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.

腺病毒六邻体及其在病毒与宿主相互作用中的作用。 项目摘要/摘要： 腺病毒载体(Ad)是美国临床试验中第二常用的治疗载体 许多先天和后天人类疾病，包括癌症。尽管到目前为止还没有找到治疗的方法 播散性转移性肿瘤，目前公认的播散性转移可以 可能通过系统性的传递途径，如血管系统，以允许接触到所有 身体部位可能存在转移性肿瘤。然而，在使用这一路线来实现系统性 腺病毒传递，超过90%的给药载体剂量被肝脏迅速隔离，导致 对病毒灭活，降低肝外基因转移的效果，并引发全身固有 免疫和炎症反应。尽管Ad细胞感染的体外模型假定 我们的体内分析表明，Ad纤维和五色子蛋白在介导病毒进入细胞中的关键作用 证明在血管内分娩后，主要的Ad衣壳蛋白--六邻体--起主要作用 驱动病毒在肝脏和肝细胞转导中的机制作用。重要的是，我们的 初步研究强烈表明，循环抗体与接触溶剂的特异性相互作用 超变量六邻体环机械地定义了病毒与库普弗细胞的相互作用，导致病毒 困在肝脏并失活。此外，我们的初步研究也表明，只有 同时灭活腺病毒与肝细胞、肝窦内皮细胞、 库普弗细胞允许病毒在血管内后从隔离在肝脏中逃逸 送货。尽管在肝细胞转导或与其相互作用时减弱的Ad载体 Kupffer细胞已经被描述，到目前为止，还没有发表的研究直接和明确地提供 有证据表明，这些载体在血管内注射后不久就逃脱了肝脏隔离。基于 不同肝细胞隔室等功能隔离腺病毒的新概念 布拉德，在这项提案中，我们将填补我们对Ad Hexon在引导病毒生物- 血管内分娩后的分布和传染性。通过结构上的低温电子的结合- 显微镜(冷冻-EM)、分析和定点突变的计算方法 我们将1)确定腺病毒六邻体与低亲和力天然相互作用的表面区 抗体(IgM)和高亲和力小鼠和人抗体(Ig G)。我们还将确定 Hexon hvr1在病毒感染、复制和Kupffer细胞捕获中的环变化。最后，使用一个集合 具有修饰的五子体和六子体的独特载体，我们将3)开发新型的六元突变病毒， 将避免库普弗细胞陷阱，并抵抗与病毒特异性抗体后血管内中和 送货。我们在本申请中提出的假设和数据驱动的研究将极大地推动我们的 了解Ad Hexon-宿主细胞和因子在体内的相互作用，并最终导致 防止血液中的广告隔离的新策略的实验验证。概念性和 这些策略的实验验证将代表着朝着发展SAFE迈出的重要一步 以及有效的系统适用的Ad载体，用于人类的多种治疗应用。