Project Summary/AbstractDiabetes has reached epidemic proportions in the United States and globally and impaired diabetic woundhealing is a significant and growing clinical problem. The long-term goal of our research is to develop smallmolecule therapeutics to effectively promote healing of diabetic wounds. Diabetic wounds are deficient instromal derived factor-1 (SDF-1) a potent chemokine involved in progenitor cell recruitment angiogenesisand granulation tissue formation mediated through binding to the CXCR4 receptor and the establishment of achemotactic gradient. Our previous research data confirmed that targeting SDF-1/CXCR4 signaling pathwayhas great potential to improve diabetic wound healing. Therefore screening small molecule agonists that canactivate CXCR4 receptor and its downstream pathway will provide a novel therapy for diabetic wound healingand has great potential for clinical application and commercialization. In our previous RO1 (R01DK105010Identifying CXCR4 receptor agonists to improve diabetic wound healing) we screened the entire NIH SMRlibrary of >370k molecules using our robust testing funnel and identified 303 lead compounds. These leadcompounds were further validated with by counter-screen secondary chemotaxis/migration assays and theirability to correct abnormal expression of microRNA-146a 15b and 29a resulting in the identification of 2 leadscaffolds. In an exciting preliminary study we tested the ability of our lead scaffold CAG1 to improve diabeticwound healing following injection into murine diabetic wounds. We found that a single CAG1 injection resultedin a significant improvement in the rate of diabetic wound closure but the optimal formulation and mechanismsof correction remain to be determined. The objective of this work is to determine the ability of our novel smallmolecule CXCR4 agonists to correct the diabetic wound healing impairment in vivo optimize the formulationand pharmacokinetics in vitro and in vivo using a medicinal chemistry approach determine the mechanisms ofaction and then extend these observations to the clinically relevant porcine model. The following SpecificAims are proposed: Specific aim 1: Conduct lead optimization of the 5-aryl oxazole and triazolothiadiazine series using structure- activity- relationship medicinal chemistry approaches Determinecompound in vitro EC50 values in primary and functional assays KD values in a CXCR4 binding assayand determine effect on human diabetic fibroblast expression of miR15b and miR29a. Specific aim 2:Perform in vitro stability skin permeability and metabolism studies and determine the mechanisms ofdiabetic wound healing correction in a murine model of wound healing for compounds advancing totier 4. Specific aim 3: Identify a CAG1 or CAG2 lead compound in tier 5 and Validate that CXCR4agonist corrects the diabetic wound healing impairment and is non-toxic in a pre-clinical porcinemodel.