Abstract Ischemia reperfusion injury (IRI) causes endothelial inflammation and microvascular rarefaction that leads to adverse kidney graft outcomes in organ transplant. Direct treatment of endothelial cells (EC) can reduce the impact of IRI on the health of the graft, but there is a lack of EC targeted therapies that can effectively intervene and alleviate the various modes of dysfunctional endothelial response. The goal of this work is to develop a therapeutic strategy that addresses the two key modes of endothelial damage in response to IRI: dysfunctional inflammation in ECs and damage to capillary networks, in a site-specific and temporary manner. We propose that therapeutic siRNA can be delivered directly to endothelial cells using polymeric nanoparticles (NPs), which provide a customizable platform to enhance the cell penetration and to sustain the delivery of nucleic acids. In Aim 1, we will determine the NP characteristics utilizing a novel family of PACE polymers that enable maximum and sustained siRNA to endothelial cells in order to reduce adhesion molecule expression upon inflammatory activation. In Aim 2, we will translate this knowledge of structure/function relationship of the NP to rationally design siRNA-mediated knockdown of adhesion molecules in relevant models of 3D human vasculature and evaluate the long-term effect after transplantation in vivo. In the R00 phase of the award, the principles determined in Aim 1 and 2 to impact endothelial-NP interaction will be applied to polymer NPs delivered within a hydrogel delivery vehicle to the renal cortex. Aim 3 will investigate the potential of endothelial-tailored siRNA-NPs to locally deliver anti-fibrotic siRNAs within an ECM-derived hydrogel to IRI- damaged renal cortex in vivo. Dr. Laura Bracaglia has earned her PhD in Bioengineering and is currently a postdoctoral fellow in the Department of Biomedical Engineering at Yale University. In her training so far, she has studied NP and drug delivery methods in human tissue models that provide translatable insights into vascular inflammation. To successfully accomplish the specific aims of this proposal, Dr. Bracaglia has identified that she will need additional training in the 1) chemical and polymer science aspects involved in the development of NPs. In addition, the impact of the proposed work would be enhanced with training and expertise in 2) vascular immune biology, 3) renal pathology and response to injury, and 4) translation to human immunology. To train in these areas, Dr. Bracaglia has assembled a team of expert mentors who can provide clinical perspective and technical expertise. In addition, she has planned key course work and set milestones for progress in scientific and professional goals. This proposed training in the K99 mentored phase will support meeting the initial goals of this work. NP treatment strategies developed during the mentored phase, together with her expertise in the development of ECM-based biomaterials...