Abstract: Abstract:Objective To determine whether HSV1-tk/GCV combining with temozolomide (TMZ) would enhance antitumor effect in malignant glioma in vitro. Methods Human U251 glioblastoma cells were transfected with the recombinant retrovirus which contained HSV1-tk gene.After Screening and identifying the positive ones， the transfected and non-transfected cells were mixed at a 1∶9 ratio.The cells were incubated in the presence of GCV (2,5,10,20,40μM), both GCV(2,5,10,20,40μM) and 50μM TMZ, or non-drug for 72h respectively.Cell viability was accessed by MTT.Cell cycle progression and apoptosis ratio was determined by using flow cytometry. Results The ratio of the OD570 was significant different between the GCV group (≥5μM),GCV+TMZ group(≥2μM) and untreated control group respectively (P<0.01).The survival curve of GCV+TMZ group was significantly shifted to the left compare with the GCV group. The GCV IC50 of two groups were 17.3μM (GCV group) and 8.1μM (GCV+TMZ group) respectively (P<0.01).The flow cytometry showed that apoptosis of two groups increased significantly (P<0.01) and cells were arrested in G2~M phases. Conclusion HSV1- tk /GCV suicide gene therapy system has a powerful antitumor effect and a bystander effect for glioma in vitro.It has a synergistic anticancer effect when combined with TMZ,And GCV can induct the cells apoptosis.