Fibroblast growth factor-2-retargeted adenoviral vector for selective transduction of primary glioblastoma multiforme endothelial cells.
OBJECTIVE
Adenovirus transduction in gene therapy is dependent on the expression of the coxsackie virus-adenovirus receptor (CAR) for initial binding and on the integrin receptors (avb3, avb5) for viral internalization. Low and variable expression of CAR may be responsible for the low transduction rates seen with native adenoviral vectors. The goal of this study was to demonstrate increased transduction efficiency by retargeting the adenovirus with a fibroblast growth factor (FGF) ligand, FGF-2.


METHODS
The retargeted adenoviruses were used to transduce human glioblastoma multiforme (GBM)-derived ECs (tumor-associated brain endothelial cells [TuBECs]), in which there is minimal CAR expression but a high expression of FGF receptor (FGFR). The results demonstrate that the transduction efficiency of TuBECs can reach as high as 80% when one uses an FGF2-conjugated adenovirus containing green fluorescent protein (FGF2-AdGFP) yet be only 5% when one uses the native adenovirus (AdGFP). The TuBECs were transduced with either a native adenovirus (AdHSV-TK) or a retargeted adenovirus (FGF2-AdHSV-TK), both of which carry the suicide herpes simplex virus-thymidine kinase (HSV-TK) gene. Administered as a cytotoxic prodrug, ganciclovir induced a significant decline in the proliferation rate and increased apoptosis in TuBECs treated with the retargeted adenovirus, compared with its effect on TuBECs treated with the native adenovirus. Increased transduction efficiency was determined by performing GFP-based flow cytometry, and the expression of the TK protein by the retargeted adenovirus was assessed by performing an immunohistochemical analysis focused on HSV-TK. The mechanism of cytotoxicity was determined to be apoptosis by performing a terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay.


CONCLUSIONS
Fibroblast growth factor-2-retargeted adenoviral vectors may be used to increase the transduction of GBM-derived endothelial cells, enabling a new and efficient antiangiogenesis strategy for the treatment of malignant gliomas.
Gupta
Vinay
V
Department of Pathology, K. Norris Jr. Comprehensive Cancer Center, University of Southern California, Los Angeles, California 90033, USA.
Wang
Weijun
W
Sosnowski
Barbara A
BA
Hofman
Florence M
FM
Chen
Thomas C
TC
D000256:Adenoviridae / Q000235:genetics; D020533:Angiogenesis Inhibitors / Q000235:genetics; D000998:Antiviral Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D001932:Brain Neoplasms / Q000098:blood supply / Q000503:physiopathology / Q000628:therapy*; D002478:Cells, Cultured; D004351:Drug Resistance / Q000235:genetics; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology / Q000821:virology; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000494:pharmacology* / Q000627:therapeutic use; D015774:Ganciclovir / Q000494:pharmacology; D041722:Genes, Transgenic, Suicide / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000187:drug effects* / Q000235:genetics / Q000627:therapeutic use; D005909:Glioblastoma / Q000098:blood supply / Q000503:physiopathology / Q000628:therapy*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000494:pharmacology / Q000627:therapeutic use; D006801:Humans; D009389:Neovascularization, Pathologic / Q000235:genetics / Q000517:prevention & control / Q000628:therapy*; D050130:Oncolytic Virotherapy / Q000379:methods*; D017468:Receptors, Fibroblast Growth Factor / Q000187:drug effects / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000494:pharmacology / Q000627:therapeutic use; D013937:Thymidine Kinase / Q000235:genetics; D014161:Transduction, Genetic / Q000379:methods
D020533:Angiogenesis Inhibitors; D000998:Antiviral Agents; D017468:Receptors, Fibroblast Growth Factor; D011993:Recombinant Fusion Proteins; D016222:Fibroblast Growth Factor 2; D049452:Green Fluorescent Proteins; D013937:Thymidine Kinase; D015774:Ganciclovir