Mobile website
The Construction and Biological Effects of Double Targeting Tissue-Specific HSV-tk/GCV Anti-Tumor System in vitro
- Article
- Figures
- Metrics
- Preview PDF
- Reference
- Related
- Cited by
- Materials
Abstract:
Objective: To construct the potent targeting gene delivery and expression system, and to investigate the special killing effect of HSV-tk/GCV system on human liver cancer cells in vitro mediated by it. Methods: The anti-TfR ScFv-GAL4 fusion protein expression vector ScFv-GAL4-pET28a and the eukaryocyte expression plasmid pEBAF/tk-GAL4rec were constructed by recombinant DNA technology. After the induction by IPTG, we got the anti-TfR ScFv-GAL4 fusion protein as delivery vector to transfect pEBAF/tk-GAL4rec into the human liver cancer cells line HepG2,SMMC7721 and lung cancer cells line A549 which all over-express TfR via receptor-mediated endocytosis. The positive cell clones were selected by hygromycin and were named HepG2/tk,SMMC7721/tk and A549/tk respectively. Then MTT method was used to determine the killing effect of GCV on them. Results: The constructions of the ScFv-Gal4 fusion protein and the recombination expression plasmid pEBAF/tk-GAL4rec were confirmed by double enzyme digestion, SDS-PAGE and sequencing. In the experiment of cytotoxicity effect, the HepG2/tk cells that over-secrete AFP (845 ng/ml) were highly sensitive to the toxicity of GCV, whereas the SMMC7721/tk cells that under-secrete AFP (2 ng/ml) were slightly sensitive to GCV, and the A549/tk cells that don t secrete AFP were not.Conclusions:The double-targeting and tissue-specific HSV-tk/GCV anti-tumor system has been constructed successfully, and it displays a good targeting to tumor cells.
Keywords:
Project Supported:
Clc Number:
