Percutaneous transluminal coronary angioplasty (PTCA) and coronary artery bypass graft surgery (CABG) are each used to treat approximately 400,000 coronary artery disease patients annually in the United States. PTCA and saphenous vein grafts in CABG are both complicated by proliferation of smooth muscle cells in the vessel subintima, in a process termed neointimal hyperplasia. Neointimal hyperplasia leads to significant coronary restenosis in approximately 30% of PTCA cases, and engenders significant vein graft stenoses--requiring some sort of intervention--in 30% of CABG cases within two years of surgery. The annual cost of PTCA restenosis alone exceeds $2 billion. This work proposes to test the hypothesis that transient gene therapy with G protein-coupled receptor kinases (GRKs) at the time of PTCA or saphenous in graft placement may substantially alleviate neointimal hyperplasia. GRKs are a family of enzymes which initiate either desensitization or signal termination for many G protein-coupled receptors, several of which have been implicated in the pathology of vascular smooth muscle cell proliferation and restenosis: endothelin A and B, angiotensin II (type I), thrombin, and thromboxane A2 receptors. Over-expression of GRK2 can drastically reduce signaling through all of these receptors in transfected cell systems. Systematically administered antagonists of the endothelin and angiotensin II (type I) receptors significantly reduce neointimal hyperplasia in animal models of vascular injury. This proposal therefore seeks to develop recombinant adenoviruses for expressing GRKs 2, 3, and 5 transiently in smooth muscle cells, and to test each of these recombinant viruses in rabbit and human smooth muscle cells for the ability to reduce cellular responses to agonists for the receptors listed above, as assessed by inositol phosphate signaling, cellular migration, DNA synthesis, and cell proliferation. Subsequently, GRK adenoviruses and adeno-associated viruses will be used ex vivo to treat rabbit jugular veins, which will be grated across the rabbit's ligated carotid artery in a well-established model of neointimal hyperplasia. Four weeks after surgery, vein grafts will be evaluated to determine the GRK's ability (a) to attenuate neointimal hyperplasia in vivo, by histology, and (b) to attenuate agonist-stimulated hyperplasia by transiently over-expressed GRKs in the rabbit model of vascular injury would have therapeutic implications for restenosis in PTCA and vein graft failure in CABG.
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