Induction of mucosal SIV immunity in non human primates by secreted Hsp-Gp96

描述（由申请人提供）：细胞免疫和记忆是清除病毒和防止病毒感染所必需的。CD8 CTL和NK细胞的细胞免疫是通过激活先天免疫反应、树突状细胞（DC）成熟和抗原向CD8 T细胞交叉递呈而启动的。刺激细胞免疫的抗病毒疫苗必须模仿这一过程。刺激抗体反应的佐剂被有效地使用，但是我们对佐剂刺激CTL免疫的知识是有限的。我们已经证明，内质网驻留热休克蛋白gp96是一种由MHC-I转运到呈递的肽的伴侣，是激活DC、NK和同源CD8 T细胞的天然佐剂。DC细胞和巨噬细胞具有gp96肽复合物的受体，并在gp96结合时被激活。Gp96及其相关肽被APC吸收，肽段被运送到内质网并用于给mhc - 1分子充电。与DC摄取的完整蛋白交叉启动相比，DC的伴随激活导致同源CD8 T细胞的交叉启动增强超过百万倍。为了利用这种独特的佐剂作用和转运相关肽的能力，我们通过用lgG1的Fc部分取代KDEL er保留信号，制备了一种可分泌的gp96， gp96-lg。我们假设基于细胞的gp96-lg疫苗通过在体内长时间分泌gp96-lg肽，模拟病毒复制并提供与减毒病毒相当的免疫刺激。在小鼠模型系统中，我们已经证明gp96-lg转染，表达抗原的肿瘤细胞在体内分泌gp96-lg，刺激先天DC和NK以及适应性同源细胞CD8 CTL免疫反应，并在没有淋巴结的情况下独立于CD4帮助产生特异性CD8记忆。gp96-lg疫苗在上皮内、固有层和Peyer’s patch中产生全身和强粘膜免疫CD8 CTL。由于其独特的性质，我们现在计划在非人类灵长类动物模型中评估gp96-疫苗对SIV粘膜和全身细胞免疫的免疫原性（R21）。此外，我们将检验siv -gp96疫苗对后续病毒攻击的保护能力（R33）。为了最大限度地提高成功的机会，迈阿密（Podack/Pahwa）的研究小组与美国国立卫生研究院（NIH）的专家（Franchini实验室）进行了合作，将基础免疫学家与人类和非人类灵长类动物HIV/SIV发病机制方面的专家聚集在一起。

DESCRIPTION (provided by applicant): Cellular immunity and memory is required for clearance of viruses and for protection from viral infection. Cellular immunity by CD8 CTL and NK cells is initiated through activation of the innate immune response, maturation of dendritic cells (DC) and antigen cross presentation to CD8 T cells. Anti viral vaccines stimulating cellular immunity have to imitate this process. Adjuvants for the stimulation of antibody responses are used effectively however our knowledge about adjuvants for the stimulation of CTL immunity is limited. We have demonstrated that the endoplasmic reticulum resident heat shock protein gp96, a chaperone for peptides transported to be presented by MHC-I, is a natural adjuvant for the activation of DC, NK and cognate CD8 T cells. DC and macrophages have receptors for gp96-peptide complexes and become activated upon gp96-binding. Gp96 together with its associated peptides is taken up the APC, the peptide moiety is transported to the ER and used to charge MHC-I molecules. Concomitant activation of DC results in more than million fold enhanced cross priming of cognate CD8 T cells when compared to cross priming by intact protein taken up by DC. To take advantage of this unique adjuvant effect and the ability to transport relevant peptides, we have made a secretable form of gp96, gp96-lg, by replacing the KDEL ER-retention signal with the Fc portion of lgG1. We hypothesize that cell-based gp96-lg vaccines, by prolonged in vivo secretion of gp96-lg-peptide, imitate viral replication and provide immune stimuli comparable to attenuated viruses. In model systems in mice we have shown that gp96-lg transfected, antigen expressing tumor cells secrete gp96-lg in vivo and stimulate the innate DC and NK as well as adaptive, cognate cellular CD8 CTL immune response and generate specific CD8 memory independent of CD4 help and in the absence of lymph nodes. Both systemic and strong mucosal immunity in intraepithelial, lamina propria and Peyer's patch CD8 CTL is generated by gp96-lg vaccines. Because of their unique properties we now plan to evaluate the gp96- vaccines in non-human primate models for SIV for their immunogenicity for mucosal and systemic cellular immunity (R21). In addition we will examine the protective power of SIV-gp96-vaccines against subsequent viral challenge (R33). To maximize the chances of success, the team in Miami (Podack/Pahwa) has entered into a collaboration with experts (Franchini lab) at the NIH bringing together basic immunologists with experts in human and non human primate HIV/SIV pathogenesis.