Transcellular transfer of active HSV-1 thymidine kinase mediated by an 11-amino-acid peptide from HIV-1 Tat.
Suicide gene therapy using herpes simplex virus type-1 (HSV-1) thymidine kinase (TK) is a widely exploited approach for gene therapy of cancer and other hyperproliferative disorders. Despite its popularity, clinical success has been so far hampered mostly by the relative inefficiency of TK gene transfer and its limited bystander effect. Here we report that fusion of TK to an 11-amino-acid peptide from the basic domain of the HIV-1 Tat protein (Tat11) imparts cell membrane translocating ability to the enzyme and significantly increases its cytotoxic efficacy. In cells expressing Tat11-TK, this protein is found extracellularly, associated with cell surface heparan sulfate proteoglycans, and is released into the cell culture medium. Based on its interaction with HSPGs, the protein is then internalized by neighboring, nonexpressing cells, which become susceptible to cell death when treated with the nucleoside analogue acyclovir. As a consequence, co-cultures of wild-type cells with cells expressing Tat11-TK show increased sensitivity to ACV through a mechanism involving apoptosis. Modification of TK by fusion with Tat11 might constitute an important step for the optimization of TK suicide gene strategy for gene therapy of cellular proliferation.
Tasciotti
Ennio
E
Molecular Medicine Laboratory, International Center for Genetic Engineering and Biotechnology (ICGEB), Trieste, Italy.
Zoppè
Monica
M
Giacca
Mauro
M
D000212:Acyclovir / Q000494:pharmacology; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000998:Antiviral Agents / Q000627:therapeutic use*; D017209:Apoptosis; D005343:Fibrinolytic Agents / Q000494:pharmacology; D015696:Gene Products, tat / Q000235:genetics*; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods; D015497:HIV-1 / Q000235:genetics; D006493:Heparin / Q000494:pharmacology; D018259:Herpesvirus 1, Human / Q000201:enzymology*; D006801:Humans; D007668:Kidney / Q000166:cytology; D008546:Melanoma, Experimental / Q000188:drug therapy / Q000235:genetics; D016685:Mitomycin / Q000494:pharmacology; D009842:Oligopeptides / Q000235:genetics*; D010587:Phagocytosis; D010957:Plasmids; D011509:Proteoglycans / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D013937:Thymidine Kinase / Q000235:genetics*; D014161:Transduction, Genetic; D014407:Tumor Cells, Cultured / Q000187:drug effects; D054322:tat Gene Products, Human Immunodeficiency Virus
D000903:Antibiotics, Antineoplastic; D000998:Antiviral Agents; D005343:Fibrinolytic Agents; D015696:Gene Products, tat; D009842:Oligopeptides; D011509:Proteoglycans; D011993:Recombinant Fusion Proteins; D054322:tat Gene Products, Human Immunodeficiency Virus; D016685:Mitomycin; D006493:Heparin; D013937:Thymidine Kinase; D000212:Acyclovir