PMID: 16511352Mar 3, 2006Paper

Intracellular localization and sustained prodrug cell killing activity of TAT-HSVTK fusion protein in hepatocelullar carcinoma cells

Molecules and Cells
Limin CaoGuanxin Shen

Abstract

Gene therapy with nonviral vectors using the suicide gene/prodrug activating system of herpes simplex vi-rus type-1 thymidine kinase (HSV1-TK)/ganciclovir (GCV) is inefficient in killing malignant tumor cells due to two major factors: (a) an unsatisfactory by-stander effect; (b) short-lived expression of the pro-tein. 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 nu-cleus. The transduced HepG2 cells are killed by exo-genously added GCV and have bystander effects on untransduced HepG2 cells. Most importantly, the in-troduced 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 nu-clear transcription machinery. Our results indicate that TAT fusion proteins traffic intercellularly and have enhanced stability and prodrug cell killing activ-ity. We conclude that TAT has potential for enhancing enzym...Continue Reading

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