Structure-guided design of protease-resistant, lipopeptide inhibitors of SARS-CoV-2

SARS-CoV-2 蛋白酶抗性脂肽抑制剂的结构指导设计

基本信息

批准号：
10679139
负责人：
Ariel Jade Kuhn
金额：
$ 6.87万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-03-01 至 2026-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10679139
关键词：
2019-nCoV Address Amino Acid Substitution Amino Acids Animal Model Antiviral Agents Biodistribution Biological Assay Biological Availability C-terminal COVID-19 COVID-19 therapeutics Cell membrane Cells Cessation of life Chemicals Cholesterol Circular Dichroism Collaborations Coronavirus Crystallization Crystallography Data Development Dose Drug Kinetics Economics Engineering Etiology Exhibits Fluorescence Polarization Foundations Goals HIV Half-Life Hybrids In Vitro Infection Learning Length Mediating Membrane Fusion Membrane Proteins Middle East Respiratory Syndrome Modification Molecular Molecular Conformation N-terminal Para-Influenza Virus Type 3 Pathogenesis Peptide Hydrolases Peptides Positioning Attribute Predisposition Productivity Property Prophylactic treatment Protein Engineering Proteins Proteolysis Research Research Personnel Resistance Resolution SARS coronavirus SARS-CoV-2 infection SARS-CoV-2 inhibitor SARS-CoV-2 variant Side Site Structure Surface Techniques Therapeutic Agents Treatment Efficacy Universities Vaccines Variant Vertebral column Viral Viral Physiology Virus Virus Diseases Virus Inhibitors Vulnerable Populations Wisconsin Work X ray diffraction analysis X-Ray Crystallography analog career coronavirus disease design experience improved in vivo inhibitor insight interdisciplinary approach mimicry novel novel strategies novel therapeutics pandemic disease protein expression viral entry inhibitor virology

项目摘要

PROJECT SUMMARY Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), the etiological agent of coronavirus disease (COVID-19), has spurred an unprecedented global pandemic. Infection surges necessitate new therapeutic agents that are effective against a rapidly changing virus. Antivirals that inhibit viral entry into host cells have proven effective against other viruses with similar mechanisms of pathogenesis. The long-term objective of this highly collaborative proposed research is to develop potent peptides inhibitors of SARS-CoV-2 infection that operate by blocking structural rearrangements of the spike protein required for viral entry into host cells. For SARS-CoV-2 infection to occur, the viral surface spike (S) protein, a homotrimer, rearranges to form an energetically favored postfusion state. In this postfusion conformation, two helical domains, the N-terminal (HRN) and C-terminal (HRC) heptad repeats, associate to form a 6-helix bundle (6HB). Peptides derived from the HRC can inhibit formation of the 6HB, and thus SARS-CoV-2 infection. However, conventional peptides, composed entirely of α-amino acid residues, are highly susceptible to proteolytic degradation, which necessitates frequent and high dosing. The Gellman lab, in collaboration with virologists Prof. Anne Moscona and Prof. Matteo Porotto at Columbia University, has demonstrated that site- selective incorporation of backbone modifications, in combination with cholesterol conjugation, can decrease proteolytic sensitivity while maintaining high antiviral potency. Our team recently found that such lipopeptides can inhibit SARS-CoV-2 infection in biological assays and animal models, and that these inhibitors are effective against SARS-CoV-2 variants, SARS-CoV-1 and MERS. Building on this foundation, my proposed project seeks to develop lipopeptides containing backbone modifications that display high antiviral potency and resist proteolysis. Aim 1 will produce potent inhibitors of SARS-CoV-2 (and other coronaviruses) that contain backbone modifications and resist proteolysis. Aim 2 will evaluate the stability of 6HB formation between inhibitor candidates and the native SARS-CoV-2 HRN. Aim 3 will elucidate critical structural interactions between the HRC mimics and the native HRN. My hypothesis is that site-selective incorporation of backbone modifications into HRC-based designs will increase both antiviral activity and half-life in vivo, improving therapeutic efficacy. The proposed research, which will be conducted under the guidance of Prof. Sam Gellman at the University of Wisconsin, will provide me with experience in macromolecular X-ray crystallography, molecular design, protein engineering & expression, and virology. Through structure-guided engineering and sophisticated and multi-pronged assay implementation, these efforts could generate effective pan-variant therapeutics for COVID-19.

项目概要严重急性呼吸综合征冠状病毒 2 型 (SARS-CoV-2)，冠状病毒的病原体疾病（COVID-19）引发了前所未有的全球大流行，需要新的感染激增。有效对抗快速变化的病毒的治疗剂，抑制病毒进入宿主。细胞已被证明可以有效对抗具有类似发病机制的其他病毒。这项备受瞩目的合作研究的长期目标是开发有效的肽 SARS-CoV-2 感染的抑制剂，通过阻断刺突蛋白的结构重排来发挥作用病毒进入宿主细胞需要病毒表面刺突 (S) 蛋白。同源三聚体，重新排列以形成能量有利的融合后状态。在这种融合后构象中，两个。螺旋结构域、N 端 (HRN) 和 C 端 (HRC) 七肽重复序列结合形成 6 螺旋束 (6HB)。源自 HRC 的肽可以抑制 6HB 的形成，从而抑制 SARS-CoV-2 感染。然而，传统的肽完全由α-氨基酸残基组成，非常容易受到蛋白水解降解，这需要与盖尔曼实验室合作进行频繁和高剂量的治疗。哥伦比亚大学病毒学家 Anne Moscona 教授和 Matteo Porotto 教授已经证明，选择性掺入主链修饰，与胆固醇缀合相结合，可以减少我们的团队最近发现这种脂肽具有蛋白水解敏感性，同时保持高抗病毒效力。在生物测定和动物模型中可以抑制 SARS-CoV-2 感染，并且这些抑制剂是有效的在此基础上，我提出的项目旨在针对 SARS-CoV-2 变种、SARS-CoV-1 和 MERS。开发含有主链修饰的脂肽，显示出高抗病毒效力并抵抗目标 1 将产生包含骨架的 SARS-CoV-2（和其他冠状病毒）的有效抑制剂。修饰和抗蛋白水解作用目标 2 将评估抑制剂之间 6HB 形成的稳定性。目标 3 将阐明候选者和天然 SARS-CoV-2 HRN 之间的关键结构相互作用。 HRC 模仿者和原生 HRN。我的假设是，将主链修饰选择性地纳入基于 HRC 的设计中将增加抗病毒活性和体内半衰期，提高治疗效果。将在威斯康星大学 Sam Gellman 教授的指导下进行，将为我提供具有大分子 X 射线晶体学、分子设计、蛋白质工程和表达方面的经验，以及通过结构引导工程和复杂的多管齐下的检测。如果实施的话，这些努力可以为 COVID-19 产生有效的泛变异疗法。