Mechanisms of Aggregation in Light Chain Amyloidosis

描述（由申请人提供）：我们拟议研究的总体目标是剖析导致轻链淀粉样变性（AL）沉积形成的分子机制。AL是一种由血浆B细胞异常增殖引起的毁灭性疾病。这些肿瘤细胞分泌单克隆免疫球蛋白轻链，这些轻链错误折叠为截短的轻链，进入重要器官的淀粉样蛋白沉积物中，导致组织损伤和器官衰竭。在免疫球蛋白可变结构域中积累的体细胞突变使得这些蛋白质比它们的非淀粉样蛋白对应物更不稳定。免疫球蛋白轻链和重链均在成熟的血浆B细胞中产生，在那里它们结合形成分泌到循环中的抗体。在大多数AL患者中，即使当重链存在于浆细胞中时，也仅轻链被分泌到循环中，这表明未能与重链结合。目前尚不清楚是什么原因导致与重链、细胞区室或导致蛋白质截短的事件的结合丧失，以及截短是否是触发淀粉样蛋白形成的重要不稳定事件。最后，鉴于AL丰富的突变多样性，没有全面的研究已经做了了解AL中蛋白质折叠，稳定性和淀粉样变性的突变位置的作用。我们假设，AL蛋白不稳定是由于采样的部分未折叠状态触发的一个或多个以下事件：体细胞突变，从免疫球蛋白重链解离，和/或蛋白水解裂解。目的1研究可变区、截短区和全长AL蛋白的结构和稳定性。我们还将进行AL突变到种系残基的系统性恢复，以鉴定导致淀粉样蛋白形成倾向的突变。目的2将表征AL蛋白的淀粉样蛋白形成。我们将研究硫酸钠稳定淀粉样蛋白生成中间体和加速淀粉样蛋白形成的作用。目的3将研究AL的细胞生物学。我们将确定HC基因的完整性，AL蛋白与重链二聚化的能力，并将研究细胞培养中轻链的内化，以确定是否发生细胞内蛋白水解。这项工作将增加我们对AL发病机制的理解，帮助我们制定更好的管理和治疗策略。

DESCRIPTION (provided by applicant): The overall goal of our proposed research is to dissect the molecular mechanisms that cause Light chain amyloidosis (AL) deposits to form. AL is a devastating disease caused by the abnormal proliferation of plasma B cells. These tumor cells secrete monoclonal immunoglobulin light chains that misfold as truncated light chains into amyloid deposits in vital organs, causing tissue damage and organ failure. Somatic mutations that accumulate in the immunoglobulin variable domain make these proteins less thermodynamially stable than their non-amyloidogenic counterparts. Both immunoglobulin light and heavy chains are produced in the mature plasma B cell where they associate to form antibodies that are secreted into circulation. In most AL patients, only light chain is secreted into circulation, even when the heavy chain is present in the plasma cell, suggesting a failure to associate with the heavy chain. Many questions remain unanswered in regards to AL. It is not clear what causes the loss of association with the heavy chain, the cellular compartment or event that leads to the truncation of the protein, and whether or not the truncation is an important destabilzing event that triggers amyloid formation. Finally, given the rich mutational diversity of AL, no comprehensive study has been done to understand the role of the location of mutations in protein folding, stability and amyloidogenicity in AL. We hypothesize that AL protein destabilization is due to sampling of partially unfolded states triggered by one or more of the following events: somatic mutations, dissociation from the immunoglobulin heavy chain, and/or proteolytic cleavage. Aim 1 of this work will study the structure and stability of variable domain, truncated and full length AL proteins. We will also perform a systematic restoration of AL mutations to germline residues to identify the mutations responsible for the propensity to form amyloid. Aim 2 will characterize amyloid formation by AL proteins. We will study the role of sodium sulfate stabilizing amyloidogenic intermediates and accelerating amyloid formation. Aim 3 will study the cell biology of AL. We will determine the integrity of the HC gene, the ability of AL proteins to dimerize with heavy chains and will study internalization of light chains in cell culture to determine if intracellular proteolysis is occurring. This work will increase our understanding of the mechanism of AL pathogenesis, helping us delineate better strategies for its management and cure.