Objective To explore the effects of trifluoperazine (TFP) on the growth and proliferation of Ishikawa cells of human endometrial carcinoma in vitro and their possible mechanisms. Methods The inhibitory effect of TFP of different concentrations on the growth of Ishikawa cells of human endometrial carcinoma was detected by the MTT assay. The half maximal inhibitory concentration (IC50) of the drug was calculated and the optimal time was selected. The effect of TFP on the cell cycle of Ishikawa cells was measured by the flow cytometer. The regulatory effect of TFP on the expressions of forkhead box transcription factor O1 (FOXO1) and Kruppellike factor 6 (KLF6) in Ishikawa cells was determined by the Real-Time PCR. Results TFP of different concentrations would inhibit the growth of Ishikawa cells. The inhibitory effect depended on the dose and time. The IC50 value of TFP was 16.56 μmol/L. The optimal time was 24 h for being treated by TFP of 20 μmol/L. After Ishikawa cells were treated by TFP of 20 μmol/L for 24 h, the proportion of S phase cells was significantly decreased (P<0.01) and the expression of KLF6 mRNA was significantly up-regulated (P<0.05). Conclusion The intervention of TFP in vitro can inhibit the growth and proliferation of Ishikawa cells of endometrial carcinoma. The mechanism may be relevant to up-regulate the expression of the tumor suppressor gene KLF6.