Advancing small molecule CXCR4 agonists for diabetic wound healing

项目摘要/摘要 糖尿病在美国和全球已经达到流行的程度，糖尿病患者的伤口受损 治愈是一个重大且日益严重的临床问题。我们研究的长期目标是开发小型 有效促进糖尿病伤口愈合的分子疗法。糖尿病创面缺乏 基质衍生因子-1α(SDF-1α)是一种强大的趋化因子，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。我们以前的研究数据证实，靶向SDF-1α/CXCR4信号通路 在促进糖尿病创面愈合方面有很大潜力。因此，筛选小分子激动剂可以 激活CXCR4受体及其下游通路将为糖尿病创面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， 确定CXCR4受体激动剂以促进糖尿病伤口愈合)，我们筛选了整个NIH SMR 370K分子库使用我们强大的测试漏斗，并确定了303个先导化合物。这些线索 通过反筛选、二次趋化/迁移试验进一步验证了化合物，并对其 纠正microRNA-146a、15b和29a异常表达的能力，导致识别2铅 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CAG1改善糖尿病的能力 小鼠糖尿病伤口注射后伤口愈合。我们发现一次CAG1注射导致 在明显改善糖尿病创面闭合率的同时，优化配方和作用机制 修正的幅度仍有待确定。这项工作的目标是确定我们的小说Small的能力 分子CXCR4激动剂体内纠正糖尿病创面愈合障碍，优化处方 并利用药物化学方法进行体外和体内药代动力学研究，确定其作用机制 行动，然后将这些观察扩展到临床相关的猪模型。以下是具体的 提出目标：具体目标1：对5-芳基恶唑和三氮唑进行先导优化 用结构-活性-关系药物化学方法测定噻二嗪系列 化合物体外EC50值初步测定和功能测定，Kd值测定CXCR4结合试验 并测定miR15b、miR29a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、皮肤通透性和代谢研究，并确定其作用机制 糖尿病小鼠伤口愈合模型中的糖尿病伤口愈合矫正 第4级。具体目标3：确定第5级中CAG1或CAG2先导化合物并验证CXCR4 激动剂纠正糖尿病创面愈合损害，临床前猪无毒 模特。

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.