Abstract: Objective: To investigate the in vitro effect of pirarubicin (THP) in combination with hyperthermia on gastric cancer cell and discuss the acting mechanism. Methods: Simulation model was established in vitro with gastric cancer cell line MGC-803. Cell viability and growth was measured by means of MTS-PMS assay. Cell-doubling time was calculated. Cell cycle were analyzed by flow cytometry(FCM) and apoptosis status was analyzed using Annexin V-FITC kit. Results: Strong synergistic effect was observed after cancer cell line MGC-803 was treated with THP in combination with elevated hyperthermia between 39℃ and 45℃, which is superior to cisplatin. Dose-effect and time-effect relationship were also observed when treated with elevated THP concentrations and prolonged treating time. Slowed cell growth and delayed cell doubling time were evident after treated with THP in combination with hyperthermia (p<0.01). Furthermore, studies on cell cycle suggested that the cells at all cycle stages could be strongly affected after treatment with THP in combination with hyperthermia, revealing significantly increased S and decreased G2/M stage fraction (p<0.05). Cell death mainly consisted of necrosis and apoptosis. Enhanced apoptosis status treated with combination method was positively correlated with elevated temperature, which was significantly higher compared to group of simple thermotherapy or of medication alone (p<0.01). Conclusions: Synergistic effects of THP in combination with hyperthermia is evident. Cytotoxicity of THP has been considerably enhanced by mild hyperthermia on gastric cancer cells. THP may be a potential therapeutic drug for intra-peritoneal chemohyperthermia.