Use of Epstein-Barr virus nuclear antigen-1 in targeted therapy of EBV-associated neoplasia.
To target expression of toxic genes to Epstein-Barr virus (EBV)-associated tumor cells, we have developed an EBV-driven enzyme prodrug system (EDEPS) that takes advantage of the trans-activating properties of EBNA1, a latent protein expressed in all EBV-containing cells, to direct expression of cytosine deaminase (CD) at high levels in those cells only. Plasmids were constructed in which the CD gene or a luciferase reporter gene were cloned downstream of the herpes simplex virus thymidine kinase (tk) promoter and the family of repeats (FR) sequence from the oriP region of EBV. Analysis of luciferase activity after transient transfection into a panel of EBV-negative or -positive human cell lines showed that the presence of the FR element enhanced transcription from the tk promoter in all EBV-positive cell lines, whereas transcription from tk was repressed in all EBV-negative cell lines, including B, T, and fibroblast cell lines. In clonogenicity assays following transfection with the CD vector, the presence of 5-fluorocytosine (5-FC) in the culture medium completely abolished cell growth in EBV-positive cell lines, but did not affect the growth of EBV-negative cell lines. This vector system should have wide applicability in that it allows targeted expression of any gene of interest to tumors that carry EBV, irrespective of the role EBV plays in their pathogenesis.
Judde
J G
JG
Lymphoma Biology Section, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Spangler
G
G
Magrath
I
I
Bhatia
K
K
D000956:Antigens, Viral / Q000627:therapeutic use*; D001483:Base Sequence; D002470:Cell Survival / Q000235:genetics; D043525:Cytosine Deaminase; D004268:DNA-Binding Proteins / Q000627:therapeutic use*; D019309:Epstein-Barr Virus Nuclear Antigens; D005437:Flucytosine / Q000378:metabolism; D005472:Fluorouracil / Q000378:metabolism / Q000633:toxicity; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D004854:Herpesvirus 4, Human / Q000235:genetics / Q000276:immunology*; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000628:therapy* / Q000821:virology; D009700:Nucleoside Deaminases / Q000235:genetics* / Q000378:metabolism; D011355:Prodrugs / Q000378:metabolism; D015534:Trans-Activators / Q000627:therapeutic use*; D014162:Transfection; D014407:Tumor Cells, Cultured
D000956:Antigens, Viral; D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D011355:Prodrugs; D015534:Trans-Activators; D005437:Flucytosine; D008156:Luciferases; D009700:Nucleoside Deaminases; D043525:Cytosine Deaminase; D005472:Fluorouracil