Hepatoma cell-specific ganciclovir-mediated toxicity of a lentivirally transduced HSV-TkEGFP fusion protein gene placed under the control of rat alpha-fetoprotein gene regulatory sequences.
Suicide gene therapy combining herpes simplex virus thymidine kinase gene transfer and ganciclovir administration can be envisioned as a powerful therapeutical approach in the treatment of hepatocellular carcinoma; however, safety issues regarding transgene expression in parenchyma cells have to be addressed. In this study, we constructed LATKW, a lentiviral vector expressing the HSV-TkEGFP gene placed under the control of the promoter elements that control the expression of the rat alpha-fetoprotein, and assayed its specific expression in vitro in hepatocarcinoma and nonhepatocarcinoma human cell lines, and in epidermal growth factor stimulated human primary hepatocytes. Using LATKW, a strong expression of the transgene was found in transduced hepatocarcinoma cells compared to a very low expression in nonhepatocarcinoma human cell lines, as assessed by Northern blot, RT-PCR, FACS analysis and ganciclovir-mediated toxicity assay, and no expression was found in lentivirally transduced normal human hepatocytes. Altogether, these results demonstrate the possibility to use a lentivirally transduced expression unit containing the rat alpha-fetoprotein promoter to restrict the HSV-TK-mediated induced GCV sensitivity to human hepatocarcinoma cells.
Uch
Rathviro
R
Département de thérapie cellulaire et génique, Etablissement Français du Sang Alpes-Méditerranée, Marseille, France.
Gérolami
René
R
Faivre
Jamila
J
Hardwigsen
Jean
J
Mathieu
Sylvie
S
Mannoni
Patrice
P
Bagnis
Claude
C
D000818:Animals; D015152:Blotting, Northern; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism / Q000473:pathology* / Q000628:therapy*; D016923:Cell Death; D045744:Cell Line, Tumor; D015774:Ganciclovir / Q000633:toxicity*; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism / Q000473:pathology; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013937:Thymidine Kinase / Q000235:genetics* / Q000378:metabolism*; D014161:Transduction, Genetic; D014162:Transfection; D000509:alpha-Fetoproteins / Q000235:genetics*
D008164:Luminescent Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D000509:alpha-Fetoproteins; D049452:Green Fluorescent Proteins; D013937:Thymidine Kinase; D015774:Ganciclovir