Human prostate cancer is second cause of male death every year in the western countries. Prostate cancer incidence rate is increased in eastern countries such as Japan, Singapore and Korea. Recently, much attention has been paid on chemotherapeutic compounds from natural product as anti-cancer agents. In the present study, marine-derived fungus Aspergillus fumigatus was cultivated and isolated three secondary metabolites (Cyclotryprostatin A (C1), Tryprostatin B (C2), and Demethoxyfumitremorgin (C3)) from marine fungus strain-150 (MFS-150) culture broth extract. The structures of compounds were elucidated via Low resolution electron ionization mass spectrometer (LREIMS), 1D, and 2D nuclear magnetic resonance (NMR) spectra. To investigate the effect of secondary metabolites in inhibition of cell proliferation and induction apoptosis on prostate cancer cells, PC3 cells were treated with different concentrations (25, 50, 100 μM) and various time intervals (24, 48, 72h) of Cyclotryprostatin A (C1), Tryprostatin B (C2), and Demethoxyfumitremorgin (C3). Among them, Demethoxyfumitremorgin (C3) compound has shown anti-proliferative effect on PC3 cells whereas other compounds did not shows significant cytotoxicity. Furthermore, we tested the cell proliferation and apoptosis activity of Demethoxyfumitremorgin C (C3) in PC3 cells and results shows that significant inhibition on cell proliferation in dose and time dependent manner. The level of cell cycle arrest relative protein expression, phospho-cdc2, cyclin A, D, E, CDK 2, and 4 were decreased by Demethoxyfumitremorgin C (C3) in a dose-dependent manner. In addition, Demethoxyfumitremorgin C (C3) regulated the anti- and pro-apoptotic proteins like Bcl-2, Bcl-xL and Bax. Also, relative proteins for apoptosis, cleaved PARP, p-p53, pp21, caspase-3, -8, and -9 were increased by Demethoxyfumitremorgin C (C3). Taken together, these results implicated that Demethoxyfumitremorgin C (C3) has anticancer effect on prostate cancer PC3 cells via suppress cell proliferation through cell cycle arrest at G1 phase and induction of apoptosis via regulating pro- and anti-apoptotic protein expressions in caspase-dependent pathway. Thus, Demethoxyfumitremorgin C (C3) could be used for therapeutic agent for treatment of prostate cancer.