Gene transfer for the treatment of neovascular ocular disease (an American Ophthalmological Society thesis).
OBJECTIVE
As vasoproliferative diseases account for a substantial fraction of worldwide blindness and share the activation of the angiogenic pathway as a common etiology, the expression of antiangiogenic proteins offers a promising means of treatment. This study was designed to develop viral vectors, harboring angiostatic genes, for the study and treatment of experimental proliferative ocular disease. A variety of methods (in vitro, ex vivo tissue, and in vivo) were employed to model the process of proliferation and test the effectiveness of these reagents.


METHODS
Antiangiogenic genes included single genes as well as hybrid genes that fused the active elements of different genes. Genes studied included the soluble vascular endothelial growth factor receptor (sKDR), soluble neuropilin (sNRP-1), tissue inhibitor of metalloproteinase-1 (TIMP-1), plasminogen gene fragments (Kringle 1-3, 1-4, and 1-5), and soluble receptors for advanced glycosylation end products (sRAGE) genes, as well as the Endo:Ang, MIG:IP10, and Endo:Kringle5 fusion genes. All genes were cloned into a lentiviral vector system and were used to produce replication deficient lentiviral particles. These viral particles were used to transduce a variety of ocular cells and tissues to test viral transfer efficiency and transgene expression. In vivo systems were employed to explore the potential of these genes as antiangiogenic agents in models of corneal and retinal neovascular disease.


RESULTS
Recombinant lentiviral particles, capable of transducing cell lines germane to eye disease (ocular endothelial, epithelial, and fibroblast cells), were successfully produced. These vectors were demonstrated to be effective in long-term transformation of cells and tissues. In vivo experiments confirmed that at least three different potentially angiostatic genes were successful in aborting the angiogenic process in the ocular models tested.


CONCLUSIONS
Lentiviral vectors are a viable means to deliver angiostatic genes to tissues of the eye. Some angiostatic genes appear to have a stronger and longer-lasting effect than others in modulating the angiogenic pathway.
Stout
John Timothy
JT
Casey Eye Institute, Oregon Health and Science University, Portland, Oregon, USA.
D020533:Angiogenesis Inhibitors / Q000235:genetics*; D000818:Animals; D002829:Choroid / Q000166:cytology; D016510:Corneal Neovascularization / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D004195:Disease Models, Animal; D004596:Electroretinography; D004797:Enzyme-Linked Immunosorbent Assay; D005347:Fibroblasts / Q000378:metabolism; D005451:Fluorescein Angiography; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D015497:HIV-1 / Q000235:genetics; D006801:Humans; D007498:Iris / Q000166:cytology; D008252:Macaca fascicularis; D008297:Male; D010857:Pigment Epithelium of Eye / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D011914:Rats, Inbred BN; D017207:Rats, Sprague-Dawley; D015861:Retinal Neovascularization / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D020533:Angiogenesis Inhibitors; D049452:Green Fluorescent Proteins