IMMUNE RESPONSES TO CLYCOSYLATION-MODIFIED SIV VACCINES

人类和猿猴免疫缺陷病毒(分别为艾滋病毒和SIV) 含有高度糖基化的表面和跨膜包膜 糖蛋白。其他来源糖蛋白的碳水化合物部分 众所周知，来源在免疫原性中起着重要作用。然而， 关于HIV和SIV碳水化合物的作用，人们知之甚少 对这些病毒的免疫反应。有关此角色的信息 可能对设计安全有效的艾滋病疫苗很重要。 因此，这项提案的目标是诱导恒河猴 (猕猴)对糖基化修饰的完整SIV的免疫应答 疫苗，并将这些反应的有效性与 天然糖基化的SIV。重点将放在幽默回答上。 病毒将在感染SIVmac251的H9细胞中合成。vt.N- 糖基化将使用糖蛋白处理进行修饰 葡甘香胺(葡萄糖苷酶I抑制剂) 脱氧甘露糖苷酶I抑制剂和苦马豆素(甘露糖苷酶 II抑制剂)或使用神经氨酸酶酶法去除唾液酸。 三种不同的高甘露糖型、非唾液酸型、 非岩藻糖化的碳水化合物部分应在以下情况下出现 抑制剂，神经氨酸酶治疗自然合成病毒 应该产生复合型的、所需的碳水化合物部分。SIV， 在细胞培养中自然合成的未经处理的，将被用作 控制力。将对病毒粒子进行生物活性检查，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。补骨脂素/紫外线灭活全病毒疫苗将 作为猕猴的佐剂给药。一般免疫反应将是 用ELISA法和免疫印迹法检测。特异性功能免疫 需要监测的反应有：1)。中和抗体效价，2)。 补体介导的、抗体依赖的增强活性，3)。 抗合胞抗体效价，4)。T细胞增殖反应。 最后，将用活SIV对猕猴进行挑战，以确定疫苗 功效。这些研究被视为更大努力的第一阶段 旨在充分解决可能存在的潜在危险和好处 通过修改潜在的SIV和HIV的正常糖基化而创建 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.