Background: The cytotoxic activity of sea cucumber glycosides against different types of cells and cell lines, including human tumor cell lines, has been studied for many years. However, the molecular mechanism(s) of the antitumor action of triterpene glycosides on cancer cells remain unclear. This article reports a continuation of investigations of triterpene glycoside cucumarioside A₂-2 isolated from the Far-Eastern sea cucumber Cucumaria japonica. It describes a study of glycoside anticancer activity in vivo and glycoside interaction with mouse Ehrlich carcinoma cells in vitro. Methods: The cytotoxicity of cucumarioside A₂-2 and its effect on apoptosis, the cell cycle, DNA biosynthesis and p53 activity, and glycoside anticancer action against Ehrlich carcinoma cells were studied. Results: Cucumarioside A₂-2 influences tumor cell viability at micromolar concentrations. The EC₅₀ for glycoside estimated by nonspecific esterase assay and MTT assay was 2.1 and 2.7 μM, respectively. Cucumarioside A₂-2 at a subcytotoxic range of concentrations exhibits a cytostatic effect by blocking cell proliferation and DNA biosynthesis in the S phase. It may induce apoptosis in tumor cells in a caspase-dependent way, bypassing the activation of the p53-dependent segment. Conclusion: The anticancer and proapoptotic properties of cucumarioside A₂-2 may be due to direct interaction of the glycoside with tumor cells. The in vivo anticancer effect of cucumarioside A₂-2 may be associated with the ability of the drug to arrest the cell cycle in the synthetic phase and induce programmed tumor cell death.