In situ gene-modified DCs for an HIV-1 T cell vaccine

描述（由申请人提供）：尽管已经开发了针对许多病毒源性传染病的保护性疫苗接种策略，但二十年来，对HIV-1具有广泛活性的疫苗仍然难以捉摸。 细胞介导的免疫已经涉及（i）HIV-1复制的调节，（ii）疾病进展的延长，和（iii）针对感染的保护，论证了它们作为成功免疫策略的组成部分的重要性。 树突状细胞（DC）可以通过MHC I类和II类途径呈递抗原，并且是细胞介导的免疫应答的发展的中心。 因此，目前正在开发基于DC的疫苗作为引发强烈的抗原特异性T细胞应答的试剂，但这些方案通常需要DC的离体遗传修饰，然后再输注回自体或MHC匹配的宿主，在那里它们成熟，迁移到淋巴结，并将候选抗原呈递给CD4和/或CD8 T细胞以引发抗原特异性应答。 虽然离体遗传操作方法在实验环境中建立概念验证方面是有用的，但对于开发用于该领域的预防性疫苗是不切实际的。 如果DC的转导可以在原位完成，DC靶向疫苗作为预防剂的广泛使用将是更可行的。 然而，原位转导的关键组成部分是载体有效靶向DC而不干扰DC功能。 为此，我们已经产生了一种腺病毒（Ad）载体系统，通过CD40受体特异性靶向DC，并已表明，CD40靶向的Ad有效地转导DC在体外不干扰DC功能。 原位转导的一个潜在限制是足够数量的DC细胞必须在疫苗接种部位以实现有效免疫。 因此，我们建议通过使用一种新的技术，释放趋化因子的聚合物棒，在CD40靶向疫苗接种之前将DC招募到皮下区域。 在这个提议中，我们将评估CD40靶向的Ad载体表达HIV-1抗原在小鼠体外和原位疫苗接种模型，纳入化学引诱剂释放聚合物的效力。 首先，我们将优化原位DC募集的参数，并确定这在多大程度上有助于原位CD40靶向转导的效率。 接下来，在存在或不存在DC募集的情况下，用非靶向和靶向CD40的Ad载体免疫后，将比较抗原特异性CD4和CD8 T细胞应答。 我们的假设是，CD40靶向的Ad将最佳地在皮下募集到免疫部位后使DC活化，导致诱导稳健的T细胞免疫。 能够实现靶向原位免疫的DC，随后诱导强大的抗原特异性T细胞免疫将是HIV-1疫苗开发的里程碑。 这项研究的成功将推动未来在非人灵长类动物模型中进行体内研究，以显示保护性免疫力和HIV-1感染高危人群的人体临床试验。

DESCRIPTION (provided by applicant): Although protective vaccination strategies have been developed against many infectious diseases of viral origin, a vaccine with broad activity against HIV-1 has remained elusive for two decades. Cell-mediated immunity has been implicated in (i) modulation of HIV-1 replication, (ii) protraction of disease progression, and (iii) protection against infection, arguing their importance as components of a successful immunization strategy. Dendritic cells (DC) can present antigens via MHC class I and II pathways and are central to the development of cell-mediated immune responses. Thus, DC-based vaccines are currently being developed as agents to elicit strong, antigen-specific T cell responses, but these protocols usually require ex vivo genetic modification of DCs, followed by re-infusion back into an autologous or MHC-matched host where they mature, migrate to lymph nodes, and present the candidate antigen to CD4 and/or CD8 T cells to elicit antigen-specific responses. Although the ex vivo genetic manipulation approach has been useful in establishing proof-of-concept in an experimental setting, it is not practical for the development of prophylactic vaccines to be used in the field. If transduction of the DCs could be accomplished in situ, the broad use of DC-targeted vaccines as prophylactic agents would be more feasible. However, a critical component of in situ transduction is efficient targeting of the vector to DCs without perturbation of DC function. To this end, we have generated an adenovirus (Ad) vector system that specifically targets DCs via the CD40 receptor and have shown that the CD40-targeted Ad efficiently transduces DCs in vitro without interfering with DC function. A potential limitation of in situ transduction is that sufficient numbers of DC cells must be at the vaccination site to achieve effective immunization. Therefore, we propose to recruit DCs into subcutaneous locales prior to CD40-targeted vaccination by using a novel technology, chemoattractant-releasing polymer rods. In this proposal, we will evaluate the potency of CD40-targeted Ad vectors that express HIV-1 antigens in murine in vitro and in situ vaccination models that incorporate chemoattractant-releasing polymers. Initially, we will optimize the parameters of in situ DC recruitment and determine the degree to which this contributes to the efficiency of in situ CD40-targeted transduction. Next, antigen-specific CD4 and CD8 T cell responses will be compared following immunization with untargeted and CD40-targeted Ad vectors in the presence or absence of DC recruitment. Our hypothesis is that the CD40-targeted Ad will optimally transduce DCs following subcutaneous recruitment to the immunization locale, resulting in the induction of robust T cell immunity. The ability to achieve targeted in situ immunization of DCs followed by the induction of robust antigen-specific T cell immunity will be a milestone in HIV-1 vaccine development. The success of this study will be the impetus for future in vivo studies in non-human primate models to show protective immunity and human clinical trials for populations at high risk of HIV-1 infection.