Development of dendritic cell vaccine against uveitis

葡萄膜炎（眼内炎性疾病）是美国约10%严重视力障碍的原因。目前的治疗是基于局部或全身皮质类固醇，有或没有二线药物，如环孢素A或抗代谢药。这些药物的严重不良反应是开发毒性更小和更特异性的葡萄膜炎治疗方法的动力。实验性自身免疫性葡萄膜炎（EAU）是一种特征良好的人类葡萄膜炎动物模型，对猴和啮齿动物EAU的研究已经鉴定出视网膜蛋白S-抗原和IRBP作为葡萄膜炎的推定自身抗原。人们对开发针对葡萄膜炎的治疗性疫苗非常感兴趣，特别强调对S-抗原、IRBP和/或其他推定的葡萄膜原（恢复蛋白、视蛋白）的耐受性诱导。我们先前表明STAT通路的激活是发育调节的，并且在树突状细胞（DC）分化和成熟中起作用。例如，STAT 6信号传导途径在前体DC（pDC）和未成熟DC（iDC）中被组成性激活，但随着这些DC分化成成熟DC（mDC）而下降，并且STAT 6激活的下降与SOCS蛋白的上调相关（J Immunol.172：2307-15）。相比之下，STAT 1信号传导促进DC成熟，并且在mDC中最稳健。然而，与STAT 6通路不同，STAT 1信号传导不受SOCS蛋白的反馈调节，表明STAT 1和STAT 6通路在发育和成熟DC中受到不同的调节。因此，STAT 1和STAT 6似乎分别是mDC和iDC的谱系标志物。在本财政年度，我们的工作重点是利用树突状细胞群中STAT蛋白的差异利用，通过特异性删除DC制剂中的STAT 1基因来产生高度纯化的iDC。因此，我们已经产生了几种siRNA构建体，并正在分析siRNA技术沉默STAT 1表达或通过沉默SOCS基因上调STAT 6的效率和可行性。我们的最终目标是用IRBP或S抗原的免疫致病性表位负载纯化的iDC，以用作针对葡萄膜炎的治疗性疫苗。

Uveitis (intraocular inflammatory diseases) is the cause of approximately 10% of severe visual handicap in the United States. Current therapy is based on topical or systemic corticosteroid, with or without second line agents such as cyclosporine A or anti-metabolites. Serious adverse effects of these drugs are the impetus for development of less toxic and more specific therapies for uveitis. Experimental autoimmune uveitis (EAU) is a well-characterized animal model of human uveitis and studies of monkey and rodent EAU have led to identification of the retinal proteins, S-Antigen and IRBP, as putative autoantigen of uveitis. There is significant interest in developing a therapeutic vaccine against uveitis, with particular emphasis on tolerance induction to S-Antigen, IRBP and/or other putative uveitogens (recoverin, opsin). We previously showed that activation of STAT pathways is developmentally regulated and plays a role in dendritic cell (DC) differentiation and maturation. For example, the STAT6 signaling pathway is constitutively activated in precursor DCs (pDCs) and immature DCs (iDCs) but declines as these DCs differentiate into mature DCs (mDCs) and the decline in STAT6 activation correlates with upregulation of SOCS proteins (J Immunol. 172:2307-15). In contrast, STAT1 signaling promotes DC maturation and is most robust in mDCs. However, unlike the STAT6 pathway, STAT1 signalling is not under feedback regulation by SOCS proteins, indicating that STAT1 and STAT6 pathways are distinctly regulated in developing and mature DC. Thus, STAT1 and STAT6 appear to be lineage markers of mDCs and iDCs, respectively. In this fiscal year the thrust of our work has been to exploit the differential utilization of STAT proteins in dendritic cell populations to generate highly purified iDC by specifically deleting STAT1 gene in DC preparations. We have therefore generated several siRNA constructs and are analyzing the efficiency and feasibility of siRNA technology to silence expression of STAT1 or upregulate STAT6 by silencing SOCS genes. Our ultimate goal is to load the purified iDCs with immunopathogenic epitopes of IRBP or S-Antigen for use as therapeutic vaccines against uveitis.