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Intracellular Localization and Sustained Prodrug Cell Killing Activity of TAT-HSVTK Fusion Protein in Hepatocelullar Carcinoma Cells  

Cao, Limin (Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology)
Si, Jin (The Key Laboratory of Molecular Biology, Jiangsu Institute of Parasitic Diseases)
Wang, Weiyu (Department of General Surgery, Tongji Hospital)
Zhao, Xiaorong (Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology)
Yuan, Xiaomei (Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology)
Zhu, Huifen (Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology)
Wu, Xiaolong (Department of General Surgery, The Third Affiliated Hospital of NanTong University)
Zhu, Jianzhong (Department of General Surgery, The Third Affiliated Hospital of NanTong University)
Shen, Guanxin (Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology)
Publication Information
Molecules and Cells / v.21, no.1, 2006 , pp. 104-111 More about this Journal
Abstract
Gene therapy with nonviral vectors using the suicide gene/prodrug activating system of herpes simplex virus type-1 thymidine kinase (HSV1-TK)/ganciclovir (GCV) is inefficient in killing malignant tumor cells due to two major factors: (a) an unsatisfactory bystander effect; (b) short-lived expression of the protein. To study the capacity of the protein transduction domain (PTD) of HIV-1 TAT protein to enhance HSV1-TK/GCV cancer gene therapy, we constructed three fusion proteins TAT-TK, TK-TAT and TK. TAT-TK retained as much enzyme activity as TK, whereas that of TK-TAT was much lower. TAT-TK can enter HepG2 cells and much of it is translocated to the nucleus. The transduced HepG2 cells are killed by exogenously added GCV and have bystander effects on untransduced HepG2 cells. Most importantly, the introduced recombinant protein is stable and remains functional for several days at least, probably because nuclear localization protects it from the cytoplasmic degradation machinery and provides access to the nuclear transcription machinery. Our results indicate that TAT fusion proteins traffic intercellularly and have enhanced stability and prodrug cell killing activity. We conclude that TAT has potential for enhancing enzyme prodrug treatment of liver cancers.
Keywords
Bystander Effect; Cancer Gene Therapy; Ganciclovir; HIV-1 TAT; Intercellular Translocation; Protein Transduction; Thymidine Kinase;
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