Characterization of HIV-1 TAT peptide as an enhancer of HSV-TK/GCV cancer gene therapy
O Rautsi1, S Lehmusvaara, A Ketola
1Department of Biotechnology and Molecular Medicine, AI Virtanen Institute for Molecular Sciences, University of Kuopio, Kuopio, Finland. outi.rautsi@uku.fi
Abstract:
Cancer suicide gene therapy based on herpes simplex virus type I thymidine kinase (HSV-TK) and ganciclovir (GCV) suffers from the lack of efficacy in clinical use, which is mostly due to low gene-transfer efficiency and absence of bystander effect in tumors. We have previously demonstrated the enhancement of GCV cytotoxicity by fusing the HSV-TK with the cell penetrating peptide from HIV-1 transactivator protein transduction domain (TAT PTD). Despite the earlier promising results, we found that the triple fusion protein HIV-1 transactivator protein transduction domain-thymidine kinase suicide gene-green fluorescent protein marker gene (TAT-TK-GFP) increased GCV cytotoxicity only in 3/12 of different human tumor cell lines. Extended GCV exposure enhanced the cytotoxic effect of HSV-TK/GCV gene therapy, but the difference between TK-GFP and TAT-TK-GFP was not statistically significant. The modest improvement on cell killing mediated by TAT PTD in Chinese hamster ovary cells appeared to be associated with cell-surface heparan sulfate proteoglycan (HSPG) composition. However, TAT-mediated increased cell death did not correlate with the density of cell-surface HSPG expression in different tumor cell lines. In conclusion, although some degree of enhancement by TAT was shown in certain tumor cells in vitro, it is unlikely that TAT peptide linked to a suicide protein could be a useful booster of in vivo gene therapy trials.
Insights
Herpes simplex virus type I thymidine kinase (HSV-TK) suicide gene therapy shows limited efficacy. Fusing HSV-TK with HIV-1 transactivator protein transduction domain (TAT PTD) offered only modest improvements in certain tumor cells, limiting its in vivo potential.
Area of Science:
- Oncology
- Gene Therapy
- Molecular Biology
Background:
- Cancer suicide gene therapy using herpes simplex virus type I thymidine kinase (HSV-TK) and ganciclovir (GCV) faces challenges in clinical efficacy.
- Low gene-transfer efficiency and lack of bystander effect in tumors contribute to its limitations.
Purpose of the Study:
- To evaluate the efficacy of fusing HSV-TK with the HIV-1 transactivator protein transduction domain (TAT PTD) to enhance GCV cytotoxicity.
- To investigate the correlation between TAT-mediated enhancement and cell-surface heparan sulfate proteoglycan (HSPG) composition.
Main Methods:
- Constructed a triple fusion protein: HIV-1 transactivator protein transduction domain-thymidine kinase suicide gene-green fluorescent protein marker gene (TAT-TK-GFP).
- Assessed GCV cytotoxicity in various human tumor cell lines treated with TAT-TK-GFP.
- Analyzed the association of TAT-mediated cell death with cell-surface HSPG expression.
Main Results:
- The TAT-TK-GFP fusion protein only enhanced GCV cytotoxicity in 3 out of 12 tested human tumor cell lines.
- Extended GCV exposure improved the cytotoxic effect, but the difference between TK-GFP and TAT-TK-GFP was not statistically significant.
- TAT-mediated cell death did not consistently correlate with cell-surface HSPG density across different tumor cell lines.
Conclusions:
- While TAT PTD showed some in vitro enhancement in specific tumor cells, its overall benefit as a booster for HSV-TK/GCV gene therapy is questionable.
- The findings suggest that TAT peptide linked to a suicide protein is unlikely to be a significant advantage in in vivo gene therapy trials.
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