2D affinity and frequency of antigen specific Tregs

 描述(由申请人提供)：调节性T细胞(Treg)对于适当控制免疫反应至关重要，并在自身免疫性疾病的调节中发挥明显的作用。Tregs进一步强调了它们的重要性，它提供了一种抗原特异性疗法，目前正在临床前和临床试验中进行评估。尽管它们具有毋庸置疑的价值，但很难评估Treg抗原的特异性。事实上，许多实验方案利用非特异性TCR刺激，如抗CD3/抗CD28来激活它们，而典型的特异性评估，如肽：MHC II类四聚体和功能读数在应用于Tregs时并不是特别有效。由于TCR的特异性和亲和力对任何T细胞功能反应都是至关重要的，我们需要了解什么亲和力范围最有利于Treg的最佳使用以及它们的分化。我们的实验室在检验问题方面具有独特的地位。 由于我们已经开发了一种基于二维(2D)微吸管技术的新型检测系统，该系统提供了目前可用于评估这些参数的最灵敏的手段。关于胸腺来源的Tregs(TTreg)亲和力的流行观点是，它们的谱系由亲和力最高的TCR组成 负选择；然而，我们的初步数据已经确定了髓鞘少突胶质细胞糖蛋白(MOG)特异性tTregs的一系列亲和力。MOG支持胸腺来源的MOG特异性Tregs的发展，而不是驱动胸腺细胞的负选择。我们认为这些Treg将用于治疗，并在我们的初步研究的基础上，提出以下两个目标来追踪Treg在慢性和复发/缓解性脱髓鞘疾病中的特异性、位置、动力学、稳定性和效力。目的1：确定脱髓鞘疾病中MOG特异性Treg的亲和力和频率-目标2：确定MOG的Treg特异性和谱系稳定性。

 DESCRIPTION (provided by applicant): Regulatory T cells (Tregs) are critical for proper control of the immune response and play a clear role in regulation of autoimmune disease. Further highlighting their importance, Tregs provide an antigen specific therapy that is currently being evaluated in preclinical and clinical trials. Although they are of unquestioned value, it is difficult to assess Treg specificity for antigen. In fact, many experimental protocols make use of non-specific TCR stimuli such as anti-CD3/anti-CD28 for their activation, and typical assessments of specificity such as peptide: MHC Class II tetramers and functional readouts are not especially effective when applied to Tregs. As TCR specificity and affinity are crucial to any T cell functional response, we need to understand what affinity range works best for optimal Treg use as well as for their differentiation. Our lab is in a unique position to examine questions on TCR specificity and affinity because we have developed a novel assay system based on two dimensional (2D) micropipette technology that provides the most sensitive means currently available to assess these parameters. The prevalent view of thymic derived Tregs (tTreg) in terms of affinity is that their repertoires are comprised of the highest affinity TCRs that escaped negative selection; however, our preliminary data has identified a range of affinities of tTregs specific for myelin oligodendrocyte glycoprotein (MOG). Instead of driving thymocyte negative selection, MOG supports development of thymic derived MOG specific Tregs. We propose that these Tregs will be of therapeutic use and based on our preliminary studies, propose the following two aims to track Treg specificity, location, kinetics, stability, and potency during chronic and relapsing/remitting demyelinating disease. Aim 1: To define MOG-specific Treg affinity and frequency during demyelinating disease- Aim 2: To determine Treg specificity and lineage stability for MOG-