, sequestosome 1 (SQSTM1/p62), and LC3 II was increased. Confocal laser scanning microscopy further revealed decreased Beclin 1 and increased LC3 levels following PEITC treatment. After CQ pre-treatment, the expression levels of SQSTM1/p62 and LC3 II were elevated. PEITC induces protective autophagy via the adenosine monophosphate-activated protein kinase/mammalian target of rapamycin (AMPK/mTOR) signaling pathway in GBM 8401 cells. Elucidation of the molecular mechanism of PEITC in GBM 8401 cells in vitro may provide a potential therapeutic approach for glioblastoma in the future.