Non-Integrating Lentiviral Vectors: Potential as Vaccines

描述（由申请人提供）：尽管在HIV-1治疗方面取得了重大进展，但HIV-1/艾滋病的流行仍在继续增长，这突出了开发有效疫苗的必要性。虽然减毒复制能力的HIV-1疫苗存在不可接受的风险，但先前对SIV减毒株的研究已经导致了关于灵长类动物模型中疫苗效力的几个重要观察结果。在病毒感染的情况下，低水平病毒抗原的慢性呈现提供了保护。在逆转录病毒感染期间，除了负责产生新病毒子代的整合前病毒外，高水平的未整合的染色体外DNA（E-DNA）在感染细胞中积累。E-DNA已被证明即使在没有可检测到的血浆病毒血症的情况下也能在体内持续存在。我们和其他人已经表明，E-DNA在体外非分裂细胞中是稳定的，并且具有转录活性，可产生功能性病毒蛋白。然而，尽管产生病毒蛋白，E-DNA并不维持病毒复制。该提议的中心假设是，从HIV-1 E-DNA持续产生病毒蛋白是表达病毒蛋白并诱导免疫应答的有效且安全的方式。病毒蛋白在感染周期的背景下呈现，具有安全特征，包括缺乏复制病毒的产生和缺乏整合。 为了验证这一假设，我们将测试整合酶缺陷型慢病毒载体在专职抗原呈递细胞（APC）（包括树突状细胞（DC））中产生病毒E-DNA和蛋白质的效率。还将评价这些转导细胞被效应T细胞识别并诱导效应T细胞应答的能力。最后，我们将使用小鼠模型来确定从肌内递送的IN缺陷型载体产生E-DNA和病毒蛋白的动力学，以及确定表达是否与可测量的免疫应答相关。这些可行性研究将确定是否应利用IN缺陷型载体进行疫苗开发。

DESCRIPTION (provided by applicant): Despite major advancements in HIV-1 therapeutics, the HIV-1/AIDS epidemic continues to grow, highlighting the need for the development of an effective vaccine. Although an attenuated replication competent HIV-1 vaccine presents unacceptable risks, prior work with attenuated strains of SIV has led to several important observations concerning vaccine efficacy in the primate model. Chronic presentation of low levels of viral antigens in the setting of a viral infection provides protection. During retroviral infection, high levels of unintegrated extrachromosomal DNA (E-DNA) accumulate in the infected cells in addition to integrated provirus responsible for production of new viral progeny. E- DNA has been shown to persist in vivo even in the absence of detectable plasma viremia. We, as well as others, have shown that E-DNA is stable in vitro in non-dividing cells and is transcriptionally active producing functional viral proteins. However, despite the production of viral protein, E-DNA does not sustain viral replication. The central hypothesis of this proposal is that sustained viral protein production from HIV-1 E-DNA is an efficient and safe way to express viral proteins and induce an immune response. Viral proteins are presented in the context of an infectious cycle with the safety features including the lack of production of replicating virus and the absence of integration. To test this hypothesis, we will test the efficiency of integrase defective lentiviral vectors to produce viral E-DNA and proteins in professional antigen presenting cells (APC) including dendritic cells (DC). The ability of these transduced cells to be recognized by and induce a response in effector T-cells will be evaluated as well. Finally we will use a murine model to determine the kinetics of E-DNA and viral protein production from IN-defective vectors delivered intramuscularly as well as determine if the expression is associated with a measurable immune response. These feasibility studies will establish whether IN-defective vectors should be exploited for vaccine development.