Antitumor effect of cytosine deaminase/5-fluorocytosine suicide gene therapy system mediated by Bifidobacterium infantis on melanoma
Antitumor effect of cytosine deaminase/5-fluorocytosine suicide gene therapy system mediated by Bifidobacterium infantis on melanoma
复制标题
婴儿双歧杆菌介导的胞嘧啶脱氨酶/5-氟胞嘧啶自杀基因治疗系统对黑色素瘤的抗肿瘤作用
DOI：
10.1111/j.1745-7254.2005.00094.x
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发表时间：
2005-05-01
影响因子：
8.2
通讯作者：
Wang, SR
中科院分区：
文献类型：
--
作者：
Yi, C;Huang, Y;Wang, SR
目的：构建双歧杆菌介导的CD/5-氟胞嘧啶（CD/5-FC）靶向基因治疗系统，观察CD/5-FC自杀基因治疗系统对黑色素瘤的体内外治疗效果。将CD/pGEX-1 LamdaT重组质粒转染双歧杆菌，与5-FC厌氧孵育。收集上清液，加入黑色素瘤B16-F10细胞中，观察对B16-F10细胞的杀伤作用。小鼠接种黑色素瘤B16-F10细胞建立动物模型。结果：从6.2kb的重组质粒中提取出两个大小约为4.9kb和1.3kb的片段，分别与pGEX-1 LamdaT质粒和CD基因大小相当。测序结果表明，所提取的重组质粒中插入的基因的全长和核苷酸序列与CD基因完全一致。在体外培养条件下，上清处理的B16-F10细胞形态明显受损，细胞生长受到明显抑制。结论：外源CD基因已正确插入到pGEX-1 LambdaT质粒中，并转入到双歧杆菌Bifidobacteriums中。婴儿双歧杆菌介导的CD/5-FC自杀基因治疗系统在体外和体内对黑色素瘤均表现出良好的抗肿瘤作用。
AbstractAim:To construct a Bifidobacterium infantis/CD targeting gene therapy system and observe the antitumor effect of cytosine deaminase/5-fluorocytosine (CD/5-FC) suicide gene therapy system mediated by Bifidobacterium infantis on melanoma in vitro and in vivo.Methods:A recombinant CD/pGEX-1LamdaT plasmid was transfected into Bifidobacterium infantis by electroporation. Bifidobacterium infantis transfected by recombinant CD/pGEX-1LamdaT plasmid was incubated with 5-FC anaerobically. Then the supernatant fluid was collected and added to melanoma B16-F10 cells to observe the killing effect for B16-F10 cells. Mice were inoculated with melanoma B16-F10 cells to establish animal models. The mice were then injected with 5-FC and Bifidobacterium infantis transfected by recombinant CD/pGEX-1LamdaT plasmid.Results:Two segments of approximate 4.9 kb and 1.3 kb were extracted from the 6.2 kb recombinant plasmid, which were equal to the size of the pGEX-1LamdaT plasmid and CD gene, respectively. Sequencing results showed that the full length and sequence of nucleotide acid of the inserted gene in extracted recombinant plasmid was completely identical to the CD gene. In vitro, B16-F10 cells treated by supernatant fluid were remarkably damaged morphologically, and the cell growth was significantly inhibited. Experiments on the mice melanoma model showed that after treatment with a combination of transfected Bifidobacterium infantis and 5-FC, the tumor volume was significantly inhibited compared with controls.Conclusion:The foreign gene, CD gene, was correctly inserted into pGEX-1LambdaT plasmid and transferred into Bifidobacterium infantis. CD/5-FC suicide gene therapy system mediated by Bifidobacterium infantis demonstrated a good antitumor effect on melanoma in vitro and in vivo.