临床数据表明重组型腺相关病毒载体（rAAV）衣壳蛋白能引起炎症反应和杀伤性T细胞反应，显著降低基因治疗疗效。由于不清楚rAAV靶向哪些免疫细胞，过去的研究集中于获得性免疫应答，而对先天性免疫反应知之甚少。近年来，我们以及他人的工作成果均表明rAAV能够高效感染树突状细胞。进一步，我们的预试验结果提示rAAV能显著提高免疫调控因子糖皮质激素受体（GR）的磷酸化与核定位。因此，本项目将以树突状细胞为研究对象，探索通过GR信号通路抑制rAAV先天性免疫反应的详细分子机制。首先，我们将通过共定位、分子克隆等方法研究衣壳蛋白不同结构域激活GR的作用。并以此改造病毒载体，使之更有效地激活GR。其次，我们将在体内外实验中通过芯片技术等高通量方法阐明由rAAV激活的GR对下游靶基因表达的影响，以及与其它免疫相关信号通路的对话。综上，我们的研究将为临床用药降低rAAV免疫应答提供理论基础和实验依据。

Recombinant adeno-associated viral vectors (rAAV) have been used in a number of gene therapy clinical trials. Inflammatory responses have been reported in those trials that were triggered by the rAAV capsid. These responses induced cytotoxic T cell activity and eventually reduced gene therapy efficacy. Due to the lack of information about the immune cells targeted by rAAV, previous research focused on the adaptive immune responses. On the other hand, the mechanism of innate immune responses is largely unclear. In recent years, we and others documented that rAAV can efficiently transduce dendritic cells. Furthermore, our pilot experiment suggested that rAAV transduction can significantly enhance the phosphorylation and nuclear translocation of glucocorticoid receptor (GR), a well-known innate immune regulator. Thus, the current proposal aims to explore the detailed molecular mechanisms using dendritic cells. First, through techniques such as sub-cellular co-localization, molecular cloning, etc, we will study various rAAV capsid domains to demonstrate their ability of GR activation. The capsid will then be modified accordingly through site-specific mutagenesis, in order to further increase the GR activator function. Second, we plan to reveal the effect of rAAV-activated GR on the expression of its downstream target genes, both in vitro and in vivo, using high-throughput techniques such as Chip assays. In addition, the cross talk between GR and other immune-related signal pathways will also be investigated. To summarize, our proposed study will provide not only the answers of fundamental virological questions, but also the pre-clinical data for future gene therapy clinical trials.