Tumor-specific gene expression using regulatory elements of the glucose transporter isoform 1 gene.
In order to achieve tumor-specific targeting of adeno-associated virus (AAV)-mediated gene expression, the promoter of the glucose transporter isoform 1 (GLUT1) gene was cloned upstream of the enhanced green fluorescence protein (EGFP) and the herpes simplex virus thymidine kinase (HSVtk) gene. FACS analysis performed at 48 h after transient infection with rAAV/cytomegalovirus (CMV)egfp viral particles revealed an increase of fluorescence in all the cell lines tested. However, EGFP expression under control of the GLUT1 promoter element (rAAV/GTI-1.3egfp) was limited to the tumor cells and oncogene-transformed cells. Evidence for phosphorylation of the HSVtk substrates ganciclovir (GCV) and 125I-deoxycytidine was found in all transfected tumor cell lines compared to noninfected controls (HCT116: 111%; MH3924A: 130%; HaCaT-RT3: 257% increase), but not in HaCaT and HUVEC cells. Furthermore, tumor cells and the oncogene-transformed (ras) cell line HaCaT-RT3 showed a GCV-induced reduction in cell number (HCT116: -71%; MH3924A: -43% and HaCaT-RT3: -31%). No statistically relevant cytotoxic effect was observed in HaCaT (6% decrease) and HUVEC cells (2% decrease). Furthermore, a reduction of 3H-thymidine incorporation into the DNA was seen after treatment with GCV (HCT116: 38%; MH3924A: 33% and HaCaT-RT3: 37% decrease). In a therapy study of HSVtk-expressing tumors with GCV, we achieved total tumor remission.
Sieger
Stephanie
S
Clinical Cooperation Unit Nuclear Medicine, Heidelberg, Germany.
Jiang
Shiming
S
Kleinschmidt
Jürgen
J
Eskerski
Helmut
H
Schönsiegel
Frank
F
Altmann
Annette
A
Mier
Walter
W
Haberkorn
Uwe
U
D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D003841:Deoxycytidine / Q000378:metabolism / Q000493:pharmacokinetics / Q000494:pharmacology; D000229:Dependovirus / Q000502:physiology; D005434:Flow Cytometry; D015774:Ganciclovir / Q000493:pharmacokinetics / Q000494:pharmacology; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D051379:Mice; D009004:Monosaccharide Transport Proteins / Q000235:genetics*; D009368:Neoplasm Transplantation; D009928:Organ Specificity; D020033:Protein Isoforms / Q000235:genetics; D051381:Rats; D011912:Rats, Inbred ACI; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism
D009004:Monosaccharide Transport Proteins; D020033:Protein Isoforms; D003841:Deoxycytidine; D013937:Thymidine Kinase; D015774:Ganciclovir