Influenza HA in SHIV VLPS for mucosal vaccination

描述(申请人提供)：最近，我们利用杆状病毒表达系统成功地生产出含有SIV Gag和HIV Env的猿人免疫缺陷病毒样颗粒(SIV VLP)。此外，我们还将流感病毒表面糖蛋白血凝素(HA)整合到SIV VLP中。我们的中心假设是，利用HA与上呼吸道粘膜具有高亲和力的优势，使用含有流感HA的Shiv VLP作为粘膜疫苗将增强系统和粘膜对HIV感染的免疫应答。本研究的主要目的是在小鼠鼻腔免疫模型中研究SHIV VLP中HA的内在佐剂作用的有效性和机制。具体而言，我们提出：1)。目的：研究猪传染性支气管炎病毒VLP与流感血凝素(HA)结合作为黏膜疫苗在增强抗HIV免疫应答中的作用。拟议的实验将调查：a)流感病毒HA作为粘膜疫苗的内置佐剂作用是否比可溶性流感HA在增强系统和粘膜免疫方面更有效；以及b)流感病毒HA的受体结合或膜融合活性影响其对新城疫病毒VLP的佐剂活性。2)。目的：研究与流感病毒血凝素(HA)结合在SHIV VLP中对树突状细胞(DC)结合、激活、细胞因子产生和抗原提呈的作用。我们建议调查：a)。HA/SIV VLP是否具有增强的DC结合、内化和亚细胞定位能力；b)。HA/SIV VLP对DC的激活和细胞因子的产生是否有增加作用；HA/SIV VLP激活的DC是否能促进初始T细胞的增殖；HA/SIV VLP是否能增加DC与CD8+T细胞的抗原交叉提呈效率，以及相关的细胞内途径。3)。目的：在没有CD4+T细胞帮助的情况下，确定在SIV VLP中与流感血凝素(HA)结合在特异性B细胞结合、激活和抗体产生中的作用。实验旨在调查：a)。HA/SIV VLP是否增强其与初始B细胞的结合能力；b)。HA/SIV VLP对幼稚B细胞增殖是否有促进作用；HA/SIV VLP激活的DC是否具有更强的黏附幼稚B细胞的能力；HA/SIV VLP激活的DC是否促进初始B细胞的增殖；鼻腔相关淋巴组织(NALT)中是否存在B细胞的活化和分化以及细胞毒性CD8+的形成，鼻腔相关淋巴组织(NALT)是CD4+T细胞缺陷小鼠鼻内接种HA/SHV VLP后的诱导部位。该项目代表了一种开发有效和安全的艾滋病毒疫苗的新方法。了解HA/Shiv VLP增强小鼠免疫反应的细胞和分子机制对于未来在非人类灵长类动物模型和人类试验中设计和测试成功的HIV疫苗至关重要。

DESCRIPTION (provided by applicant): Recently, we have successfully produced simian-human immunodeficiency virus-like particles (SHIV VLPs) which contain SIV Gag and HIV Env by using a baculovirus expression system. Furthermore, we have incorporated the influenza virus surface glycoprotein, hemagglutinin (HA), into SHIV VLPs. Taking advantage of HA having a high affinity to bind to the mucosa of the upper respiratory track, our central hypothesis is that use of SHIV VLPs containing influenza HA as a mucosal vaccine will enhance both systemic and mucosal immune responses against HIV infection. The major focus of this project is to investigate the efficiency and mechanisms of the built-in adjuvanticity of HA in SHIV VLPs for intranasal immunization in a mouse model. Specifically, we propose: 1). To determine the role of incorporation with influenza HA in SHIV VLPs as a mucosal vaccine in enhancement of immune responses against HIV. Proposed experiments will investigate: a) whether the built-in adjuvanticity of influenza HA in SHIV VLPs as a mucosal vaccine is more potent than soluble influenza HA in enhancement of both systemic and mucosal immunity; and b). whether the receptor binding or membrane fusion activity of influenza HA affects its adjuvanticity for SHIV VLPs. 2). To determine the role of incorporation with influenza HA in SHIV VLPs in dendritic cell (DC) binding, activation, cytokine production, and antigen presentation. We propose to investigate: a). whether HA/SHIV VLPs have an increased ability to DC binding, internalization, and subcellular localization; b). whether HA/SHIV VLPs have an increased effect on DC activation, and cytokine production; c). whether HA/SHIV VLP-activated DCs increase naive T cell proliferation; and d). whether HA/SHIV VLPs increase the efficiency of antigen cross-presentation of DCs to CD8+ T cells and what are the associated intracellular pathways. 3). To determine the role of incorporation with influenza HA in SHIV VLPs in specific B cell binding, activation, and antibody production without CD4+ T cell help. Experiments are designed to investigate: a). whether HA/SHIV VLPs increase their ability to bind to naive B cells; b). whether HA/SHIV VLPs have an increased effect on naive B cell proliferation; c). whether HA/SHIV VLP-activated DCs have an increased ability to adhere to naive B cells; d). whether HA/SHIV VLP-activated DCs increase naive B cell proliferation; and e). whether B cell activation and differentiation and cytotoxic CD8+ formation occur in nasal-associated lymphoid tissue (NALT), an inductive site after intranasal immunization with HA/SHIV VLPs in CD4+ T-cell-deficient mice. This project represents a novel approach to develop an effective and safe HIV vaccine. Understanding the cellular and molecular mechanisms of HA/SHIV VLP-enhanced immune responses in mice is critical for the future design and testing of a successful HIV vaccine in non-human primate models and in human trials.