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

描述（申请人提供）：我们已经证明CMV/SIV载体可以1）再感染CMV+恒河猴（RM），2）在再感染期间，引发具有强“效应记忆”（TEM）偏好的有效和持久的SIV特异性CD 4+和CD 8 + T细胞应答，和3）在用高致病性SIVmac 239病毒进行有限剂量直肠攻击后，完全保护约50%的接种RM免于进行性感染。在这些RM中表现出的保护在其灵敏度和程度上与先前的疫苗不同，受保护的RM在初始感染时在血浆中表现出不同大小的病毒爆发，随后立即控制到不可检测的水平。尽管在受保护的RM中偶尔观察到血浆中的病毒斑点，但这些斑点随着时间的推移而下降，并且在1年后，保护不受CD 8+或CD 4+细胞耗竭的影响，并且使用超灵敏巢式PCR进行的广泛组织分析显示仅罕见地检测到~单拷贝SIV核酸并且没有活的SIV。保护与疫苗接种阶段产生的总SIV特异性CD 8 + TEM相关，并且在没有记忆应答的情况下发生。这些数据表明一种新的保护模式与非常早期的控制一致，可能发生在病毒进入的位点和/或病毒复制和扩增的早期位点，并涉及组织驻留的CD 8 + TEM。因此，CMV载体和“TEM”疫苗概念为HIV/AIDS疫苗开发提供了一种强有力的新方法。然而，由于CMV的致病潜力，完全复制型CMV载体不太可能用于人类。因此，一个中心目标是开发保持免疫原性和功效，但在人类中使用是安全的CMV载体。幸运的是，初步的数据表明，复制/传播缺陷型CMV可以是强免疫原性的。在这里，我们将探讨复制/传播缺陷型CMV载体的致病性和传播性可以在多大程度上减弱，同时保留免疫原性和疗效。提出了三个具体目标：1）设计、构建和体外表征复制/扩散缺陷型CMV/SIV载体，2）确定复制/扩散缺陷型CMV/SIV载体在胎儿RM中的致病性，和3）确定复制/扩散缺陷型CMV/SIV载体在幼年RM中的免疫原性和脱落，并将最佳复制/扩散缺陷型载体设计用于功效试验，以确定其保护免受限制剂量直肠SIVmac 239攻击的能力。 公共卫生相关性：2007年，全世界约有250万人新感染艾滋病毒（南部非洲某些地区的流行率超过15%），人们普遍认为，有效的预防性疫苗是控制艾滋病毒/艾滋病流行的唯一实际手段。我们已经证明，巨细胞病毒（CMV）载体可以利用组织驻留效应记忆T细胞，以打击艾滋病病毒感染的早期，并可以保护恒河猴从粘膜挑战后进行性感染。本申请中提出的工作将提高CMV载体的安全性，并为这种新型疫苗方法的进一步临床开发提供关键信息。

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.