Development and In Vivo Characterization of Safety-Enhanced RhCMV/SIV Vectors

描述（由申请人提供）：我们已经证明CMV/SIV载体可以1)再次感染CMV+恒河猴（RM）， 2)在再次感染期间，引发有效和持续的SIV特异性CD4+和CD8+ T细胞反应，具有很强的“效应记忆”（TEM）偏好，以及3)在高致病性SIVmac239病毒的有限剂量直肠攻击后，完全保护约50%的接种过的恒河猴免受进进性感染。在这些RM中表现出的保护在其突发性和程度上与以前的疫苗不同，受保护的RM在初始感染时表现出不同大小的血浆病毒爆发，随后立即控制到无法检测到的水平。虽然在受保护的RM中偶尔会观察到血浆中的病毒突变，但随着时间的推移，这些突变会随着时间的推移而下降，并且在1年后，CD8+或CD4+细胞耗尽对保护不产生影响，并且使用超灵敏巢式PCR进行广泛的组织分析仅显示很少检测到~单拷贝SIV核酸，没有活的SIV。保护作用与疫苗阶段产生的siv特异性CD8+ TEM总量相关，且无遗忘反应。这些数据表明了一种与早期控制一致的新型保护模式，可能发生在病毒进入位点和/或病毒复制和扩增的早期位点，并涉及组织驻留CD8+ TEM。因此，巨细胞病毒载体和“TEM”疫苗概念为艾滋病毒/艾滋病疫苗开发提供了一个强有力的新途径。然而，由于巨细胞病毒的致病潜力，完全复制的巨细胞病毒载体不太可能先进到人类使用。因此，中心目标是开发既能保持免疫原性和有效性，又能安全用于人类的巨细胞病毒载体。幸运的是，初步数据表明，复制/扩散缺陷的cmv具有很强的免疫原性。在这里，我们将探讨在保留免疫原性和有效性的同时，复制/传播缺陷CMV载体在致病性和传播性方面可以减弱的程度。提出了三个具体目标：1)设计、构建复制/传播缺陷CMV/SIV载体并进行体外表征；2)确定复制/传播缺陷CMV/SIV载体在胎儿RM中的致病性；3)确定复制/传播缺陷CMV/SIV载体在幼年RM中的免疫原性和脱落情况，并将最佳复制/传播缺陷载体设计用于功效试验，以确定其对有限剂量直肠SIVmac239攻击的保护能力。

DESCRIPTION (provided by applicant): We have demonstrated that CMV/SIV vectors can 1) re-infect CMV+ rhesus macaques (RM), 2) during re-infection, elicit potent and persistent SIV-specific CD4+ and CD8+ T cell responses with a strong "effector memory" (TEM) bias, and 3) completely protect ~50% of vaccinated RM from progressive infection after limiting dose rectal challenge with the highly pathogenic SIVmac239 virus. The protection manifested in these RM is distinct from previous vaccines in its abruptness and extent, with protected RM exhibiting a viral burst in plasma of varying size upon initial infection, followed by immediate control to undetectable levels. Although occasional viral blips in plasma are observed in protected RM, these decline with time, and after 1 year, protection is unaffected by CD8+ or CD4+ cell depletion, and extensive tissue analysis with ultrasensitive nested PCR has shown only rare detection of ~ single copy SIV nucleic acid and no viable SIV. Protection correlates with the total SIV-specific CD8+ TEM generated during the vaccine phase, and occurs without an anamnestic response. These data indicate a novel pattern of protection consistent with very early control, likely taking place at the site of viral entry and/or early sites of viral replication and amplification, and involving tissue-resident CD8+ TEM. Thus, CMV vectors and the "TEM" vaccine concept offer a powerful new approach to HIV/AIDS vaccine development. However, fully replicative CMV vectors are unlikely to be advanced to human use due to the pathogenic potential of CMV. A central goal is therefore to develop CMV vectors that maintain immunogenicity and efficacy, but are safe to use in humans. Fortunately, preliminary data indicates that replication/spread-deficient CMVs can be robustly immunogenic. Here, we will explore the extent to which replication/spread-deficient CMV vectors can be attenuated with respect to pathogenicity and transmissibility, while retaining immunogenicity and efficacy. Three specific aims are proposed: 1) to design, construct and in vitro characterize replication/spread-deficient CMV/SIV vectors, 2) to determine the pathogenicity of replication/spread-deficient CMV/SIV vectors in fetal RM, and 3) to determine the immunogenicity and shedding of replication/spread-deficient CMV/SIV vectors in juvenile RM, and to take the optimal replication/spread deficient vector design to an efficacy trial to determine its ability to protect against limiting dose rectal SIVmac239 challenge. PUBLIC HEALTH RELEVANCE: Worldwide, ~2.5 million new HIV infections occurred in 2007 (with prevalence rates in some areas of southern Africa exceeding 15%), and it is generally agreed that an effective prophylactic vaccine is the only practical means by which the HIV/AIDS epidemic can be controlled. We have demonstrated that cytomegalovirus (CMV) vectors can harness tissue-resident effector memory T cells to combat the AIDS virus very early in infection, and can protect rhesus monkeys from progressive infection after mucosal challenge. The work proposed in this application will enhance the safety of CMV vectors, and provide crucial information for the further clinical development of this novel vaccine approach.