Mechanisms of Aggregation in Light Chain Amyloidosis

描述（由申请人提供）：我们提出的研究的总体目标是剖析导致轻链淀粉样变性（AL）沉积物形成的分子机制。 AL 是一种由血浆 B 细胞异常增殖引起的破坏性疾病。这些肿瘤细胞分泌单克隆免疫球蛋白轻链，这些轻链作为截短的轻链错误折叠成重要器官中的淀粉样蛋白沉积物，导致组织损伤和器官衰竭。免疫球蛋白可变域中积累的体细胞突变使这些蛋白质的热力学稳定性低于其非淀粉样变蛋白的对应物。免疫球蛋白轻链和重链均在成熟的血浆 B 细胞中产生，它们在其中结合形成分泌到循环中的抗体。在大多数 AL 患者中，即使重链存在于浆细胞中，也只有轻链被分泌到循环中，这表明无法与重链结合。关于 AL 的许多问题仍未得到解答。目前尚不清楚是什么原因导致与重链、细胞区室或事件的关联性丧失，从而导致蛋白质截短，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，尚未进行全面的研究来了解突变位置在 AL 中蛋白质折叠、稳定性和淀粉样蛋白生成性中的作用。我们假设 AL 蛋白不稳定是由于以下一种或多种事件触发的部分未折叠状态的采样所致：体细胞突变、免疫球蛋白重链解离和/或蛋白水解裂解。这项工作的目标 1 将研究可变域、截短和全长 AL 蛋白的结构和稳定性。我们还将对种系残基进行 AL 突变的系统恢复，以确定导致淀粉样蛋白形成倾向的突变。目标 2 将表征 AL 蛋白形成的淀粉样蛋白。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。目标 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，并将研究细胞培养物中轻链的内化以确定是否发生细胞内蛋白水解。这项工作将增进我们对 AL 发病机制的了解，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.