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- W1520396523 abstract "Top of pageAbstractINTRODUCTIONSuicide gene therapy using herpes simplex thymidine kinase gene combined to prodrug ganciclovir (GCV) is one of the most prominent approaches in cancer gene therapy. However, efficacy of the therapy is often insufficient due to the poor rate of gene transfer and due to the limited bystander effect. Recent studies have shown that a protein transduction domain from HIV-1 transactivator protein (TAT-PTD) can translocate through plasmamembrane and deliver proteins into cell. Aim of this study was to make intercellularly spreading HSV-TK protein with the aid of cell penetrating peptide TAT and thus enhance the cytotoxic effect of HSV-TK/GCV gene therapy.METHODSWe constructed a novel triple fusion protein containing TAT protein transduction domain (TAT-PTD), HSV-TK suicide gene and GFP marker gene (TAT-TK-GFP) to study effect of TAT-PTD on HSV-TK/GCV-suicide gene therapy. Sensitivity to ganciclovir was tested in different cell lines by comparing cells expressing TAT-TK-GFP to cells expressing TK-GFP. Biodistribution of TAT-TK-GFP from transfected cell to surrounding cells was analyzed by direct fluorescence microscopy or by immunofluorescence staining.RESULTSWe found that TAT-peptide enhances cytotoxic effect of HSV-TK/GCV-suicide gene therapy in rapidly dividing cells e.g. in BTC4 rat glioma cell line or in SKOV3.ip1 ovarian cancer cell line. However, enhancement was abolished after serum deprevation. Intercellular spreading of TAT-TK-GFP-protein was not visualized by direct fluorescence microscopy or immunofluorescence staining. Control fusion protein, VP22-GFP, contrast to TAT-fusion protein showed spreading from transfected cell to adjacent cells after methanol fixation.CONCLUSIONThis study shows that the triple fusion gene TAT-TK-GFP is useful tool for investigating the utility of TAT fusion proteins in suicide gene therapy. Our data with TAT-TK-GFP demonstrate that HSV-TK/GCV cytotoxicity can be enhanced in rapidly growing cancer cells, despite the fact that intercellular spreading cannot be visualized." @default.
- W1520396523 created "2016-06-24" @default.
- W1520396523 date "2004-05-01" @default.
- W1520396523 modified "2023-10-18" @default.
- W1520396523 title "HIV-1 TAT protein transduction domain-mediated enhancement of HSV-TK/ganciclovir suicide gene therapy" @default.
- W1520396523 doi "https://doi.org/10.1016/j.ymthe.2004.06.525" @default.
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