PROJECT SUMMARY Peripheral artery disease (PAD), an atherosclerotic disease leading to peripheral artery obstruction, affects over 6.5 million people in the US. PAD reduces blood flow, causing limb pain, non-healing wounds, and in extreme cases even death. Percutaneous transluminal angioplasty (PTA), involving inflation of a balloon at the site of blockage to re-open the arterial lumen, is a common treatment for PAD. However, vascular wall stress associated with balloon distension (sometimes in combination with stent placement) damages and activates stress response in the tissue, leading to restenosis, a re-narrowing of the artery. Because of these complications, most PTA-treated vessels fail within the first 12 months. Restenosis is driven by the pathological process intimal hyperplasia (IH), characterized by vascular smooth muscle cells (VSMCs) switching from a contractile to synthetic phenotype, causing them to become more migratory, proliferative, and active in secreting extracellular matrix (ECM) and inflammatory cytokines. Synthetic VSMC activities produce neointima that closes back off the vessel and that serves as a fertile ground for advanced atherosclerosis or even thrombosis. Current therapeutic strategies to reduce IH focus solely on inhibiting VSMC proliferation through the use of drug coated balloons and stents. However, these methods have not shown promise in improving vessel patency in clinical trials. I have been pursuing inhibition of the p38-MK2 “stress response” signaling pathway as a more comprehensive IH therapy that targets the underlying mechanisms that drive the pathological VSMC phenotype switch. In my recent studies, inhibition of MK2 through the use of a novel peptide therapeutic (MK2i) electrostatically complexed with a pH-sensitive endosomolytic polymer to form nanopolyplexes (MK2i-NPs) proved to be promising in the inhibition of VSMC proliferation, phenotype switching, and IH. In the context of vascular bypass grafts, a rabbit vascular transplant model showed that a single intra-operative treatment with MK2i-NPs (topically to explanted tissue) inhibited neointima formation up to 28 days after surgery and mitigated VSMC phenotype switching during the acute (7 day) stress response phase post-transplant. We hypothesize that in vivo, catheter-based delivery of MK2i in conjunction with balloon angioplasty will prevent VSMC phenotype switching and protect against restenosis and vessel failure. To model in vivo delivery from a specialized catheter, I will develop an ex vivo bioreactor system and apply this system to evaluate the effects of pressure on MK2i penetration and retention in the vessel wall. Additionally, we will use the bioreactor to evaluate pharmacokinetics and pharmacodynamics of MK2i in arteries ex vivo. Finally, based on the pressure conditions optimized in initial work, we will apply a specialized occlusion perfusion catheter to create a “chamber” within the lumen of the vessel through which the MK2i-NPs ...