Low Molecular Weight Protein Nephrotoxicity

描述(由申请人提供)： 在过去的二十五年中，在免疫球蛋白游离轻链(以下简称FLC)引起的肾小管间质损伤的特征以及FLC介导的肾小管损伤的结构-功能关系的阐明方面，低分子量蛋白质肾毒性研究取得了显著进展。分子发病机制已被详细描述，增加了创造新的治疗方法的可能性，旨在中断病理过程，防止与单克隆性Flc沉积相关的进行性肾功能丧失。氧化应激是细胞功能和命运的关键决定因素，尤其是在疾病状态下。已发表的和本实验室的初步研究表明，Flcs的内吞作用产生过氧化氢，并诱导细胞内氧化应激，促进肾上皮细胞的激活和损伤。第一个目标将进一步表征FLC对导致细胞凋亡或细胞存活的近端肾小管上皮细胞信号事件的影响。本实验室发表了证明管腔内管型形成的机制和功能意义的开创性观察，最近未发表的研究为管型形成导致急性肾损伤的概念性实验提供了关键证据，并可以通过使用与Tamm-Horsfall蛋白(THP)上的FLC结合结构域(LCBD)相结合的竞争肽来预防。这些研究允许将第二个目标的唯一重点放在FLC与LCBD的相互作用上，而THP是管型肾病的直接原因。这一应用的主要理论是，FLC所致的肾小管间质损伤是一个复杂的过程，涉及近端肾小管细胞的激活和损伤以及管腔内管型的形成。这些事件可以独立发生，也可以协同作用，产生急性肾损伤和慢性肾脏疾病的临床症状。我们将追求两个目标：目标1.确定在近端肾小管上皮细胞暴露于FLC后，细胞内信号转导事件发生改变并影响细胞存活。需要检验的具体假设是，Flc的内吞作用进入近端小管，激活c-Src(c-src的60 kDa产物，也称为pp60c-src)和凋亡信号调节蛋白1(ASK1)信号通路，这两个信号通路激活下游分子，共同作用于平衡细胞生存和凋亡。目的2.明确Flc与THP上的FLC结合域(LCBD)之间的分子相互作用及其功能意义。需要检验的假设是，小分子非肽和多肽化合物有效地与结合在THP上的LCBD的FLC竞争，从而预防管型肾病。该项目的长期目标--了解这些过程中涉及的分子机制--提供了开发新的治疗方法的潜力，旨在改善单克隆性FLC过度生产的情况下的肾功能衰竭，如多发性骨髓瘤。好了！

DESCRIPTION (provided by applicant): Low Molecular Weight Protein Nephrotoxicity Significant advances have been made over the past twenty-five years in the characterization of tubulointerstitial renal injury caused by immunoglobulin free light chains (abbreviated FLC in this application) and in elucidation of the structure-function relationships involved in FLC-mediated tubular damage. The molecular pathogenesis has been described in some detail, increasing the potential to create novel treatments designed to interrupt pathological proceses and prevent progresive los of renal function related to deposition of monoclonal FLCs. Oxidative stres is a critical determinant of cell function and fate, especially in disease states. Published and preliminary studies from this laboratory demonstrated that endocytosis of FLCs generates hydrogen peroxide and induces intracellular oxidative stress that promotes renal epithelial cell activation and injury. The first aim will characterize further the effects of FLC on proximal tubular epithelial cell signaling events that lead to apoptosis or cell survival. The seminal observations that demonstrated the mechanism and functional significance of intraluminal cast formation have been published by this laboratory, and recent unpublished studies have provided critical proof of concept experiments that cast formation causes acute kidney injury and can be prevented with the use of a competitor peptide that binds to the FLC binding domain (LCBD) on Tamm-Horsfall protein (THP). These studies permit the singular focus of the second aim on the interactions of FLC with the LCBD on THP as the proximate cause of cast nephropathy. The overarching theory of this application is that tubulointerstitial renal damage from FLC is a complex process that involves both activation and injury of proximal tubular cells and intraluminal cast formation. These events can occur independently or synergize to produce the clinical syndromes of acute kidney injury and chronic kidney disease. Two aims will be pursued: Aim 1. Define the intracellular signaling events that are altered and effect cell survival following exposure of proximal tubular cells to FLC. The specific hypothesis to be tested is that endocytosis of FLCs into the proximal tubule activates c-Src (60-kDa product of c- src, also known as pp60c-src) and apoptosis signal-regulating kinase 1 (ASK1) signaling kinases, which activate downstream molecules that work in concert to balance cell survival and apoptosis. Aim 2. Define the molecular interaction between FLCs and the FLC binding domain (LCBD) on THP and the functional significance of this interaction. The hypothesis to be tested is that small molecular weight nonpeptide and peptide compounds effectively compete with FLC binding to the LCBD on THP and prevent cast nephropathy. The long-term goal of this project - understanding the molecular mechanisms involved in these processes - offers the potential to develop new treatments designed to ameliorate renal failure in the setting of overproduction of monoclonal FLC, such as multiple myeloma. !