目的：探讨慢病毒介导的酵母菌胞嘧啶脱氨酶（yeast cytosine deaminase，YCD）自杀基因对人白血病Jurkat细胞的杀伤作用和旁观者效应。方法：制备YCD-GFP慢病毒上清，转染Jurkat细胞（即Jurkat/YCD-GFP细胞），流式细胞术检测YCD-GFP基因的表达。体外观察前体药物5-氟胞嘧啶（5-fluorocytosine，5-FC）对Jurkat/YCD-GFP细胞的杀伤情况和旁观者效应；建立Jurkat/YCD-GFP荷瘤小鼠模型，体内观察5-FC对移植瘤细胞的杀伤作用和旁观者效应。以上实验均以Jukut/GFP细胞为对照。结果：成功制备YCD-GFP慢病毒和感染YCD-GFP慢病毒的Jurkat/YCD-GFP细胞，YCD-GFP慢病毒对Jurkat细胞的感染率达96.93%。232 μmol/L的5-FC作用72 h可使（95.61±2.07）% 的Jurkat/YCD-GFP细胞死亡，且上清中5-FC含量下降80%左右。体外实验显示，Jurkat/YCD-GFP和Jurkat细胞效靶比1∶10混合培养\[（68.69±4.97）％ vs （97.87±111）％，P<0.05）\]和Transwell 1〖DK〗∶25分层培养\[（1.46±0.27）×105 vs （3.00±0.16）×105，P<0.01\]均存在旁观者效应。体内实验显示，YCD/5-FC自杀基因系统对Jurkat/YCD-GFP荷瘤小鼠亦具有较强的杀伤作用和旁观者效应（P<0.05）。结论：慢病毒介导的YCD/5-FC自杀基因系统对人Jurkat白血病细胞具有显著的杀伤作用和旁观者效应

Objective: To investigate the cytotoxicity and bystander effects of lentiviral vector-mediated yeast cytosine deaminase (YCD) suicide gene against the human leukemia Jurkat cells. Methods:YCD-GFP lentiviral vector was prepared and transfected into Jurkat cells (Jurkat/YCD-GFP cells). The expression of YCD-GFP gene was examined by flow cytometry. The cytotoxicity and bystander effects of pro-drug 5-fluorocytosine (5-FC) on Jurkat/YCD-GFP cells were measured in vitro. Nude mice bearing-Jurkat/YCD-GFP cell tumors were established and the cytotoxicity and bystander effects of 5-FC on tumor cells were measured in vivo. Results: YCD-GFP lentiviral vector and Jurkat/YCD-GFP cells (Jurkat cells infected by YCD-GFP lentiviral vector) were successfully prepared; the infection rate of YCD-GFP lentiviral vector on Jurkat cells was 96.93%. 5-FC (232 μmol/L for 72h) killed (95.61±2.07)% of Jurkat/YCD-GFP cells, and 5-FC level in the culture supernatant decreased by about 80%. Bystander effect of Jurkat/YCD-GFP cells could be observed both in mixed culture (effect∶target=1∶10, \[68.69± 4.97\]% vs \[97.87±1.11\]%, P<0.05) and in Transwell culture (effect∶target=1∶25, \[1.46±0.27\]×105 vs \[3.00±0.16\]×105，P<0.01) . In vivo experiment results demonstrated that YCD/5-FC suicide gene system had strong cytotoxicity and bystander effects in Jurkat/YCD-GFP tumor bearing mice (P<0.05). Conclusion: The lentiviral vector-mediated YCD/5-FC suicide gene system has marked cytotoxicity and bystander effects against human leukemia Jurkat cells.

国家自然科学基金资助项目（No. 30871100，No.30172347）；上海市科委重点项目基金资助（No.8JC1406500）