DELIVERY OF BIOPOLYMERS USING CATIONIC PEPTIDES

这个提议的长期目标是使用短的、阳离子的 多肽作为一种有效的生物聚合物输送方法，如完整的 自身抗原、免疫优势肽和反义寡核苷酸 进入抗原提呈细胞和T淋巴细胞的细胞质中 诱导抗原特异性无能和/或影响分泌物的特征 细胞因子用于实验性变态反应性脑脊髓炎(EAE)的治疗。 我们已经证明了一个肽的共轭，对应于九个 从HIV Tat蛋白到蛋白质抗原的氨基酸导致 它们被各种细胞快速摄取，并允许分子 进入MHC第一类和第二类生物合成途径，结果是 它们的抗原性和免疫原性显著增加。在……里面 此外，除了将抗原输送到MHC I类和11类分子外，我们还 建议使用TAT多肽将PNA反义试剂转移到 调节炎症特征的细胞因子的产生。这个 该项目中概述的研究试图探索治疗 特异性抑制促炎细胞因子表达的可能性 通过为转录激活因子NF-kappaB引入反义PNA 转化为小鼠T细胞克隆。如果成功，这些方法可以被用作 小鼠体内减轻中枢炎症反应的治疗策略 老鼠的世界。这个项目有三个具体目标：1.证明 TAT多肽与完整髓鞘碱性蛋白的偶联 蛋白脂蛋白及其免疫优势决定簇和 改变的多肽配体极大地增强了它们诱导 抗原特异性耐受性和/或修改细胞因子谱 自身抗原特异性克隆，2.证明Tat结合蛋白 多肽进入抗原提呈细胞，如树突状细胞和 小的静息B细胞，并探索是否通过移植 抗原负载细胞更有效地诱导免疫刺激 或耐受性优于用多肽或蛋白质免疫，以及3。 证明多种促炎蛋白的表达 使用HIV-1 TAT标记的反义PNA可显著降低 体外抗核因子-kappaB的65kodalton亚基 反义PNA构建物直接注射可减轻炎症反应 变成患有流行性乙型脑炎的啮齿动物。

The long term objective of this proposal is to use short, cationic peptides as a efficient method of delivering biopolymers, such as intact autoantigens, immunodominant peptides, and antisense oligonucleotides into the cytoplasm of antigen presenting cells and T lymphocytes to induce antigen specific anergy and/or affect the profile of secreted cytokines as a therapy for experimental allergic encephalomyelitis (EAE). We have demonstrated that conjugation of a peptide, corresponding to nine amino acids from the HIV tat protein, to protein antigens results in their rapid uptake by a variety of cells and permits the molecules to enter both the MHC class I and ll biosynthetic pathways, which results in a dramatic increase in their antigenicity and immunogenicity. In addition, to delivering antigens to the MHC class I and ll molecules, we propose to use the tat peptide to transport PNA antisense reagents into cells to modulate cytokine production characteristic of inflammation. The studies outlined in this project seek to explore the therapeutic potential of specifically suppressing proinflammatory cytokine expression by introducing antisense PNA for the transcriptional activator NF-kappaB into murine T cell clones. If successful, these methods could be used as therapeutic strategies in mice in vivo to reduce inflammation in the CNS of mice. This project has three specific aims, 1. demonstrate that conjugation of the tat peptide to intact myelin basic protein and proteolipid protein as well as their immunodominant determinants and altered peptide ligands dramatically increases their ability to induce antigen specific tolerance and/or modify the cytokine profile of the autoantigen specific clones, 2. demonstrate that tat conjugated proteins and peptides enter antigen presenting cells, such as dendritic cells and small resting B cells, and explore whether adoptive transfer of the antigen loaded cells more efficiently induces either immune stimulation or tolerance than immunization with the peptides or proteins, and 3. demonstrate that the expression of a variety of proinflammatory proteins can be dramatically reduced with HIV-1 tat conjugated antisense PNA against the 65 kilodalton subunit of NF-kappaB in vitro and that the antisense PNA constructs can reduce inflammation when directly injected into rodents with EAE.