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

基本信息

批准号：
8852065
负责人：
PHOEBE L STEWART
金额：
$ 58.75万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
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/8852065
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 Health 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 receptor scavenger receptor vector

项目摘要

DESCRIPTION (provided by applicant): 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 an 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.

描述（由申请人提供）：腺病毒载体（AD）是美国临床试验中第二次最常用的载体，用于治疗包括癌症在内的许多先天和获得的人类疾病。尽管到目前为止尚未发现用于传播转移性肿瘤疾病的治疗方法，但目前可以接受，可以通过诸如脉管系统（例如脉管系统）的全身递送途径来治疗传播转移，以允许进入所有身体部位的转移性肿瘤可能是转移性肿瘤的。但是，在使用这种途径实现全身性腺病毒递送后，超过90％的施用载体剂量会被肝脏迅速隔离，导致病毒失活，降低了肝外基因转移的功效，并触发了全身性的先天性免疫和炎症反应。尽管AD细胞感染的体外衍生模型假设AD纤维和Penton蛋白在将病毒进入细胞中的关键作用，但我们的体内分析表明，在血管内递送后，主要的AD Capsid蛋白（己酮）在尿液中扮演了尿液中的主要机械机械作用，在尿液中扮演了尿液中的主要机械作用。重要的是，我们的初步研究强烈表明，循环抗体与暴露于溶剂的高变量己孔环的特定相互作用机械地定义了与库普弗细胞的病毒相互作用，从而导致肝脏中的病毒捕获和失活。此外，我们的初步研究还表明，只有同时灭活腺病毒与肝细胞，正弦内皮细胞和kupffer细胞的相互作用允许病毒在血管内递送后在肝脏中被隔离。尽管在肝细胞转导或与库普弗细胞的相互作用时已衰减的AD载体已被描述为迄今为止，尚未发表过直接有明确的证据，表明此类载体在血管内注射后不久就会逃脱肝脏螯合。基于不同肝细胞室在血液中隔离广告中的新型概念，在此提案中，我们将填补对AD己酮在指导病毒生物分布的作用的知识，使血管内递送后感染性具有感染性。通过结构冷冻电子显微镜（冷冻-EM）和分析的计算方法和位于定位的诱变方法，在本提案中，我们将1）确定与低亲和力自然抗体（IGM）和高亲和力小鼠和人类抗体和人类抗体（IGG）相互作用的腺病毒己酮的表面区域。我们还将2）确定Hexon HVR1环变异在病毒感染，复制和库普弗细胞捕获中的作用。最后，使用一组带有修饰的五孔和己酮的唯一矢量，我们将3）开发新型的己孔氧化病毒，这些病毒将避免库普弗细胞捕获并抵抗血管内递送后病毒特异性抗体中和中和中和中和。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提高我们对AD六链球菌的理解 - 体内的宿主细胞和因子相互作用，并最终应导致对防止血液中AD隔离的新型策略的实验验证。这些策略的概念和实验验证将代表为在人类中众多治疗应用开发安全有效的全身性AD向量迈出的重大步骤。