The Implications of Dystrophin-Specific T cells for DMD gene Correction

Ducheime肌营养不良症(DMD)小鼠和狗模型的原则验证研究证实，基因替代疗法是一种很有前途的治疗策略。试图将从临床前学到的原理应用到临床方案中，并没有预测到针对突变下游肌肉纤维上的新表位的抗肌营养不良蛋白特异性T细胞。在一个案例中，这些基因表达在突变纤维上，这一发现与预测这些纤维具有耐受作用的公理相反。另一种治疗模式，庆大霉素诱导的突变抑制，被证明同样令人困惑，因为治疗后从血液和肌肉中分离出抗肌萎缩蛋白特异的T细胞。这些观察结果需要进一步的研究才能在DMD的基因纠正策略上取得成功。在目标1中，我们将表征DMD患者血液和肌肉中Dstrophin特异性T细胞的特性，这些患者具有良好的特征突变 确定有多少患者表现出对dystrophin的细胞免疫，并确定同源自身表位在突变的dystrophin蛋白中的位置。我们将研究抗肌萎缩蛋白特异性的CD4+和CD8+T细胞的效应器功能。在目标2中，我们将特别关注糖皮质激素在调节T细胞反应中的作用，这是一项针对幼稚受试者的指定三个月治疗计划。在这里，我们预计T细胞表型从效应/炎症转变为调节/抑制作用。在目标3中，我们将使用AAVS.MCK.microdystrophin进行血管递送临床基因转移研究。我们可以通过血管传递转基因到恒河猴特定的腿部肌肉来实现高水平的肌肉纤维转导。这为临床疗效奠定了基础。这项研究的纳入标准包括目前根据先前经验确定的免疫屏障，并将增加项目1和2出现的发现。

PROJECT SUMMARY (See instructions): The Implications of Dystrophin-Speclfic T cells for DMD Gene Correction Proof-of-principle studies in mouse and dog models of Ducheime muscular dystrophy (DMD) have established that gene replacement therapy is a promising treatment strategy. Attempts to apply the tenets learned from pre-clinical to clinical protocols did not predict dystrophin-specific T cells targeting novel epitopes on muscle fibers downstream of the mutation. In one case these were expressed on revertant fibers, a finding contrary to the axiom that forecasts a tolerizing role for these fibers. Another treatment paradigm, gentamicin-induced mutation suppression, proved equally confounding because dystrophin-specific T cells were isolated fi-om the blood and muscle following treatment. These observations require further study to achieve success in gene correction strategies for DMD. In Aim 1 we will characterize the properties of dystrophin-specific T cells in the blood and muscle of DMD patients with well characterized mutations to determine how many patients exhibit cellular immunity to dystrophin and define the location of cognate selfepitopes within the mutated dystrophin protein. We will examine the effector fimctions of CD4+ and CD8+ T cells that are dystrophin specific. In Aim 2 we will look specifically at the role of glucocorticoids in modulating T cell response in a designated three-month treatment program of naive subjects. Here we anticipate a T cell phenotype change fi-om effector/inflammatory to a regulatory/suppressor role. In Aim 3 we will perform a vascular delivery clinical gene transfer study using AAVS.MCK.micro-dystrophin. We can achieve high levels of muscle fiber transduction through vascular delivery of transgene to specific leg muscles in the rhesus macaque. This sets the stage for clinical efficacy. The study inclusion criteria include currently identified immune barriers based on prior experience and will add findings that emerge from Projects 1 and 2.