EVALUATION OF HIV GAG DNA VACCINE TO MAX T HELPER/CTL

描述：（改编自申请人的摘要）我们的总体目标是 确定CD 8+细胞毒性T淋巴细胞（CTL）和CD 4 + T细胞 可以针对高度保守的内部病毒体引发辅助应答 HIV-1 Gag蛋白为了实现这一目标，一个新的发展是， 构建DNA表达载体，使我们能够在Rev RRE独立和物种独立。使用这种新方法，我们 将系统地评估表达，和免疫反应， 不同的亚细胞靶向形式的HIV-1 Gag，以最大限度地诱导 小鼠对Gag的CTL和辅助性T细胞应答。 我们将实现两个具体目标：（1）。能力的表征 各种HIV-1 Gag表达载体诱导CD 8 + CTL。HIV-1 Gag表达 这些载体使我们能够在小鼠细胞中表达HIV-1 Gag分子， 已生成Rev和RRE缺失。衍生的Gag表达载体， 其靶向于不同的细胞内和细胞外区室 用于抗原加工和呈递的细胞将被构建和评估。 这些不同形式的Gag包括：（a）野生型Gag，其组装成 （B）从细胞中有效释放的突变体Gag， 靶向质膜并保留在细胞质中;（c）突变型Gags， 未能组装成颗粒;（d）快速降解的突变型Gags 细胞内，（e）分泌但非颗粒形式的Gag;（f）细胞表面 加格的固定形式。(A)各种形式的HIV-11 Gag分子将被 在p815细胞中表达并作为CTL靶细胞进行比较。(B)DNA载体 将对表达各种形式的HIV-1 Gag分子的人进行CTL检测 小鼠中的诱导。（二）、各种HIV-1 Gag的能力表征 将优化T辅助应答的表达载体，特别是那些 可以增强CTL反应。(A)不同形式的HIV Gag DNA疫苗 还将评价上述的免疫抑制剂对T辅助应答的诱导 DNA免疫小鼠后，通过测量T细胞增殖和细胞因子 体外Gag刺激后的反应。(B)CTLA-4-HIV-1 Gag融合体 将评估由单个DNA疫苗载体编码的分子。这 该策略将允许Gag分子特异性靶向抗原呈递 细胞的MHC 11类呈递。(C)使用DNA的组合研究 从该特定目的鉴定的载体与从 具体目标1.

DESCRIPTION: (Adapted from Applicant's Abstract) Our overall goal is to determine the mechanisms by which CD8+ cytotoxic T lymphocyte (CTL) and CD4+ T helper responses can be elicited against highly conserved internal virion proteins of HIV-1 Gag. Toward this goal, a novel development is the construction of DNA expression vectors, which enable us to express Gag in a Rev and RRE independent and species independent fashion. Using this new method, we will systematically evaluate the expression of, and immune response to, different subcellularly targeted forms of HIV-1 Gag to maximize the induction of CTL and T helper responses against Gag in mice. We will carry out two specific aims: (1). Characterization of the ability of various HIV-1 Gag expression vectors to induce CD8+ CTL. HIV-1 Gag expression vectors which enable us to express HIV-1 Gag molecules in mouse cells in the absence of Rev and RRE have been generated. Derivative Gag expression vectors, which are targeted to different intracellular and extracellular compartments for antigen processing and presentation will be constructed and evaluated. These different forms of Gag include: (a) wild type Gag that assembles into particles which are released from cells efficiently; (b) mutant Gags that fail to target to plasma membrane and remain in the cytoplasm; (c) mutant Gags that fail to assemble into particles; (d) mutant Gags that are rapidly degraded intracellularly, (e) secreted but non-particle forms of Gag; (f) cell surface anchored forms of Gag. (A) Various forms of HIV-11 Gag molecules will be expressed in p815 cells and compared as CTL target cells. (B) DNA vectors expressing various forms of HIV-1 Gag molecules will be tested for CTL induction in mice. (2). Characterization of the ability of various HIV-1 Gag expression vectors that will optimize T helper response especially those that could enhance CTL response. (A) The different forms of HIVA Gag DNA vaccine described above will also be evaluated for induction of T helper responses after DNA immunization in mice by measuring T cell proliferative and cytokine responses after Gag stimulation in vitro. (B) A CTLA-4-HIV-1 Gag fusion molecule encoded by a single DNA vaccine vector will be evaluated. This strategy will allow specific targeting of Gag molecules to antigen presentation cells for MHC class 11 presentation. (C) Combination studies by using DNA vectors identified from this specific aim with DNA vectors identified from specific aim 1.