1 Project Summary: Anti-integrins are considered as promising alternative for angiogenesis related disease 2 conditions including cancer, Crohn’s, and retinal diseases. However, an optimal nanomaterial design by 3 considering the application and target cell receptor expression is extremely vital for the success of the 4 formulation. Although polymeric nanocarriers have demonstrated considerable potential associated with anti- 5 integrin peptide delivery, the peptide valency and surface distance are rarely co-optimized to enhance 6 therapeutic ligand clustering. Hence, the primary goal of this proposal is to develop and validate a novel 7 bioresponsive hydrogel depot for the sustained delivery of micelles carrying therapeutic peptides at an optimal 8 valency for therapeutic efficacy. A key innovation of this proposal is that we will use self-assembled poly(ethylene 9 glycol)-b-poly(propylene sulfide) (PEG-b-PPS) filomicelles (FM) that use cylinder-to-sphere thermodynamic 10 transitions for sustained in situ generation and delivery of peptide-loaded micelles (MC). In this proposed work, 11 the peptide’s valency and distance from the MC surface will be co-optimized for an efficient therapeutic effect. 12 As proof of concept, the current proposal will use an anti-integrin heptapeptide that showed therapeutic efficiency 13 in vitro using HUVEC cells. With the strong initial results, the objective of the current proposal is to study the 14 mechanistic investigation of the FM-hydrogel depot through in vitro, ex vivo and in vivo investigations and 15 generate preliminary data for future translational grant mechanisms involving appropriate animal disease 16 models. Ex vivo model in this proposal will assist in selecting the optimum formulation and reduce the use of 17 animals and align with the current grant mechanism. The following specific aims will be achieved: 18 AIM 1: Synthesize, optimization and In vitro characterization of FM-depots for the sustained release of MCs 19 displaying a co-optimized surface display of anti-integrin peptide (aANG-P). Aim 1a will address the development 20 of FM-depots with aANG-P constructs, verify control over the MC release rate and confirm that peptide valency 21 and bioactivity remain unchanged following the FM-to-MC transition. Aim 1b will study the inflammatory response 22 induced by FM depot using primary macrophages and confirm of therapeutic activity of the released MC using 23 HUVECs and macrophage cells. 24 AIM 2: Ex vivo and in vivo tissue distribution and bioactivity of aANGP-MC released from aANG-P FM-depots. 25 Aim 2a will study the micellar release kinetics, and tissue distribution from aANGP-FM injected suprachoroidal 26 space of excised rabbit whole eye and study the bioactivity using choroid sprouting assay and CAM assay. Aim 27 2b will confirm the tissue distribution of the aANGP-FM depot using Sprague Dawley rat model and study the 28 inflammatory response after injecting at suprachoroidal space...