Targeting Immunogenicity to the MPER Hinge and C-helix for BNAb Elicitation

项目摘要--总体 虽然显然需要疫苗来诱导针对该病毒的广谱中和抗体(BNAbs) 高度变异的逆转录病毒HIV-1能够阻止人与人之间的传播和全球传播，是实现这一目标的手段 仍然难以捉摸。在自然条件下产生的针对gp160三聚体易损部位的抗体 感染进一步推动逆转录病毒变异，使个体中的准种多样化。一个明显的例外是 膜近端外区(MPER)部位是隐形的，主要浸泡在脂质中，只有 在半融合/融合过程中被发现。因此，免疫压力不会混淆或助长病毒。 逃走。然而，自然产生的抗体的缺乏被认为是由 在最早的B细胞检查点删除鉴于脂质反应性，多特异性，甚至自身反应性 某些抗MPER的单抗。相反，我们最近的小鼠免疫原性研究已经产生了生发 利用脂质体排列的MPER制备中心来源和T细胞依赖的亲和力成熟的抗MPER抗体 无临床后遗症或明显抗脂反应的节段。这些发现与我们的研究结果相一致。 评估了在HIV-1患者中自然产生的人类抗MPER抗体的更高流行率。此PO1 专注于产生具有强大bNab特异性的抗MPER反应，创造出 捕获在三聚体解离转变过程中产生的天然MPER构象状态，从 紧密的环状gp160碱基到平坦的病毒膜上的异构体，具有最大的MPER暴露。这 目标应使用#年项目1中Reherz小组(DFCI)的分子免疫学方法来实现 与欧文集团(MIT)在项目2中的生物材料专业知识和利用技术相结合 组分A-D、核磁共振(Wagner，Harvard)、电子显微镜(EM)(Walz，Rockefeller)、EPR(Song， NHMFL)和微雕(Love，MIT)。免疫原设计的优化将通过以下方式实现 合成肽核酸在纳米疫苗脂膜上MPER定位的研究 (PNA)茎、酰基链和TM链段连接体，并通过核磁共振、EPR、EM和Biacore测量进行审查。在……里面 此外，最佳的纳米级MPER段间距和组织将涉及未承诺的常见 BNAbs的祖先(UCA)。抗体附着的载体空间将由生物材料配方控制 聚合物聚乙二醇组分的“空间云”和纳米盘包埋gp160在 异源免疫策略。单细胞微刻法挽救诱生单抗的趋近角 将进行评估，并与自然产生的bNAbs进行比较。将化学佐剂联合交付给 激活刺痛和ICOS通路将增强生发中心的CD4TFH，以驱动体细胞 超突变。用散装血清分析评估疫苗诱导的浆细胞长期发育 用常规小鼠免疫球蛋白和单个骨髓(BM)浆细胞微雕，以及完整的免疫球蛋白. Locus人源化小鼠(Kymouse)将允许在相关的临床前模型系统中调整免疫原。

Project Summary - Overall While there is an obvious need for vaccines eliciting broadly neutralizing antibodies (BNAbs) against the highly mutable retrovirus HIV-1 to stem person-to-person and global spread, the means to achieve this goal have remained elusive. Antibodies produced against trimeric gp160 sites of vulnerability during natural infection drive retroviral mutation further, diversifying quasispecies in individuals. One apparent exception is the membrane proximal external region (MPER) site, which is stealth, largely immersed in lipid and only revealed during hemifusion/fusion. Hence, immune pressures are not confounding or contributing to viral escape. However, a paucity of naturally arising antibodies has been postulated to be a consequence of deletion at the earliest B cell checkpoints in view of lipid reactivity, polyspecificity and even autoreactivity of certain anti-MPER mAbs. To the contrary, our recent mouse immunogenicity studies have generated germinal center-derived and T cell-dependent, affinity-matured anti-MPER antibodies using liposome-arrayed MPER segments without clinical sequelae or significant anti-lipid reactivity. These findings are consistent with the re- evaluated greater prevalence of naturally arising human anti-MPER antibodies in HIV-1 patients. This PO1 focuses on engendering anti-MPER responses with potent BNAb specificities, creating immunogens that capture native MPER conformational states arising during transitions of trimer unwinding, going from the compact torus-like gp160 base to conformers on the flat viral membrane with maximum MPER exposure. This goal shall be achieved using molecular immunology approaches of the Reinherz group (DFCI) in Project 1 in conjunction with the biomaterials expertise of the Irvine group (MIT) in Project 2 and utilizing technology components A-D, NMR (Wagner, Harvard), electron microscopy (EM) (Walz, Rockefeller), EPR (Song, NHMFL) and microengraving (Love, MIT), respectively. Optimization of immunogen design will be achieved by orientation of the MPER on lipid membranes in nanovaccines through the use of synthetic peptide nucleic acid (PNA) stalks, acyl chain and TM segment linkers and vetted by NMR, EPR, EM and BIAcore measurements. In addition, optimal nanoscale MPER segment spacing and organization will engage uncommitted common ancestors (UCAs) of BNAbs. Vector space of antibody attachment will be controlled by biomaterial formulation of polymer poly(ethylene glycol) (PEG) "steric clouds" and utilization of nanodisc-embedded gp160 in heterologous immunization strategies. Approach angles of elicited mAbs rescued by single-cell microengraving will be assessed and compared with those of naturally arising BNAbs. Co-delivery of chemical adjuvants to activate the STING and ICOS pathways shall augment CD4 TFH in germinal centers to drive somatic hypermutation. Assessment of vaccine-induced long-term plasma cell development by analysis of bulk serum IgG and single bone marrow (BM) plasma cell microengraving using conventional mice as well as complete Ig- locus humanized mice (KyMouse) will allow for immunogen tuning in a relevant pre-clinical model system.