Unmasking the roles of viral glycoproteins in oral transmission of KSHV

项目摘要 全球每年报告的卡波西肉瘤（KS）新发病例超过44，000例，其中84%发生在 非洲这种和其他卡波西肉瘤相关疱疹病毒（KSHV）诱导的恶性肿瘤主要发生在 获得性或医源性免疫缺陷的人。虽然KSHV可以在其他人体中检测到， 液体，其在具有和不具有性传播感染风险的群体中的唾液中的频繁检测（例如， 儿童）表明口腔是主要的获得部位。然而，KSHV口服的机制 体内传播，特别是病毒进入所需的关键病毒包膜糖蛋白（GPS）， 仍然没有解决。已经确定了几种KSHV-宿主相互作用，但所有先前的实验都是 在体外进行，由于之前缺乏适当的动物模型，尚未在体内验证。通过 通过与威斯康星州国家灵长类动物研究中心的合作，我们的实验室可以获得常见的 绒猴（Callithrix jacchus，CJ），最近开发的KSHV非人灵长类动物模型，其对KSHV易感。 KSHV口腔感染，并在免疫抑制下获得KS样皮肤病变。的目的 本申请的目的是阐明在体内引发原发性口腔感染所需的最小GPS，作为 选择关键的全球定位系统，以开发有效的预防性候选疫苗。这个应用程序建立在博士。 奥根博最近完成了NCI K 01 CA 184388 -05关于KSHV进入机制和疫苗的研究 发展最近，我们发现在体外，KSHV糖蛋白gH/gL是病毒感染KSHV所必需的。 上皮、内皮和成纤维细胞，但不包括B细胞。值得注意的是，我们和其他人也表明， 抗KSHV糖蛋白gB、gH/gL和gpK8.1的单克隆和多克隆抗体可中和KSHV感染， 不同的人类细胞。基于这一成功，我们产生了KSHV缺失突变体， 认为对病毒进入至关重要的四种糖蛋白（gB、gH/gL、gpK8.1）和各种单克隆抗体 专门针对这些GPS。在这个项目中，我们将使用人体离体样本和CJ KSHV模型来测试 假设gB和gH/gL是KSHV体内经口传播关键。我们提出这个建议的前提是 建立在强有力的证据，1）KSHV可以感染CJ，发展KS样皮肤病变，和2）抗体对 KSHV糖蛋白gB和gH/gL在体外和离体都能中和KSHV感染。而且 允许KSHV离体感染人细胞，CJ使这些平台成为测试KSHV的理想平台 GP感染要求。成功完成拟议研究将阐明最低KSHV 在体外和体内模型中原发性感染所需的GPS，推进了我们定义 KSHV感染人类的初始步骤以及抗体在预防KSHV早期步骤中的作用 传输这将最终为预防KSHV的预防性疫苗的设计和开发提供信息 感染及其相关的癌症。

PROJECT SUMMARY More than 44,000 new cases of Kaposi sarcoma (KS) are reported globally each year, 84% of which occur in Africa. This and other Kaposi sarcoma-associated herpesvirus (KSHV)-induced malignancies predominate in people with acquired or iatrogenic immunodeficiencies. Although KSHV can be detected in other human body fluids, its frequent detection in saliva in groups both with and without risk of sexually transmitted infections (e.g., children) suggests that the oral cavity is the site of primary acquisition. However, the mechanism of KSHV oral transmission in vivo, particularly the critical viral envelope glycoproteins (gps) required for viral entry, remains unresolved. Several KSHV–host interactions have been identified, but all prior experiments were performed in vitro and have not been validated in vivo due to prior lack of an appropriate animal model. Through collaboration with the Wisconsin National Primate Research Center, our laboratory has access to the common marmoset (Callithrix jacchus, CJ), a recently developed KSHV non-human primate model that is susceptible to KSHV oral infection, and under immunosuppression acquires KS-like skin lesions. The objective of this application is to elucidate the minimum gps required to initiate primary oral infection in vivo, as a prerequisite to selecting key gps for developing an effective prophylactic vaccine candidate. This application builds on Dr. Ogembo’s recently completed NCI K01 CA184388-05 research on KSHV entry mechanisms and vaccine development. Recently, we showed that in vitro, the KSHV glycoprotein gH/gL is essential for viral infection of epithelial, endothelial, and fibroblasts cells, but not B cells. Notably, we and others have also shown that both monoclonal and polyclonal Abs to KSHV glycoproteins gB, gH/gL, and gpK8.1, can neutralize KSHV infection of diverse permissive human cells in vitro. Building on this success, we generated KSHV deletion mutants lacking the four glycoproteins thought to be critical for viral entry (gB, gH/gL, gpK8.1) and various monoclonal antibodies specific to these gps. In this project, we will use human ex vivo samples and the CJ KSHV model to test the hypothesis that gB and gH/gL are critical for KSHV in vivo oral transmission. The premise of our proposal is built on strong evidence that 1) KSHV can infect CJ, which develop KS-like skin lesions, and 2) Abs against the KSHV glycoproteins gB and gH/gL can neutralize KSHV infection in vitro and ex vivo. Furthermore, the permissiveness to KSHV infection of human cells ex vivo and CJ makes these platforms ideal to test the KSHV gp requirements for infection. Successful completion of the proposed study will elucidate the minimum KSHV gps required for primary infection in ex vivo and in vivo models, advancing our long-term goal of defining the initial steps in KSHV infection of humans and the role of antibodies in protecting against the early steps of KSHV transmission. This will ultimately inform design and development of prophylactic vaccines that can prevent KSHV infection and its associated cancers.