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

描述（由申请方提供）：清除病毒和保护免受病毒感染需要细胞免疫和记忆。CD 8 CTL和NK细胞的细胞免疫是通过激活先天免疫应答、树突状细胞（DC）的成熟和抗原交叉呈递给CD 8 T细胞来启动的。刺激细胞免疫的抗病毒疫苗必须模仿这一过程。用于刺激抗体应答的佐剂被有效地使用，然而我们关于用于刺激CTL免疫的佐剂的知识是有限的。我们已经证明，内质网驻留热休克蛋白gp 96（MHC-I转运递呈的肽的伴侣蛋白）是DC、NK和同源CD 8 T细胞活化的天然佐剂。DC和巨噬细胞具有gp 96-肽复合物的受体，并在gp 96结合后被激活。Gp 96与其相关肽一起被APC吸收，肽部分被转运到ER并用于给MHC-I分子充电。与DC吸收的完整蛋白质的交叉引发相比，DC的伴随激活导致同源CD 8 T细胞的交叉引发增强了一百万倍以上。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IgG 1的Fc部分替换KDEL ER-保留信号，制备了gp 96的分泌形式gp 96-Ig。我们假设基于细胞的gp 96-lg疫苗，通过延长gp 96-lg肽的体内分泌，模仿病毒复制并提供与减毒病毒相当的免疫刺激。在小鼠的模型系统中，我们已经表明，gp 96-Ig转染的抗原表达肿瘤细胞在体内分泌gp 96-Ig，并刺激先天性DC和NK以及适应性同源细胞CD 8 CTL免疫应答，并在不存在淋巴结的情况下产生特异性CD 8记忆，而不依赖于CD 4辅助。gp 96-lg疫苗产生上皮内、固有层和派伊尔集合淋巴结中的系统性和强粘膜免疫CD 8 CTL。由于其独特的性质，我们现在计划在非人灵长类动物模型中评估SIV的gp 96疫苗的粘膜和全身细胞免疫的免疫原性（R21）。此外，我们将检查SIV-gp 96疫苗对随后的病毒攻击（R33）的保护能力。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NIH的专家（Franchini实验室）合作，将基础免疫学家与人类和非人类灵长类HIV/SIV发病机制的专家聚集在一起。

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.