Human Autoantibodies in Anti-GBM Disease

描述（由申请人提供）：抗肾小球基底膜（GBM）疾病是由抗GBM抗体介导的自身免疫性疾病。通常的临床表现是快速进展性肾小球肾炎（RPGN），导致终末期肾病。由于终生需要肾脏替代治疗，这些患者的生活质量因此显著降低。虽然目前存在关于人抗GBM疾病的抗原（Ag）α 3（IV）NCI胶原蛋白（α 3（IV））的结构基础的重要知识;但这种自身免疫性疾病的发病机制尚未完全阐明。虽然已经确定了假定的抗原表位，但对识别这些表位的致病性人抗GBM抗体（Ab）的来源、结构和构象知之甚少，本文提出的研究旨在确定抗α 3（IV）的自身抗体如何通过结合GBM引发肾炎。使用XenoMouseII（XMII）模型的人抗GBM疾病，已在我们的实验室开发，我建议产生人单克隆抗体（mAb），针对2 pupillary致病相关抗原决定簇的抗体对GBM。通常的临床表现是快速进展性肾小球肾炎（RPGN），导致终末期肾病。由于终生需要肾脏替代治疗，这些患者的生活质量因此显著降低。虽然目前存在关于人抗GBM疾病的抗原（Ag）α 3（IV）NCI胶原蛋白（α 3（IV））的结构基础的重要知识;但这种自身免疫性疾病的发病机制尚未完全阐明。虽然推定的抗原表位已被确定，很少有人知道的起源，结构和构象的致病性人类抗GBM抗体（Ab），识别这些表位。本文提出的研究旨在确定抗α 3（IV）的自身抗体如何通过与GBM结合引发肾炎。使用我们实验室开发的人抗GBM疾病的XenoMouseII（XMII）模型，我建议产生针对α 3（IV）上的2个脓毒症致病相关表位的人单克隆抗体（mAb）（目的1）。将测定这些mAb在体外结合α 3（IV）的特异性和亲和力（目的2），并将其与致病性相关联（目的3）。致病性mAb的高变区的测序将允许预测Ag结合区的三维结构。随后，这些信息应使我能够确定高变区中的哪些氨基酸残基对于与α 3（IV）（Aim 2）和产生疾病的自身抗体（Aim 3）的相互作用是必需的。这些研究有助于深入了解抗GBM疾病的基本病理生理学。此外，这些结果可能有助于设计和评估有效的治疗方法（例如，小分子）用于治疗这种毁灭性的疾病，在未来。k 08机制将使我能够进行指导的研究培训，在基础免疫学和发病机制的自身免疫性肾小球肾炎。这些研究将在宾夕法尼亚大学Michael P. Madaio博士的实验室进行。培训和研究将大大促进我的职业目标，作为一个医生/科学家在学术环境中工作。

DESCRIPTION (provided by applicant):Anti glomerular basement membrane (GBM) disease is an autoimmune disease mediated by antibodies against the GBM. The usual clinical picture is rapidly progressive glomerulonephritis (RPGN) resulting in end stage renal disease. These patients consequently suffer a significant reduction in quality of life, due to life-long need for renal replacement therapy. While significant knowledge currently exist about the structural basis of the antigen (Ag) of human anti-GBM disease, alpha3(IV) NCI collagen (alpha3(IV)); the pathogenesis of this autoimmune disease has yet to be fully elucidated. Although putative antigenic epitopes have been identified, little is known about the origin, structure and conformation of pathogenic human anti-GBM antibodies (Ab) that recognize these epitopes.The studies proposed here seek to identify how autoantibodies against alpha3(IV) initiate nephritis through binding to the GBM. Using the XenoMouseII (XMII) model of human anti-GBM disease that has been developed in our laboratory, I propose to generate human monoclonal antibodies (mAb), directed towards 2 putatively pathogenically relevant epitopes on Antibodies against the GBM. The usual clinical picture is rapidly progressive glomerulonephritis (RPGN) resulting in end stage renal disease. These patients consequently suffer a significant reduction in quality of life, due to life-long need for renal replacement therapy. While significant knowledge currently exist about the structural basis of the antigen (Ag) of human anti-GBM disease, alpha3(IV) NCI collagen (alpha3(IV)); the pathogenesis of this autoimmune disease has yet to be fully elucidated. Although putative antigenic epitopes have been identified, little is known about the origin, structure and conformation of pathogenic human anti-GBM antibodies (Ab) that recognize these epitopes. The studies proposed here seek to identify how autoantibodies against alpha3(IV) initiate nephritis through binding to the GBM. Using the XenoMouseII (XMII) model of human anti-GBM disease that has been developed in our laboratory, I propose to generate human monoclonal antibodies (mAb), directed towards 2 putatively pathogenically relevant epitopes on alpha3(IV) (Aim 1). The specificity and affinity with which these mAb bind alpha3(IV) in vitro will be determined (Aim 2), and correlated to their pathogenicity (Aim 3). Sequencing of the hypervariable region of the pathogenic mAb will allow prediction of the 3-dimensional structure of the Ag-binding region. Subsequently, this information should enable me to determine which amino acid residues in the hypervariable region are essential for the interaction with alpha3(IV) (Aim 2) and the disease producing autoAb (Aim 3). These studies should help provide insights into the basic pathophysiology of anti-GBM disease. Additionally, these results may aide in the design and evaluation of effective therapies (e.g., small molecules) for treatment of this devastating disease, in the future.The K08 mechanism will enable me to undertake mentored research training in basic immunology and the pathogenesis of autoimmune glomerulonephritis. These studies will be conducted in Dr. Michael P. Madaio's laboratory at the University of Pennsylvania. The training and studies will greatly facilitate my career goals of working as a physician/scientist in an academic setting.