Evaluation of in vitro and in vivo efficacy of glycan-based compounds against flavivirus endothelial permeability and vascular leak

聚糖基化合物对抗黄病毒内皮通透性和血管渗漏的体外和体内功效评估

• 批准号: 10115592
• 负责人: Eva Harris
• 金额: $ 20万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-01 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10115592
• 关键词: Address; Agaricus; Americas; Anticoagulants; Antiviral Agents; Arbovirus Infections; Binding; Biological; Biological Assay; Biological Availability; Blood Vessels; Case Management; Cathepsin L; Cells; Characteristics; Collaborations; Complex; Confocal Microscopy; Congenital Abnormality; Cyclodextrins; Dengue; Dengue Infection; Dengue Virus; Disease; Electrical Resistance; Endothelial Cells; Endothelium; Epidemic; Evaluation; Extravasation; Flavivirus; Functional disorder; Glycocalyx; Glycosaminoglycans; Guillain Barré Syndrome; Heparitin Sulfate; Human; Immune; Immune Evasion; In Vitro; Infection; Inflammatory; Location; Measures; Mediating; Medical; Methods; Modality; Molecular Conformation; Molecular Weight; Morbidity - disease rate; Mus; Nonstructural Protein; Pathogenesis; Pathway interactions; Peripheral Blood Mononuclear Cell; Permeability; Pharmaceutical Preparations; Play; Polysaccharides; Prevention; Production; Proteins; Public Health; RNA replication; Role; Route; Safety; Series; Sialic Acids; Structure; Structure-Activity Relationship; Sulfate; Targeted Research; Testing; Therapeutic; Therapeutic Agents; Toxic effect; Viral Load result; Viral Nonstructural Proteins; Viral Pathogenesis; Viral Proteins; Virus; Virus Diseases; ZIKA; ZIKV disease; ZIKV infection; Zika Virus; analog; base; beta-Cyclodextrins; beta-Glucans; cytokine; cytotoxicity; design; endothelial dysfunction; fungus; glucuronyl glucosamine glycan sulfate; heparanase; human disease; improved; in vitro activity; in vivo; inhibitor/antagonist; innovation; mimetics; mortality; mosquito-borne; mouse model; novel; novel therapeutic intervention; preservation; prevent; protective effect; sugar; syndecan; viral RNA

ABSTRACT Dengue (DENV) and Zika (ZIKV) viruses are mosquito-borne viruses that are major medical and public health problems worldwide. DENV causes the most prevalent mosquito-borne viral disease of humans, and severe cases manifesting vascular leakage can be fatal. The related Zika virus (ZIKV) recently caused explosive epidemics across the Americas and has been associated with congenital birth defects and Guillain-Barré syndrome. Despite their substantial worldwide morbidity and mortality, no therapeutic agents exist for treatment of dengue or Zika. Nonstructural protein 1 (NS1) is a flaviviral protein that participates in viral RNA replication and in its secreted form plays important roles in host immune evasion and viral pathogenesis. We and others recently described novel roles for NS1 in directly triggering endothelial barrier dysfunction and inducing inflammatory cytokine production from human immune cells, contributing to vascular leak in vivo. In this proposal, we will evaluate the in vitro and in vivo efficacy of glycans against NS1-mediated pathogenesis, as well as against DENV and ZIKV infection in vivo. We have developed multiple methods to study DENV and ZIKV pathogenesis based on characterization of NS1-induced endothelial barrier dysfunction in vitro (e.g., hyper- permeability, disruption of the endothelial glycocalyx-like layer [EGL]) and in vivo, using murine models of virus- and NS1-induced disease with vascular leakage. Our preliminary results showing that a sulfated derivative of β- glucan from Agaricus brasiliensis fungus (FR-S) has a protective effect in vitro and in vivo against DENV NS1- induced endothelial hyperpermeability are promising. We have also shown that FR-S has anti-DENV and anti- ZIKV activity in vitro. In collaboration with Dr. K. Godula (UC San Diego) we determined that specific synthetic GAG-mimetic molecules efficiently bind to flavivirus NS1. Given the contribution of NS1 to flavivirus pathogenesis and our preliminary results, we hypothesize that glycans inhibit NS1-induced EGL degradation and vas- cular leakage in vivo as well as viral infection and have potential as novel treatment modalities for dengue and Zika. The approach is innovative in that we target inhibition of severe disease manifestations, in addition to antiviral activity. Aim 1 will select the most promising inhibitor(s) of NS1-induced pathophysiological pathways of DENV and ZIKV based on prevention of endothelial dysfunction in vitro. We will screen 27 glycans, including FR and FR-S, 4 GAG-mimetics, sulodexide, and 20 cyclodextrin analogues (in collaboration with Dr. T. Sohajda, CycloLab), for their ability to prevent NS1-induced hyperpermeability. Aim 2 will investigate the in vitro mechanism of action of prevention of endothelial dysfunction by the selected compounds. Aim 3 will evaluate the therapeutic potential of the most active compounds against DENV and ZIKV NS1- and virus-induced vascular leak, morbidity, and mortality in vivo. Overall, this proposal address- es a critical need, identifying novel therapeutic strategies against two major flaviviral diseases, by developing glycan-based compounds that target both the virus and pathophysiological consequences of infection.

抽象的 登革热 (DENV) 和寨卡 (ZIKV) 病毒是蚊媒病毒，是主要的医疗和公共卫生问题 DENV 是人类最常见的蚊媒病毒性疾病，并且是一种严重的全球性问题。 相关的寨卡病毒（ZIKV）最近引起了爆炸性病例，表现为血管渗漏。 美洲流行病，与先天性出生缺陷和吉兰-巴利综合征有关 尽管该综合征在世界范围内发病率和死亡率很高，但尚无治疗药物。 非结构蛋白 1 (NS1) 是一种参与病毒 RNA 的黄病毒蛋白。 复制及其分泌形式在宿主免疫逃避和病毒发病机制中发挥着重要作用。 等人最近描述了 NS1 在直接触发内皮屏障功能障碍中的新作用， 诱导人体免疫细胞产生炎症细胞因子，导致体内血管渗漏。 提案中，我们将评估聚糖对 NS1 介导的发病机制的体外和体内功效，如 以及体内对抗 DENV 和 ZIKV 感染的能力 我们开发了多种方法来研究 DENV 和 ZIKV。 发病机制基于体外 NS1 诱导的内皮屏障功能障碍的特征（例如，过度 渗透性、内皮糖萼样层 [EGL] 的破坏）和体内，使用病毒的鼠模型 我们的初步结果表明，β- 的硫酸化衍生物。 巴西蘑菇 (FR-S) 中的葡聚糖对 DENV NS1- 具有体外和体内保护作用 我们还表明 FR-S 具有抗 DENV 和抗 - 我们与 K. Godula 博士（加州大学圣地亚哥分校）合作确定了该特定合成物的体外活性。 鉴于 NS1 对黄病毒发病机制的贡献，GAG 模拟分子可有效结合黄病毒 NS1。 以及我们的初步结果，我们发现聚糖抑制 NS1 诱导的 EGL 降解和血管 体内的眼球渗漏以及病毒感染，并有可能作为新的治疗方式 该方法的创新之处在于我们的目标是抑制严重疾病的表现。 除了抗病毒活性之外，目标 1 将选择最有前途的 NS1 诱导抑制剂。 基于体外预防内皮功能障碍的 DENV 和 ZIKV 病理生理学途径。 我们将筛选 27 种聚糖，包括 FR 和 FR-S、4 种 GAG 模拟物、舒洛地昔和 20 种环糊精类似物 （与 CycloLab 的 T. Sohajda 博士合作），表彰其预防 NS1 诱导的通透性过高的能力。 2 将研究所选药物预防内皮功能障碍的体外作用机制 目标 3 将评估最活跃的化合物 DENV 和的治疗潜力。 总体而言，该提案解决了 ZIKV NS1 和病毒引起的体内血管渗漏、发病率和死亡率。 迫切需要通过开发针对两种主要黄病毒疾病的新治疗策略 基于聚糖的化合物，针对病毒和感染的病理生理后果。