Probing an Unexplored Intracellular Pathway in Diabetes Pathogenesis

摘要 糖尿病肾病和其他糖尿病并发症给患者和医疗保健带来巨大负担 系统，使得确定可操作的病因并开发有效、低成本的治疗方法成为当务之急 干预措施。非酶蛋白糖基化和晚期糖基化终末产物(AGEs)的形成 与发病机制有很大关系。AGE形成的驱动力是3-脱氧葡萄糖苷(3DG)，一种高度活性的 二羰基物种，通过破坏酶和DNA并使 脉管系统。因此，准确测量3DG水平并了解其病因的能力至关重要 阐明糖尿病的发病机制，限制其致病作用，提高临床治疗水平 并发症。内源性3DG被认为是由于糖化的缓慢崩解而非酶方式产生的 体内的蛋白质或从摄入的热加工食品中吸收的蛋白质。我们开发了新的方法来研究 果糖胺-3-激酶(FN3K)的酶活性，这种酶被认为可以修复糖化蛋白并防止 年龄，但FN3K活性的最终产物是3DG。我们发现肾脏中的3DG水平高于 之前已经预料到了。我们的核心假设是FN3K介导的细胞内3DG的形成是一个关键的致病因素 糖尿病并发症司机。具体目标1：我们将测量组织中产生的3DG与 KK.Cg-Ay/J小鼠2型糖尿病模型的发病机制具体目标2：高糖基化的影响 对糖尿病并发症敏感的组织将测量3DG水平的饮食，包括肾脏、心脏、 和糖尿病小鼠的肝脏。具体目标3：我们将确定其药效学特性和模式 葡甲胺是一种已经被证明是安全的药物，我们发现它有未知的药效， 提供肾脏保护和防止甘油三酯在糖尿病小鼠体内蓄积。具体目标4：A 我们发现的一系列FN3K拮抗剂将被表征为识别临床前药物开发 候选人。这项提案提供了几个对糖尿病具有高度意义和影响的主要创新要素 翻译研究。目标1将提供使用我们改进的方法来测量FN3K的新数据 更准确地开展活动和形成3DG，解决知识中的关键差距。目标2将探索两者之间的联系 细胞内3DG升高与富含果糖胺的西方饮食之间的关系 对于FN3K。药物安全对于任何糖尿病新药来说都是至高无上的。来自Aim 3的数据将加速 葡甲胺作为一种创新治疗方法的开发--一种被证明对慢性阻塞性肺疾病极其安全的化合物 用药--改善糖尿病肾病、脂肪肝和其他潜在的糖尿病并发症。目标 4提供了提供一流的酶抑制剂作为潜在的药物领先候选药物的机会。总而言之，这是 研究计划将阐明糖尿病发病机制中尚未探索的细胞内途径并提供 为更广泛地研究3DG在糖尿病肾病和其他糖尿病中的作用提供了前所未有的工具 并发症。

ABSTRACT Diabetic nephropathy and other diabetes complications impose enormous burdens on patients and healthcare systems, making it imperative to define actionable etiologic factors and develop effective, low-cost therapeutic interventions. Nonenzymatic protein glycation and the formation of advanced glycation end products (AGEs) are strongly implicated in pathogenesis. The driver of AGE formation is 3-deoxyglucosone (3DG), a highly reactive dicarbonyl species that also causes acute cellular toxicities by damaging enzymes and DNA and inflaming the vasculature. Accordingly, the ability to accurately measure 3DG levels and understand its etiology are paramount to elucidating pathogenesis, limiting its pathogenic effects, and improving clinical management of diabetic complications. Endogenous 3DG was deemed to arise nonenzymatically from the slow disintegration of glycated proteins in the body or absorbed from ingested heat-processed foods. We developed new methods to study the enzymatic activity of fructosamine-3-kinase (FN3K), an enzyme thought to repair glycated proteins and prevent AGE, but an end-product of FN3K activity is 3DG. We discovered that 3DG levels in kidney are higher than previously anticipated. Our core hypothesis is that FN3K-mediated 3DG formation in cells is a key pathogenic driver in diabetic complications. Specific Aim 1: we will measure 3DG arising in tissues in relationship to pathogenesis in the KK.Cg-Ay/J murine model of type-2 diabetes. Specific Aim 2: the impact of a high glycation diet on 3DG levels will be measured in tissues sensitive to diabetic complications, including in the kidney, heart, and liver of the diabetic mice. Specific Aim 3: We will define the pharmacodynamic properties and modes of action for meglumine, an agent, already proven safe, that we discovered has unrecognized medicinal effects, having provided nephroprotection and prevented triglyceride accumulation in diabetic mice. Specific Aim 4: a series of FN3K antagonists that we discovered will be characterized to identify a preclinical drug development candidate. This proposal offers several major innovative elements of high significance and impact in diabetes translational research. Aim 1 will provide new data developed with methodology we refined to measure FN3K activity and 3DG formation more accurately, addressing key gaps in knowledge. Aim 2 will explore the linkage between intracellular 3DG elevation and the consumption of ‘Western’ diets rich in fructosamines—the substrate for FN3K. Drug safety is paramount for any new diabetes drug. The data from Aim 3 will accelerate the development of meglumine as an innovative treatment modality—a compound proven extremely safe for chronic administration—to ameliorate diabetic nephropathy, fatty liver, and potentially other diabetic complications. Aim 4 offers opportunity to deliver first-in-class enzyme inhibitors as potential drug lead candidates. In summary, this research program will illuminate an unexplored intracellular pathway in diabetes pathogenesis and deliver unprecedented tools for broader research into the role of 3DG in diabetic nephropathy and other diabetes complications.