Tumor-directed gene therapy in mice using a composite nonviral gene delivery system consisting of the piggyBac transposon and polyethylenimine.
BACKGROUND
Compared with viral vectors, nonviral vectors are less immunogenic, more stable, safer and easier to replication for application in cancer gene therapy. However, nonviral gene delivery system has not been extensively used because of the low transfection efficiency and the short transgene expression, especially in vivo. It is desirable to develop a nonviral gene delivery system that can support stable genomic integration and persistent gene expression in vivo. Here, we used a composite nonviral gene delivery system consisting of the piggyBac (PB) transposon and polyethylenimine (PEI) for long-term transgene expression in mouse ovarian tumors.


METHODS
A recombinant plasmid PB [Act-RFP, HSV-tk] encoding both the herpes simplex thymidine kinase (HSV-tk) and the monomeric red fluorescent protein (mRFP1) under PB transposon elements was constructed. This plasmid and the PBase plasmid were injected into ovarian cancer tumor xenografts in mice by in vivo PEI system. The antitumor effects of HSV-tk/ganciclovir (GCV) system were observed after intraperitoneal injection of GCV. Histological analysis and TUNEL assay were performed on the cryostat sections of the tumor tissue.


RESULTS
Plasmid construction was confirmed by PCR analysis combined with restrictive enzyme digestion. mRFP1 expression could be visualized three weeks after the last transfection of pPB/TK under fluorescence microscopy. After GCV admission, the tumor volume of PB/TK group was significantly reduced and the tumor inhibitory rate was 81.96% contrasted against the 43.07% in the TK group. Histological analysis showed that there were extensive necrosis and lymphocytes infiltration in the tumor tissue of the PB/TK group but limited in the tissue of control group. TUNEL assays suggested that the transfected cells were undergoing apoptosis after GCV admission in vivo.


CONCLUSIONS
Our results show that the nonviral gene delivery system coupling PB transposon with PEI can be used as an efficient tool for gene therapy in ovarian cancer.
Kang
Yu
Y
Obstetrics and Gynecology Hospital, Fudan University, Shanghai 200011, PR China. yukang@fudan.edu.cn
Zhang
Xiaoyan
X
Jiang
Wei
W
Wu
Chaoqun
C
Chen
Chunmei
C
Zheng
Yufang
Y
Gu
Jianren
J
Xu
Congjian
C
D000818:Animals; D045744:Cell Line, Tumor; D004251:DNA Transposable Elements; D005260:Female; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D010051:Ovarian Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000628:therapy*; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011094:Polyethyleneimine / Q000378:metabolism*; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays
D004251:DNA Transposable Elements; D011094:Polyethyleneimine; D013937:Thymidine Kinase