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Tetrastigma hemsleyanum (T. hemsleyanum) has been regarded as an anticancer food in China. However, its corresponding mechanisms remains unclear. Thus, in this study, the antitumor activity of flavones-rich fraction of root of T. hemsleyanum (FRTH) was investigated in vitro and in vivo. The results indicated that FRTH could inhibit the proliferation and migration of human hepatoellular carcinomas (HepG2) cells in vitro by the phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) pathway. FRTH could increase the level of reactive oxygen species and change the mitochondrial membrane potential in HepG2 cells. In addition, FRTH treatment (300 mg/kg and 600 mg/kg, body weight) significantly suppressed tumor growth on HepG2 tumor-bearing nude mice. Besides, immunohistochemistry assays and western blotting revealed that FRTH enhanced the expression level of Bcl-2 associated X protein/B-cell lymphoma-2 (Bax/Bcl-2), cytochrome C, caspase-3, caspase-9, and cleaved-caspase-3, and downregulated the expression level of platelet endothelial cell adhesion molecule-1 (CD31), ki67, and vascular endothelial growth factor (VEGF) factor in HepG2 tumor-bearing mice. Our study suggests T. hemsleyanum as a kind of promising candidate medicine for liver cancer treatment. [ABSTRACT FROM AUTHOR]