This study investigated the effect of periplocin (PP) on prostate cancer (PCa) cells and explored the molecular mechanisms.CCK8 assay,colony formation and 3D matrix gel invasion assay were applied on in vitro cultured DU145 and PC3 cells to evaluate the effect of PP on cell growth,colony formation,migration and invasion of PCa cells;Flow cytometry,Hoechst 33258 staining and Western blot were used to measure the ability of PP induced cell apoptosis,apoptotic body formation and apoptotic proteins expression;RNA-Seq was carried out to figure out the PP-induced transcriptome expression regulation in PCa cells;DCFH-DA fluorescent probe was used to detect the effect of PP on intracellular reactive oxygen species (ROS) level;Western blot and cells expressing EGFP-LC3B fusion protein were used to detect the effect of PP on autophagy related proteins expression and autophagosome formation;A nude mouse tumor bearing model was established to detect the inhibitory effect of PP on the in vivo growth of PCa cells.The results showed that PP could inhibit the proliferation,colony formation,migration and invasion of PCa cells in a concentration dependent manner;PP could induce apoptosis,apoptotic body formation and apoptosis-related protein expression in PCa cells.The results of RNA-Seq indicated that PP treatment could alter gene expression profile of PCa cells,and the differential expression genes were mainly enriched in autophagy related signaling pathways.PP could increase the cellular ROS of PCa cells in a concentration dependent manner,up-regulate the expression of autophagy marker proteins LC3B and Beclin-1,and down-regulate the expression of autophagy degradation marker protein p62.PP could inhibit the growth of prostate cancer cells in vivo.This study suggests that PP inhibited the proliferation of PCa cells and induced apoptosis.The molecular mechanism may be that PP induced PCa cells autophagy and eventually led to cell death.