Adenovirus hexon and its role in virus interaction with the host

腺病毒六邻体及其在病毒与宿主相互作用中的作用

• 批准号: 8644629
• 负责人: PHOEBE L STEWART
• 金额: $ 59.3万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-06-01 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8644629
• 关键词: Ablation; Adenovirus Vector; Adenovirus hexon capsid protein; Adenoviruses; Affinity; Antibodies; Applications Grants; Area; Attenuated; Automobile Driving; Biology; Blood; Blood coagulation; Capsid; Capsid Proteins; Cells; Clinical; Clinical Trials; Complement Receptor; Complex; Computing Methodologies; Cryoelectron Microscopy; Data; Development; Discontinuous Capillary; Disease; Dose; Drug Kinetics; Endothelial Cells; Ensure; Epitopes; Excision; Fiber; Gene Transfer; Goals; Hepatic; Hepatocyte; Histocompatibility Testing; Human; Human body; Immune response; Immunity; Immunoglobulin G; Immunoglobulin M; In Vitro; Individual; Infection; Inflammatory Response; Injection of therapeutic agent; Integration Host Factors; Knowledge; Kupffer Cells; Lead; Link; Liver; Liver Circulation; Malignant Neoplasms; Mediating; Modeling; Molecular; Mus; Mutate; Mutation; Neoplasm Metastasis; Peptides; Play; Proteins; Publishing; RGD (sequence); Resolution; Risk; Role; Route; Serotyping; Site; Site-Directed Mutagenesis; Solvents; Structural Models; Surface; Therapeutic; Therapeutic Intervention; Tissues; Validation; Variant; Virion; Virus; Virus Diseases; Virus Inactivation; adenovirus penton protein; base; cell type; cellular transduction; hepatic sinusoid; human disease; in vivo; macrophage; neoplastic cell; novel; novel strategies; particle; prevent; public health relevance; receptor; scavenger receptor; vector

Adenovirus hexon and its role in virus interaction with the host. PROJECT SUMMARY/ABSTRACT: Adenovirus vectors (Ad) are the second most frequently used vectors in clinical trials in the US to treat numerous inborn and acquired human diseases, including cancer. Although no cure so far is found for disseminated metastatic tumor disease, it is currently accepted that disseminated metastases can potentially be treated through a systemic delivery routes, such as vasculature, to allow for access to all body sites were metastatic tumors may reside. However, upon using this route to achieve systemic adenovirus delivery, over 90% of the administered vector dose is rapidly sequestered by the liver, leading to virus inactivation, reducing the efficacy of extra-hepatic gene transfer, and triggering systemic innate immune and inflammatory responses. Although the in vitro-derived model of Ad cell infection postulates key roles for Ad fiber and penton proteins in mediating virus entry into cells, our in vivo analyses demonstrate that after intravascular delivery, the major Ad capsid protein - hexon - plays the principal mechanistic role in driving virus sequestration in the liver and hepatocyte transduction. Importantly, our preliminary studies strongly suggest that specific interactions of circulating antibodies with solvent-exposed hyper-variable hexon loops mechanistically define virus interaction with Kupffer cells, leading to virus trapping in the liver and inactivation. Furthermore, our preliminary studies also demonstrated that only simultaneous inactivation of adenovirus interactions with hepatocytes, sinusoid endothelial cells, and Kupffer cells allows for virus escape from being sequestered in the liver after intravascular delivery. Although Ad vectors that are attenuated at either hepatocyte transduction or interaction with Kupffer cells have been described, to date, there are no studies published that provide direct and definitive evidence that such vectors escape liver sequestration shortly after intravascular injection. Based on the novel concept of equifunctional role of different hepatocellular compartments in sequestering Ad from the blood, in this proposal we will fill the gap in our knowledge of the role of Ad hexon in guiding virus bio- distribution and infectivity after intravascular delivery. Through a combination of structural cryo-electron- microscopy (cryo-EM) and computational methods of analysis and site-directed mutagenesis, in this proposal we will 1) determine the surface regions of adenovirus hexon that interact with low affinity natural antibodies (IgM) and high affinity mouse and human antibodies (IgG). We will also 2) determine the role of the hexon HVR1 loop variation in virus infection, replication, and Kupffer cell trapping. Finally, using a set of unique vectors with modified pentons and hexons, we will 3) develop novel hexon-mutated viruses that will avoid Kupffer cell trapping and resist neutralization with virus-specific antibodies after intravascular delivery. Our hypothesis and data-driven studies proposed in this application will greatly advance our understanding of Ad hexon - host cell and factor interactions in vivo and should ultimately lead to the experimental validation of novel strategies to prevent Ad sequestration from the blood. Conceptual and experimental validation of these strategies would represent a major step toward the development of safe and effective systemically-applicable Ad vectors for numerous therapeutic applications in humans.

腺病毒己酮及其在与宿主的病毒相互作用中的作用。 项目摘要/摘要： 腺病毒载体（AD）是美国临床试验中第二次最常用的载体 包括癌症在内的许多先天和获得的人类疾病。虽然到目前为止尚未治愈 传播转移性肿瘤疾病，目前接受传播转移可以接受 有可能通过诸如脉管系统等系统性交付路线进行处理，以便访问所有人 身体部位可能存在转移性肿瘤。但是，使用此路线实现系统性 腺病毒递送，超过90％的施用载体剂量被肝脏迅速隔离，领先 使病毒失活，降低肝外基因转移的疗效并触发全身先天 免疫和炎症反应。尽管AD细胞感染的体外衍生模型假设 AD纤维和Penton蛋白在介导病毒进入细胞中的关键作用，我们的体内分析 证明在血管内输送后，主要的AD Capsid蛋白-Hexon-扮演主 机械作用在驱动肝脏和肝细胞转导中的病毒隔离中。重要的是，我们的 初步研究强烈表明，循环抗体与溶剂暴露的特定相互作用 超变量己孔环机械地定义了与库普弗细胞的病毒相互作用，导致病毒 捕获肝脏和失活。此外，我们的初步研究也表明 同时灭活腺病毒与肝细胞，正弦内皮细胞的相互作用， 和库普弗细胞允许病毒从血管内隔离在肝脏中 送货。尽管在肝细胞转导或与 迄今为止，已经描述了Kupffer细胞 证据表明，这种载体在血管内注射后不久逃脱了肝脏隔离。基于 不同肝细胞室在隔离广告中的新型概念 鲜血，在这一建议中，我们将填补我们对AD己酮在指导病毒生物的作用的知识的差距 血管内分娩后的分布和感染力。通过结构冷冻电子的结合 显微镜（冷冻EM）和分析和位置诱变的计算方法，在此中 提案我们将1）确定与低亲和力相互作用的腺病毒hexon的表面区域 抗体（IGM）和高亲和力小鼠和人类抗体（IgG）。我们还将2）确定 病毒感染，复制和库普弗细胞捕获中的Hexon HVR1回路变化。最后，使用一组 在带有修饰的Pentons和Hexon的独特载体中，我们将3）开发出新颖的己酮突变病毒 将避免库普弗细胞捕获和抵抗血管内病毒特异性抗体中和 送货。我们在本应用程序中提出的假设和数据驱动的研究将大大推动我们的 对AD六链的理解 - 体内的宿主细胞和因子相互作用，最终应导致 对防止血液隔离的新型策略的实验验证。概念和 这些策略的实验验证将代表开发安全的重大步骤 以及用于人类众多治疗应用的有效全身性易用的AD媒介。