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 许多先天性和后天性人类疾病，包括癌症。虽然目前还没有找到治愈方法 播散性转移性肿瘤疾病，目前公认的是播散性转移瘤可 可能通过系统性输送途径（例如脉管系统）进行治疗，以允许访问所有 身体部位可能存在转移性肿瘤。然而，当使用这种途径来实现系统性 腺病毒递送时，超过 90% 的载体剂量被肝脏迅速隔离，导致 病毒灭活，降低肝外基因转移的功效，并引发全身先天性 免疫和炎症反应。尽管Ad细胞感染的体外衍生模型假设 Ad 纤维和五邻体蛋白在介导病毒进入细胞中的关键作用，我们的体内分析 证明血管内递送后，主要的 Ad 衣壳蛋白 - 六邻体 - 起主要作用 在驱动病毒隔离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暴露于溶剂的抗体的特异性相互作用 高变六邻体环机械地定义了病毒与库普弗细胞的相互作用，从而导致病毒 捕获在肝脏中并失活。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、肝窦内皮细胞的相互作用， 库普弗细胞允许病毒在血管内逃逸后被隔离在肝脏中 送货。尽管Ad载体在肝细胞转导或与肝细胞相互作用时减弱 库普弗细胞已被描述，迄今为止，还没有发表的研究提供直接和明确的信息 有证据表明，此类载体在血管内注射后不久即可逃脱肝隔离。基于 不同肝细胞区室在隔离 Ad 方面的同等作用的新概念 血液，在这项提案中，我们将填补我们对 Ad 六邻体在引导病毒生物中的作用的知识空白。 血管内分娩后的分布和感染性。通过结构冷冻电子的组合 显微镜（冷冻电镜）以及分析和定点诱变的计算方法，在此 建议我们将 1) 确定与低亲和力天然相互作用的腺病毒六邻体的表面区域 抗体（IgM）和高亲和力小鼠和人类抗体（IgG）。我们还将 2) 确定以下角色： 病毒感染、复制和库普弗细胞捕获中的六邻体 HVR1 环变异。最后，使用一组 具有修饰的五邻体和六邻体的独特载体，我们将3）开发新型六邻体突变病毒 将避免库普弗细胞捕获并抵抗血管内病毒特异性抗体的中和 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进我们的研究 了解 Ad 六邻体 - 宿主细胞和因子在体内的相互作用，并最终应导致 防止 Ad 从血液中隔离的新策略的实验验证。概念和 这些策略的实验验证将代表朝着安全开发迈出的重要一步 以及有效的系统适用的 Ad 载体，可用于人类的众多治疗应用。