IMMUNE RESPONSES TO CLYCOSYLATION-MODIFIED SIV VACCINES

人类和猿猴免疫缺陷病毒（分别为HIV和SIV） 含有高度糖基化的表面和跨膜包膜 糖蛋白 来自其他组织的糖蛋白的碳水化合物 已知来源在免疫原性中起重要作用。 然而，在这方面， 关于HIV和SIV碳水化合物部分在 对这些病毒的免疫反应。 关于此角色的信息 对设计安全有效的艾滋病疫苗很重要。 因此，本提案的目的是在恒河猴中引出 （Macaca mulatta）对糖基化修饰的完整SIV的免疫应答 疫苗，并比较这些反应的有效性， 天然糖基化SIV。 重点将放在体液反应。 病毒将在用SIVmac 251感染的H9细胞中合成。 不，不 糖基化将使用糖蛋白加工进行修饰 抑制剂，栗精胺（葡萄糖苷酶I抑制剂），1- 脱氧甘露野尻霉素（甘露糖苷酶I抑制剂）和苦马豆素（甘露糖苷酶 II抑制剂）或通过使用神经氨酸酶酶促除去唾液酸。 三种不同的高甘露糖型，非唾液酸化， 非岩藻糖基化的碳水化合物部分应该在这些 抑制剂，而神经氨酸酶治疗天然合成的病毒 应该产生复合型的去唾液酸化的碳水化合物部分。 SIV， 在细胞培养中天然合成且未经处理的，将用作 控制 将检查病毒粒子的生物活性，并且病毒 分析蛋白质的SDS-PAGE迁移率、免疫印迹反应性和 碳水化合物含量 将在2010年12月15日至16日期间， 以佐剂的形式给猕猴服用。 一般免疫反应将是 通过ELISA和Western免疫印迹监测。 特异性功能免疫 需要监测的反应是：1）。中和抗体滴度，2）。 补体介导的抗体依赖性增强活性，3）。 抗合胞抗体滴度，和4）。T细胞增殖反应。 最后，将用活SIV攻击猕猴以确定疫苗 功效 这些研究被视为更大努力的第一阶段 旨在充分解决潜在的危害和好处， 通过修饰潜在的SIV和HIV的正常糖基化而产生 疫苗。

Human and simian immunodeficiency viruses (HIV and SIV, respectively) contain heavily glycosylated, surface and transmembrane envelope glycoproteins. The carbohydrate moieties of glycoproteins from other sources are known to play significant roles in immunogenicity. However, little is known regarding the role of HIV and SIV carbohydrate moieties in the immune response to these viruses. Information regarding this role could be important to the design of a safe and effective AIDS vaccine. Therefore, the objectives of this proposal are to elicit in rhesus macaques (Macaca mulatta) immune responses to glycosylation-modified, whole SIV vaccines, and to compare the effectiveness of these responses against naturally glycosylated SIV. Emphasis will be placed on humoral responses. Virus will be synthesized in H9 cells infected with SIVmac251. N- glycosylation will be modified using the glycoprotein processing inhibitors, castanospermine (glucosidase I inhibitor), 1- deoxymannojirimycin (mannosidase I inhibitor) and swainsonine (mannosidase II inhibitor) or by enzymatic removal of sialic acids using neuraminidase. Three different molecular species of high mannose-type, non-sialylated, non-fucosylated carbohydrate moieties should arise in the presence of these inhibitors, while neuraminidase treatment of naturally synthesized virus should yield complex-type, desialylated carbohydrate moieties. SIV, naturally synthesized in cell culture and untreated, will be used as control. Virions will be examined for biological activity, and the viral proteins analyzed for SDS-PAGE mobility, immunoblot reactivity, and carbohydrate content. Psoralen/UV-inactivated whole virus vaccines will be administered in adjuvent to macaques. General immune responses will be monitored by ELISA and Western immunoblot. Specific functional immune responses to be monitored are: 1). neutralizing antibody titers, 2). complement-mediated, antibody-dependent enhancing activity, 3). antisyncytial antibody titers, and 4). T cell proliferative responses. Finally, the macaques will be challenged with live SIV to determine vaccine efficacy. These studies are viewed as the first phase of a larger effort aimed at adequately addressing potential hazards and benefits that may be created by modifying normal glycosylation of potential SIV and HIV vaccines.