Functional Analysis of NSV-based HIV vectors

描述(申请人提供)：负链RNA病毒(NSV)载体已显示出作为多种传染病疫苗的前景。虽然它们具有相似的生命周期，但NSV载体通过非常不同的机制与天然免疫相互作用并调节天然免疫，例如I型干扰素反应。VSV是一种高度细胞病变病毒，通过其M蛋白关闭细胞基因表达。狂犬病病毒(RV)是一种非细胞病变病毒，通过其P蛋白下调STAT功能。新城疫病毒是一种禽类副粘病毒，在哺乳动物细胞中似乎没有下调天然免疫的特定机制。所有这些病毒在体内都有不同程度的复制，并在树突状细胞中引发不同的宿主信号程序。由于目前尚不清楚HIV疫苗的理想载体是什么，我们现在将比较这些病毒，并提取关于它们的特定特征如何调节免疫原性的信息。对具有相似骨架和复制周期但具有非常不同的宿主-病毒相互作用集的载体诱导的免疫原性参数的全面分析尚未进行。这一应用的总体假设是，NSV载体可以通过改变它们与先天免疫系统的相互作用而得到改进。我们假设，有效的HIV-1疫苗必须激活树突状细胞(DC)，树突状细胞是天然免疫和获得性免疫之间的重要纽带。我们将首先研究这些表达HIV-1抗原的载体在体外对DC介导的免疫的影响，并确定有望在体内诱导有效的适应性免疫的参数，如有效的抗原提呈和激活。这些初步研究将使我们能够提高载体的效率，并使我们更好地基本了解什么是诱导HIV-1特异性免疫反应的最佳载体。一旦这些参数被确定，我们将在小鼠体内研究异源Prime-Boost方法中致病性和免疫原性的载体。在老鼠身上发现的最有希望的方法将在恒河猴身上进一步分析。 项目1：检查以轮状病毒为基础的改良HIV-1疫苗(密歇根州施奈尔) 项目1描述(申请人提供)：基于狂犬病病毒(RV)的疫苗载体在小鼠和恒河猴中诱导强大的细胞和体液抗HIV反应，并可在猴子中提供对类似HIV-1疾病的保护。然而，也有证据表明，这些载体可以改进。在这里，我们对RV感染的DC的激活模式进行了详细的研究，这将指导我们构建改进的RV载体。此外，我们还将分析I型干扰素对DC激活和DC迁移过程中CCR7表达的影响。我们将在小鼠模型中分析这些载体的免疫原性和致病性，将它们与新城疫病毒和水泡性口炎疫苗载体进行比较，并优先将它们用于恒河猴的研究。具体目的有两个：1.检测重组轮状病毒疫苗载体感染DC后的激活状态，分析重组轮状病毒对I型干扰素表达的影响。验证I型干扰素增加DC激活的假设。确定CCR7表达在DC迁移中的作用。特定目的2改良新冠病毒HIV-1疫苗载体在小鼠和恒河猴中的免疫。确定优化的RV载体感染后先天免疫应答对适应性抗HIV-1免疫应答的影响。测试先天免疫反应激活会降低媒介致病性的假设。测试两种病毒载体比单一载体对表达的HIV-1抗原产生更有效的反应的假设。研究优化的病毒载体在恒河猴/SIV攻击模型系统中的效率。

DESCRIPTION (provided by applicant): Negative-stranded RNA virus (NSV) vectors have shown promise as vaccines against a variety of infectious diseases. While they have similar life-cycles, NSV vectors interact with and modulate innate immunity, such as type I interferon responses, by very different mechanisms. VSV is a highly cytopathic virus that shuts down cellular gene expression by virtue of its M protein. Rabies virus (RV) is a noncytopathic virus that down-modulates STAT function by virtue of its P protein. NDV is an avian paramyxoviruses that appears not to have specific mechanisms of down-modulating innate immunity in mammalian cells. All of these viruses replicate to different extents in vivo and elicit different host signaling programs in dendritic cells. Since it is unclear what constitutes an ideal vector for HIV vaccines, we will now compare these viruses and extract information on how their specific characteristics modulate immunogenicity. A comprehensive analysis of immunogenicity parameters induced by vectors with a similar backbone and replication cycle but with very diverse set of host-virus interactions has not been performed. The overall hypothesis for this application is that NSV vectors can be improved by altering their interactions with the innate immune system. We hypothesize that an effective HIV-1 vaccine must activate dendritic cells (DCs), which are an important link between innate and adaptive immunity. We will first study the impact of these vectors expressing HIV-1 antigens on DC-mediated immunity in-vitro, and identify parameters that would be expected to induce effective adaptive immunity in-vivo, such as efficient antigen presentation and activation. These initial studies will allow us improve the efficiency of our vectors, and provide us with a better basic understanding of what constitutes an optimal vector for the induction of HIV-1-specific immune responses. Once these parameters are determined, we will study the vectors for pathogenicity and immunogenicity in heterologous prime-boost approaches in mice. The most promising approaches identified in mice will be further analyzed in rhesus macaques. PROJECT 1: Examination of modified RV-based HIV-1 vaccines (Schnell, M) PROJECT 1 DESCRIPTION (provided by applicant): Rabies virus (RV) based vaccine vectors induce potent cellular and humoral anti-HIV responses in mice and in rhesus macaques and can provide protection against an HIV-1 like disease in monkeys. However, there is also evidence that such vectors can be improved. Here we perform a detailed study of the activation pattern of RV-infected DCs, which will guide us to construct improved RV vectors. Moreover, we will analyze the impact of type I interferons on DC activation and on CCR7 expression for DC migration. We will analyze the immunogenicity and pathogenicity of such vectors in a mouse model, compare them with Newcastle Disease virus and vesicular stomatitis-based vaccine vectors, and prioritize their use for studies in rhesus macaques. Two specific Aims are proposed: Specific Aim 1. Determine the activation state of DCs after infection with RV-based HIV-1 vaccine vectors and analyze the impact of type I interferon expression by a recombinant RV. Test the hypothesis that interferon type I increases DC activation. Determine the role of CCR7 expression on DC migration. Specific Aim 2 Immunizations of improved NSV HIV-1 vaccine vectors in mice and rhesus macaques. Determine the impact of the innate immune response on the adaptive anti-HIV-1 immune response after infection with optimized RV vectors. Test the hypothesis that activation of innate immune responses reduces vector pathogenicity. Test the hypothesis that two viral vectors induce more potent responses against the expressed HIV-1 antigens than a single vector. Study the efficiency of optimized viral vectors in the rhesus macaque / SIV challenge model system.