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Zika（Zikv）病毒是蚊子传播的病毒，是主要的医疗和公共卫生 全球问题。 DENV引起最普遍的蚊子传播的人类病毒疾病，严重 表现出血管泄漏的病例可能是致命的。相关的寨卡病毒（ZIKV）最近引起了爆炸性 整个美洲流行病，与先天性先天缺陷和吉兰·巴雷有关 综合征。尽管它们在全球范围内的发病率和死亡率很高，但尚无治疗剂 治疗登革热或寨卡病毒。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进化和病毒发病机理中起着重要作用。我们 以及其他人最近描述了NS1在直接触发内皮屏障功能障碍和 诱导人类免疫细胞的炎性细胞因子产生，导致体内血管泄漏。在这个 提案，我们将评估聚糖对NS1介导的发病机理的体外和体内效率，如 并反对体内的DENV和ZIKV感染。我们已经开发了多种研究DENV和ZIKV的方法 基于NS1诱导的内皮屏障功能障碍的表征的发病机制（例如 使用病毒的鼠模型 - NS1诱导的疾病具有血管泄漏。我们的初步结果表明β-的硫化衍生物 来自巴西脂蛋白真菌（FR-S）的葡聚糖在体外具有保护作用，并在体内对DENV NS1- 承诺诱发内皮过度过度。我们还表明，FR-S具有抗denv和抗 ZIKV活性在体外。与K. Godula博士（圣地亚哥加州大学）合作，我们确定了特定的合成 GAG模拟分子有效与黄病毒NS1结合。鉴于NS1对黄病毒发病机理的贡献 和我们的初步结果，我们假设聚糖抑制了NS1诱导的EGL降解和VAS- 体内和病毒感染的滤清液泄漏，具有新型治疗方式的潜力 登革热和寨卡。这种方法具有创新性，因为我们针对严重疾病表现的抑制 抗病毒活性。 AIM 1将选择NS1诱导的最有前途的抑制剂 基于预防体外内皮功能障碍的DENV和ZIKV的病理生理途径。 我们将筛选27种糖果，包括FR和FR-S，4个GAG-MIMETICS，悬极和20个环糊精类似物 （与Cyclolab的T. Sohajda博士合作），以防止NS1诱导的过度过敏性。目的 2将研究所选的预防内皮功能障碍的体外机理 化合物。 AIM 3将评估最活跃化合物对DENV的治疗潜力和 ZIKV NS1和病毒引起的血管泄漏，发病率和体内死亡率。总体而言，此提案地址 - ES是一个急需的需求，通过开发针对两种主要黄病毒疾病的新型治疗策略 基于聚糖的化合物靶向感染的病毒和病理生理后果。