AbstractGastric cancer (GC) is one of the most common digestive malignancies globally, and the prognosis of patients with advanced tumors remains poor. Isocryptotanshinone (ICTS), isolated from Salvia miltiorrhiza, was found to inhibit the proliferation of lung and breast cancer cells. However, whether ICTS has anticancer activities against GC is unknown. In the present study, we reported that the proliferation of GC cells was inhibited by ICTS in a dose- and time-dependent manner. After treatment with ICTS, GC cells were arrested in the G1/G0 phase of cell cycle and the apoptotic cells were induced in a dose-dependent manner. Additionally, ICTS suppressed the expression of cell cycle- and apoptosis-associated proteins (e.g., Cyclin D1, phosphorylated Rb, E2F1, Mcl-1, Bcl-2, and Survivin). ICTS inhibited the phosphorylation of STAT3 in a dose-dependent manner. Down-regulated STAT3 attenuated the expression of Cyclin D1, p-Rb, and Survivin, which remarkably increased the sensitivity of ICTS in GC cells; overexpression of STAT3 restored the cell growth and proliferation and the protein expression suppressed by ICTS. ICTS also suppressed the xenograft tumor growth in BALB/c nude mice. Together, these data indicate that ICTS inhibits GC proliferation by inducing G1/G0 cell cycle arrest and apoptosis via inhibiting the STAT3 signaling pathway.