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HSV-tk/GCV gene therapy mediated by EBV-LMP1 for EBV-associated cancer.

Lifang Y, Min T, Midan A, Ya C - J. Exp. Clin. Cancer Res. (2008)

Bottom Line: To investigate the feasibility of gene therapy in treating Epstein-Barr virus (EBV)-associated cancer by employing the suicide gene, herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV), which uses the signaling pathway through the HIV-long terminal repeat (LTR) gene which is expressed from a nuclear factor-kappaB (NF-kappaB)-binding motif-containing promoter that is regulated by EBV-latent membrane protein 1 (LMP1) via NF-kappaB.The activation of TK was increased after transfection of the pVLTR-tk into the EBV-LMP1 positive cells.After GCV treatment, the clonogenicity and survival of the cells substantially declined, and a bystander effect was also observed.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Biology Research Center, Cancer Research Institute, XiangYa School of Medicine, Central South University, ChangSha, Hunan, 410078, PR China. anglifang99@hotmail.com

ABSTRACT

Background: To investigate the feasibility of gene therapy in treating Epstein-Barr virus (EBV)-associated cancer by employing the suicide gene, herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV), which uses the signaling pathway through the HIV-long terminal repeat (LTR) gene which is expressed from a nuclear factor-kappaB (NF-kappaB)-binding motif-containing promoter that is regulated by EBV-latent membrane protein 1 (LMP1) via NF-kappaB.

Methods: First, we constructed the plasmid pVLTR-tk, which was regulated by EBV-LMP1 via NF-kappaB, and then investigated the cytotoxic effect of the pVLTR-tk/GCV on cancer cells, using MTT assays, clonogenic assays, flow cytometry, and animal experiments.

Results: The activation of TK was increased after transfection of the pVLTR-tk into the EBV-LMP1 positive cells. After GCV treatment, the clonogenicity and survival of the cells substantially declined, and a bystander effect was also observed. The LMP1 positive cells exhibited remarkable apoptosis following pVLTR-tk/GCV treatment, and the pVLTR-tk/GCV restrained tumor growth in vivo for EBV-LMP1 positive cancers.

Conclusion: The pVLTR-tk/GCV suicide gene system may be used as a new gene targeting strategy for EBV-associated cancer.

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Thymidine kinase (tk) activity assay. Tk activity of CNE1, CNE1-LMP1, and HNE2-LMP1 cells after transfection with pVLTR-tk.
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Figure 1: Thymidine kinase (tk) activity assay. Tk activity of CNE1, CNE1-LMP1, and HNE2-LMP1 cells after transfection with pVLTR-tk.

Mentions: As shown in Figure 1, the TK enzymatic activity of CNE1 cells transfected with the pVLTR-tk was similar to that of untransfected cells, with the cpm volume being about 1 × 105, indicating that the expression of the endogenous tk gene was at a low level. However, unlike the untransfected cells, the TK enzymatic activity in CNE1-LMP1 and HNE2-LMP1 cells transfected with the pVLTR-tk maintained high levels-about three times higher than that in CNE1 cells (P < 0.01)-indicating that high expression of the pVLTR-tk gene is regulated by EBV-LMP1.


HSV-tk/GCV gene therapy mediated by EBV-LMP1 for EBV-associated cancer.

Lifang Y, Min T, Midan A, Ya C - J. Exp. Clin. Cancer Res. (2008)

Thymidine kinase (tk) activity assay. Tk activity of CNE1, CNE1-LMP1, and HNE2-LMP1 cells after transfection with pVLTR-tk.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2562992&req=5

Figure 1: Thymidine kinase (tk) activity assay. Tk activity of CNE1, CNE1-LMP1, and HNE2-LMP1 cells after transfection with pVLTR-tk.
Mentions: As shown in Figure 1, the TK enzymatic activity of CNE1 cells transfected with the pVLTR-tk was similar to that of untransfected cells, with the cpm volume being about 1 × 105, indicating that the expression of the endogenous tk gene was at a low level. However, unlike the untransfected cells, the TK enzymatic activity in CNE1-LMP1 and HNE2-LMP1 cells transfected with the pVLTR-tk maintained high levels-about three times higher than that in CNE1 cells (P < 0.01)-indicating that high expression of the pVLTR-tk gene is regulated by EBV-LMP1.

Bottom Line: To investigate the feasibility of gene therapy in treating Epstein-Barr virus (EBV)-associated cancer by employing the suicide gene, herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV), which uses the signaling pathway through the HIV-long terminal repeat (LTR) gene which is expressed from a nuclear factor-kappaB (NF-kappaB)-binding motif-containing promoter that is regulated by EBV-latent membrane protein 1 (LMP1) via NF-kappaB.The activation of TK was increased after transfection of the pVLTR-tk into the EBV-LMP1 positive cells.After GCV treatment, the clonogenicity and survival of the cells substantially declined, and a bystander effect was also observed.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Biology Research Center, Cancer Research Institute, XiangYa School of Medicine, Central South University, ChangSha, Hunan, 410078, PR China. anglifang99@hotmail.com

ABSTRACT

Background: To investigate the feasibility of gene therapy in treating Epstein-Barr virus (EBV)-associated cancer by employing the suicide gene, herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV), which uses the signaling pathway through the HIV-long terminal repeat (LTR) gene which is expressed from a nuclear factor-kappaB (NF-kappaB)-binding motif-containing promoter that is regulated by EBV-latent membrane protein 1 (LMP1) via NF-kappaB.

Methods: First, we constructed the plasmid pVLTR-tk, which was regulated by EBV-LMP1 via NF-kappaB, and then investigated the cytotoxic effect of the pVLTR-tk/GCV on cancer cells, using MTT assays, clonogenic assays, flow cytometry, and animal experiments.

Results: The activation of TK was increased after transfection of the pVLTR-tk into the EBV-LMP1 positive cells. After GCV treatment, the clonogenicity and survival of the cells substantially declined, and a bystander effect was also observed. The LMP1 positive cells exhibited remarkable apoptosis following pVLTR-tk/GCV treatment, and the pVLTR-tk/GCV restrained tumor growth in vivo for EBV-LMP1 positive cancers.

Conclusion: The pVLTR-tk/GCV suicide gene system may be used as a new gene targeting strategy for EBV-associated cancer.

Show MeSH
Related in: MedlinePlus