Atherosclerosis of coronary and peripheral arteries results in significant morbidity and mortality in the U.S. and other developed countries. Autologous saphenous veins are the most commonly used conduits for bypass grafting in both coronary and peripheral artery disease, and autologous veins are commonly employed for creation of arteriovenous fistulas (AVFs) for hemodialysis. The utility of these conduits is compromised by a high incidence of occlusive vein graft disease, yet no treatments exist to prevent this problem. Vein grafting provides an ideal opportunity for gene therapy, since veins are harvested from their native territory prior to grafting and can be treated ex vivo, thereby avoiding systemic exposure to the therapeutic agent. Recent Phase III clinical trials of an oligonucleotide decoy targeting the E2F transcription factor demonstrated no benefit for the prevention of vein graft failure in peripheral or coronary artery bypass grafting (CABG). Thus, there is a critical need for novel approaches to prevent vein graft failure. PTEN is a lipid phosphatase that antagonizes the effects of phosphoinositide (PI) 3-kinase, a central regulator of pathological vascular smooth muscle cell (VSMC) growth. We demonstrated previously that overexpression of PTEN in vitro inhibits the VSMC responses necessary for intimal hyperplasia. More importantly, adenovirus (Ad)-mediated delivery of PTEN to vein grafts ex vivo resulted in significant inhibition of intimal hyperplasia in a canine model of coronary artery bypass grafting (CABG). These findings suggest that PTEN gene therapy holds great promise for the prevention of clinical vein graft disease. However, adenoviral vectors induce transgene expression only on the order of weeks and are associated with a robust inflammatory response. Moreover, it is unknown whether the short-term expression achieved with AdPTEN will result in long-term vein graft patency. In contrast, newer adeno-associated virus (AAV) vectors achieve expression that is sustained for months to years and do not induce substantial inflammation. We hypothesize that long-term PTEN expression achieved with AAV will result in long-term vein graft patency rates comparable to or better than those obtained with AdPTEN. To test this hypothesis, the Specific Aims of this proposal are to: 1) Develop AAV-PTEN and optimize transduction of vascular smooth muscle cells and vein grafts;and 2) Compare 6-month aortocoronary saphenous vein graft (SVG) patency rates and intimal hyperplasia in dogs following ex vivo vein graft treatment with AdPTEN, AAV-PTEN, or saline control. Accomplishing these Specific Aims will determine the optimal agent for further clinical development, and this agent will be subsequently tested in clinical trials of human vein graft disease. PUBLIC HEALTH RELEVANCE: Autologous veins are commonly used as bypass conduits in patients with coronary and peripheral arterial disease and to create arterio-venous fistulas (AVFs) for hemodialysis, however these conduits have a high rate of failure due to intimal hyperplasia. PTEN gene therapy holds great promise to prevent vein graft failure, and studies in this proposal will determine the optimal approach for this therapeutic strategy. The results of these studies may have a substantial impact on outcomes for patients requiring venous bypass grafting and AVF placement.