Influenza HA in SHIV VLPS for mucosal vaccination

描述（由申请人提供）：最近，我们通过使用杆状病毒表达系统成功地生产了含有SIV Gag和HIV Env的猿-人免疫缺陷病毒样颗粒（SHIV VLP）。此外，我们还将流感病毒表面糖蛋白血凝素（HA）整合到SHIV VLP中。利用HA与上呼吸道粘膜结合的高亲和力，我们的中心假设是使用含有流感HA的SHIV VLP作为粘膜疫苗将增强针对HIV感染的全身和粘膜免疫应答。本项目的主要目的是研究HA在SHIV VLP中的内在佐剂性在小鼠模型中用于鼻内免疫的效率和机制。具体而言，我们建议：1）。确定在SHIV VLP中掺入流感HA作为粘膜疫苗在增强针对HIV的免疫应答中的作用。提出的实验将研究：a）作为粘膜疫苗的SHIV VLP中流感HA的内在佐剂性是否比可溶性流感HA在增强全身和粘膜免疫方面更有效;和B）。流感HA的受体结合或膜融合活性是否影响其对SHIV VLP的佐剂性。2）。确定SHIV VLP中流感HA的掺入在树突状细胞（DC）结合、活化、细胞因子产生和抗原呈递中的作用。我们建议调查：a）。HA/SHIV VLP是否具有增加的DC结合、内化和亚细胞定位的能力; B）. HA/SHIV VLP是否对DC活化和细胞因子产生具有增加的作用; c）. HA/SHIV VLP活化的DC是否增加初始T细胞增殖;和d）. HA/SHIV VLP是否增加DC向CD 8 + T细胞的抗原交叉呈递的效率以及相关的细胞内途径是什么。3）。确定在没有CD 4 + T细胞帮助的情况下，流感HA在SHIV VLP中的掺入在特异性B细胞结合、活化和抗体产生中的作用。实验旨在研究：a）。HA/SHIV VLP是否增加它们与幼稚B细胞结合的能力; B）. HA/SHIV VLP是否对幼稚B细胞增殖具有增加的作用; c）. HA/SHIV VLP活化的DC是否具有增加的粘附至幼稚B细胞的能力; d）. HA/SHIV VLP活化的DC是否增加幼稚B细胞增殖;和e）. B细胞活化和分化以及细胞毒性CD 8+形成是否发生在鼻相关淋巴组织（NALT）中，NALT是CD 4 + T细胞缺陷小鼠中HA/SHIV VLP鼻内免疫后的诱导位点。该项目代表了开发有效和安全的艾滋病毒疫苗的新方法。 了解HA/SHIV VLP增强小鼠免疫应答的细胞和分子机制对于未来在非人灵长类动物模型和人体试验中成功设计和测试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.