Evaluation of Protective CMV Vaccines in Rhesus Macaques

描述（由申请人提供）：35年来对人类巨细胞病毒（HCMV）先天性感染具有保护作用的疫苗的研究仍未得到满足。HCMV自然史的复杂性、免疫保护相关因素的不完全定义，以及设计足够有力的临床试验时的财务和后勤因素，都导致缺乏许可的HCMV疫苗。现在，在人类和恒河猴（RM）的研究中，人们越来越认识到：(1)HCMV和恒河猴CMV （RhCMV）进入细胞（如内皮细胞和上皮细胞）的机制与进入成纤维细胞的机制不同；(2)血清免疫个体产生中和抗体，对抗介导上皮/内皮细胞向性的病毒蛋白。对HCMV的研究表明，UL128、131A、130蛋白与gH和gL共同形成复合物，通过内吞途径对上皮/内皮细胞的趋向性至关重要，缺乏这些蛋白中的任何一种都极大地限制了HCMV感染以gb介导的方式进入细胞，如成纤维细胞。来自自然史和组织培养研究的大量证据表明，疫苗策略将需要阻断内吞（UL128复合物）和成纤维细胞（gB）的进入途径，以建立足够水平的保护性免疫，以减少或预防先天性感染。HCMV的数据和我们在rMVA作为单个亚基表达RhCMV UL128的分泌形式以及描述UL128复合物在RhCMV自然史中的作用方面的进展，导致了这样的假设，即针对UL128复合物的疫苗策略将显示出对病毒在体液中脱落的增强限制。基于这一假设，接种具有UL128复合体的RhCMV-naove RM将显著降低全身感染性病毒的频率和滴度，从而降低棚病毒的水平。这一假设将通过以下目的在我们的HCMV持久性和发病机制的灵长类模型中得到严格检验。(1) UL128复合体质粒表达载体和MVA载体的构建与表征。(2)利用Aim 1构建的表达载体优化猕猴疫苗接种，表征rhcmv感染猕猴外周血和黏膜对UL128复合体的抗体反应。(3)利用Aim 2优化的初始/增强方案免疫RhCMV阴性RM，然后通过皮下或粘膜途径攻击RhCMV， RhCMV既能成纤维细胞也能进入内吞。

DESCRIPTION (provided by applicant): The 35 year quest for a vaccine that confers protective efficacy against congenital infection with human cytomegalovirus (HCMV) remains unmet. Complexities in HCMV natural history, incompletely defined correlates of immune protection and financial and logistical factors in designing sufficiently powered clinical trials all contribute to the absence of a licensed HCMV vaccine(s). There is now increased recognition from studies in both humans and rhesus macaques (RM) that (1) the mechanism of HCMV and rhesus CMV (RhCMV) entry into cells, such as endothelial and epithelial cells, is distinct from the mechanism of fibroblast entry, and (2) seroimmune individuals develop neutralizing antibodies against the viral proteins mediating epithelial/endothelial cell tropism. Studies with HCMV show that the UL128, 131A, 130 proteins together form a complex with gH and gL that is essential for epithelial/endothelial cell tropism via an endocytic pathway, and the absence of any one of these proteins greatly restricts HCMV infection to gB-mediated entry into cells, such as fibroblasts. The preponderance of evidence from natural history and tissue culture studies evidence indicates that vaccine strategies will need to block both the endocytic (UL128 complex) and fibroblast (gB) entry pathways to establish a sufficient level of protective immunity to reduce or prevent congenital infection. The data with HCMV and our progress in both expressing a secreted form of RhCMV UL128 from rMVA as a single subunit and characterizing the role of the UL128 complex in RhCMV natural history lead to the hypothesis that vaccine strategies targeting the UL128 complex will show enhanced restriction of virus shedding in bodily fluids. Based on this hypothesis, vaccination of RhCMV-naove RM with the UL128 complex will have a significantly reduced frequency and titer of systemic infectious virus, resulting in reduced levels of shed virus. This hypothesis will be rigorously tested in our primate model of HCMV persistence and pathogenesis through the following Aims. (1) Construction and characterization of plasmid expression and MVA vectors for the UL128 complex. (2) Optimization of vaccination in macaques with expression vectors constructed in Aim 1, and characterization of peripheral and mucosal antibody responses to the UL128 complex in RhCMV-infected macaques. (3) Immunization of RhCMV-negative RM by the optimized prime/boost regimen from Aim 2 followed by either a subcutaneous or mucosal route of RhCMV challenge capable of both fibroblast and endocytic entry. PUBLIC HEALTH RELEVANCE: This proposal will show in a primate RhCMV model that faithfully recapitulates human infection and expresses the monkey equivalent of the HCMV UL128 complex that a protective vaccine augmented with RhCMV UL128 complex antigens will significantly limit the capacity of challenge virus to disseminate. These studies should lead to an optimal prophylactic HCMV clinical strategy that can lead to clinically relevant reductions in the potential for both vertical and horizontal transmission of HCMV.