Background:The aims of this study were to investigate the anticancer activity of anthraquinone isolated from a soil derived filamentous bacterium Streptomyces sp. isolate ERINLG-26 and to explore the molecular mechanisms of action. Methods: Isolation of Streptomyces sp. (ERINLG-26) was performed by serial dilution using dilution plate technique. Ethyl acetate extract was taken from Streptomyces sp. isolate ERINLG-26 in MNGA medium. Anticancer properties of anthraquinoneweretested from ethyl acetate extract. Anthraquinone was also tested against COLO320 colorectal adenocarcinoma cell line using the 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Intracellular visualization was done by using a laser scanning confocal microscope. Apoptosis was measured using DNA fragmentation technique. Activation of p53 and caspasedependant pathway was evaluated in RT-PCR and western blotting analysis. Molecular- dockings were performed to investigate the binding modes of anthraquinone into p53 and caspase-3 active sites. Results: The ethyl acetate extract was subjected to fractionation by column chromatography over silica gel. The isolated compound anthraquinone showed prominent cytotoxic activity in vitro against COLO320 colorectal adenocarcinoma cell line. It showed 79.73% activity at the dose of 300 μg/mL with IC 50 value of 75 μg/mL.Treatment of the COLO320 cancer cells with isolated anthraquinone significantly reduced cell proliferation, increased formation of fragmented DNA and apoptotic body. The expression of p53 and caspase-3 were up-regulated by anthraquinone in COLO320 colorectal adenocarcinoma cell line. The molecular docking analysis revealed good activity with the ligand anthraquinone with p53 and casepase-3 targets at low energy. Conclusion: These results strongly suggest that the isolated anthraquinone induces apoptosis in COLO320 cancer cells via caspase activation and the results might provide helpful suggestions for the design of anti-tumor drugs toward colon cancer treatment.