Programming designer DNA nanostructures for blocking enveloped viral infection

编程设计 DNA 纳米结构以阻止包膜病毒感染

• 批准号: 10598739
• 负责人: Weishan Huang
• 金额: $ 20.31万
• 依托单位: LOUISIANA STATE UNIV A&M COL BATON ROUGE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-14 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10598739
• 关键词: 2019-nCoV; ACE2; Agar Gel Electrophoresis; Animal Model; Antigens; Antiviral Therapy; Area; Atomic Force Microscopy; Avidity; Binding; Biological Assay; COVID-19; Cell model; Cells; Characteristics; Collection; DNA; Detection; Development; Dose; Effectiveness; Engineering; Epithelial Cells; Epitopes; Etiology; Exhibits; Goals; HIV; Human; In Vitro; Infection; Influenza; Knock-in Mouse; Knowledge; Lung; Mechanics; Medicine; Membrane Glycoproteins; Membrane Proteins; Molecular; Molecular Genetics; Nanostructures; Nanotechnology; Pattern; Positioning Attribute; Preventive; Price; Property; Proteins; Resistance; Route; SARS-CoV-2 B.1.1.529; SARS-CoV-2 B.1.617.2; SARS-CoV-2 P.1; SARS-CoV-2 infection; SARS-CoV-2 inhibitor; SARS-CoV-2 spike protein; SARS-CoV-2 variant; Safety; Sensitivity and Specificity; Series; Shapes; Structure; Surface; Technology; Testing; Therapeutic; Toxic effect; United States National Institutes of Health; Validation; Viral; Virion; Virus; Virus Diseases; Work; anti-viral efficacy; antiviral drug development; aptamer; biomaterial compatibility; cost; cost estimate; cytotoxic; cytotoxicity; design; drug development; flexibility; future epidemic; genetic technology; in vivo; innovation; innovative technologies; mimicry; mouse model; nano; nanobodies; nanoscale; pandemic disease; particle; pathogen; post SARS-CoV-2 infection; precision medicine; prevent; programs; prophylactic; prototype; receptor; receptor binding; screening; transmission process; variants of concern; viral entry inhibitor

Project Summary/Abstract SARS-CoV-2, the etiological pathogen of COVID-19, has resulted in a pandemic. There remains an urgent need for innovative technologies which facilitate the development of affordable antiviral precision medicine. SARS-CoV-2 is an enveloped virus, and the structure of the trimeric spike protein clusters on the virion has been solved. To develop innovative, affordable, and biocompatible antiviral candidates against SARS-CoV-2 infection and transmission, we exploited the structural characteristics of viral surface proteins that can be matched at nanoscale precision by engineered DNA nanostructure platforms. Based on the structure of the SARS-CoV-2 virion and surface spike trimer layout, we have synthesized a designer DNA nanostructure (DDN) that takes the form of a macromolecular ‘net’ whose vertices are a precise mechanical match to the spacing and positioning of the spike protein matrix displayed on the virus outer surface. We hypothesize that the structural properties and the layout patterns of SARS-CoV-2 spike proteins can be exploited to design DDNs with nanoscale precision which are capable of matching and capturing intact SARS-CoV-2 virions with ultrahigh binding avidity and selectivity, thereby blocking SARS-CoV-2 infection. We have screened and found DNA aptamers and nanobodies that are specific for the spike receptor-binding domain (RBD). These spike binders can be incorporated into the ‘knots’ of the DDN net to allow the simultaneous binding of multiple DNA aptamers with multiple spikes on the viral surface in a polyvalent, pattern-matching fashion. The DNA ‘net’-aptamer prototype construct has afforded dramatic increase in SARS-CoV-2 binding avidity. This construct can work as a decoy to block viral entry into host cells and is about 1,000-fold more potent than the free aptamer. In this R21 proposal, we aim to extend this technology to enable the incorporation of multiple types of probes against spike RBD and to validate the safety and effectiveness of DDNs in antiviral therapy in vitro and in vivo. We propose two specific aims: to (1) design, synthesize, validate, and further optimize the virus-capturing avidity against various SARS- CoV-2 variants of concern (VOCs); and (2) to determine the antiviral potency and cytotoxicity of the designed DDNs during SARS-CoV-2 infections in vitro in human lung epithelial cells and in vivo in human ACE2-knockin mice. Completion of this work will help us define the antiviral potency and safety of the DNA nanostructures that are designed to perfectly match epitope layouts on the viral surface to capture and wrap live viruses. The estimated cost of DDN treatment is approximately $10/dose (a price that likely decreases at large-scale synthesis), making it an affordable therapy. This DDN platform may further contribute to the rapid development of antiviral precision medicine against emerging SARS-CoV-2 VOCs, as well as other enveloped viruses such as influenza and HIV.

项目摘要/摘要 SARS-COV-2是COVID-19的病理学病原体，导致大流行。仍然很紧急 需要创新的技术，这些技术准备了负担得起的抗病毒精确医学的开发。 SARS-COV-2是一种包裹的病毒，病毒上三聚体峰蛋白簇的结构已是 已解决。开发针对SARS-COV-2感染的创新，负担得起和生物相容性的抗病毒候选者 和传输，我们探索了病毒表面蛋白的结构特征，可以匹配 通过工程DNA纳米结构平台进行的纳米级精度。基于SARS-COV-2的结构 Virion和表面尖峰触发布局，我们合成了设计器DNA纳米结构（DDN） 大分子“网”的形式，其顶点是与间距和定位的精确机械匹配 尖峰蛋白基质显示在病毒外表面上。我们假设结构性和 SARS-COV-2尖峰蛋白的布局模式可以探索以纳米级精度设计DDN 能够匹配和捕获完整的SARS-COV-2病毒，具有超高的结合亲和力和 选择性，从而阻止SARS-COV-2感染。我们已经筛选并找到了DNA适体， 针对尖峰受体结合结构域（RBD）的纳米型。这些尖刺的粘合剂可以是 纳入DDN网的“结”，以使多个DNA适体的简单结合与 以多价，图案匹配的方式在病毒表面上有多个尖峰。 DNA'Net'-Appamer原型 构造使SARS-COV-2结合的亲戚急剧增加。该结构可以作为诱饵 阻止病毒进入宿主细胞，并且比游离apatamer的潜力高约1,000倍。在此R21提案中 我们旨在扩展这项技术，以使针对Spike RBD的多种类型的问题保险和 验证DDN在体外和体内抗病毒治疗中的安全性和有效性。我们提出了两个特定的 目的：（1）设计，合成，验证和进一步优化了捕获病毒的亲戚，以各种SARS- COV-2关注的变体（VOC）； （2）确定设计的抗病毒效力和细胞毒性 SARS-COV-2在人类肺上皮细胞中的SARS-COV-2感染期间的DDN和人ACE2-KNOCKIN的体内 老鼠。这项工作的完成将有助于我们定义DNA纳米结构的抗病毒效力和安全性 旨在完美匹配病毒表面上的情节布局，以捕获和包裹活病毒。这 DDN治疗的估计成本约为10美元/剂量（价格可能在大规模下降的价格 合成），使其成为一种负担得起的疗法。这个DDN平台可能会进一步有助于快速发展 针对新兴SARS-COV-2 VOC的抗病毒精确药物以及其他包围病毒 作为影响力和艾滋病毒。