Advancing small molecule CXCR4 agonists for diabetic wound healing

项目总结/摘要 糖尿病在美国和全球范围内已达到流行病的程度， 愈合是一个重要且日益严重的临床问题。我们研究的长期目标是开发小型 分子疗法，以有效促进糖尿病伤口的愈合。糖尿病伤口缺乏 基质衍生因子-1 α（SDF-1 α）是一种参与祖细胞募集、血管生成 和肉芽组织的形成，通过与CXCR4受体结合和建立一个 趋化梯度我们前期的研究数据证实，靶向SDF-1 α/CXCR4信号通路， 具有改善糖尿病伤口愈合的巨大潜力。因此，筛选小分子激动剂， 激活CXCR4受体及其下游通路将为糖尿病伤口愈合提供新的治疗方法， 具有很大的临床应用和商业化潜力。在我们之前的RO1（R01DK105010， 鉴定CXCR4受体激动剂以改善糖尿病伤口愈合），我们筛选了整个NIH SMR 使用我们的强大的测试漏斗，从> 370k分子库中鉴定出303种先导化合物。这些引线 化合物进一步通过反筛选、次级趋化性/迁移试验和它们的 能够纠正microRNA-146a、15b和29a的异常表达，导致鉴定出2个前导基因 脚手架在一项令人兴奋的初步研究中，我们测试了我们的铅支架CAG 1改善糖尿病的能力。 注射到鼠糖尿病伤口后的伤口愈合。我们发现单次注射CAG 1导致 在糖尿病伤口愈合率的显着改善，但最佳配方和机制， 更正的幅度仍有待确定。这项工作的目的是确定我们的小说小的能力， 分子CXCR 4激动剂可纠正体内糖尿病伤口愈合障碍，优化配方 和药代动力学在体外和体内使用药物化学方法，确定的机制， 动作，然后将这些观察结果扩展到临床相关的猪模型。以下具体 具体目标1：对5-芳基恶唑和三唑进行先导优化 噻二嗪系列药物化学构效关系测定 化合物在主要和功能试验中的体外EC50值，CXCR4结合试验中的KD值 并确定miR15 b和miR29 a对人糖尿病成纤维细胞表达的影响。具体目标2： 进行体外稳定性、皮肤渗透性和代谢研究，并确定 在小鼠伤口愈合模型中，化合物的糖尿病伤口愈合校正进展至 第4级。具体目标3：确定第5级中的CAG 1或CAG 2先导化合物，并确定CXCR 4 激动剂纠正糖尿病伤口愈合障碍，并且在临床前猪中无毒 模型

Project Summary/Abstract Diabetes has reached epidemic proportions in the United States and globally, and impaired diabetic wound healing is a significant and growing clinical problem. The long-term goal of our research is to develop small molecule therapeutics to effectively promote healing of diabetic wounds. Diabetic wounds are deficient in stromal derived factor-1α (SDF-1α), a potent chemokine involved in progenitor cell recruitment, angiogenesis, and granulation tissue formation, mediated through binding to the CXCR4 receptor and the establishment of a chemotactic gradient. Our previous research data confirmed that targeting SDF-1α/CXCR4 signaling pathway has great potential to improve diabetic wound healing. Therefore, screening small molecule agonists that can activate CXCR4 receptor and its downstream pathway will provide a novel therapy for diabetic wound healing, and has great potential for clinical application and commercialization. In our previous RO1 (R01DK105010, Identifying CXCR4 receptor agonists to improve diabetic wound healing), we screened the entire NIH SMR library of >370k molecules using our robust testing funnel and identified 303 lead compounds. These lead compounds were further validated with by counter-screen, secondary chemotaxis/migration assays, and their ability to correct abnormal expression of microRNA-146a, 15b, and 29a, resulting in the identification of 2 lead scaffolds. In an exciting preliminary study we tested the ability of our lead scaffold CAG1 to improve diabetic wound healing following injection into murine diabetic wounds. We found that a single CAG1 injection resulted in a significant improvement in the rate of diabetic wound closure, but the optimal formulation and mechanisms of correction remain to be determined. The objective of this work is to determine the ability of our novel small molecule CXCR4 agonists to correct the diabetic wound healing impairment in vivo, optimize the formulation and pharmacokinetics in vitro and in vivo using a medicinal chemistry approach, determine the mechanisms of action, and then extend these observations to the clinically relevant porcine model. The following Specific Aims are proposed: Specific aim 1: Conduct lead optimization of the 5-aryl oxazole and triazolo thiadiazine series using structure- activity- relationship medicinal chemistry approaches, Determine compound in vitro EC50 values in primary and functional assays, KD values in a CXCR4 binding assay and 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 of diabetic wound healing correction in a murine model of wound healing for compounds advancing to tier 4. Specific aim 3: Identify a CAG1 or CAG2 lead compound in tier 5 and Validate that CXCR4 agonist corrects the diabetic wound healing impairment and is non-toxic in a pre-clinical porcine model.