Reagents to Chemically Tag Specific Coronavirus Spike Proteins

化学标记特定冠状病毒刺突蛋白的试剂

• 批准号: 10259048
• 负责人: STEVEN A BENNER
• 金额: $ 25.89万
• 依托单位: FIREBIRD BIOMOLECULAR SCIENCES, LLC
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-07 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10259048
• 关键词: 2019-nCoV; Acylation; Affinity; Aminoacylation; Antibodies; Antigens; Antiviral Agents; Arboviruses; Architecture; Asia; Binding; Binding Proteins; Biological Assay; Biological Sciences; Breast Cancer Cell; COVID detection; Cell surface; Chemicals; Chemistry; Clinical Laboratory Information Systems; Coronavirus; Coronavirus Infections; Coronavirus spike protein; Crystallization; Cystic Fibrosis; DNA; Detection; Development; Diagnostic; Disulfides; Electrodes; Ensure; Environment; Europe; Evolution; Fright; Generations; Genes; Image; In Vitro; India; Individual; Information Systems; Institutional Review Boards; Laboratories; Lateral; Lung diseases; Lysine; Malignant neoplasm of liver; Measures; Medical; Middle East Respiratory Syndrome; Molecular Conformation; Norovirus; Nucleic Acids; Nucleotides; Oxidation-Reduction; Peptides; Performance; Phase; Preparation; Procedures; Protein Engineering; Proteins; Publishing; RNA; Reaction; Reagent; Regulation; Reproducibility; Research; Saliva; Sampling; Schools; Science; Severe Acute Respiratory Syndrome; Specificity; Structure; Surface; Testing; Time; Toxin; Vaccines; Virion; Virus; Work; amino group; analog; antibody test; aptamer; base; cancer cell; catalyst; covalent bond; disulfide bond; experience; genetic information; improved; innovation; instrument; instrumentation; novel coronavirus; nuclease; pandemic disease; recruit; respiratory pathogen; response; symptom treatment; tool

Reagents to Chemically Tag Specific Coronavirus Spike Proteins Firebird Biomolecular Sciences, LLC. Steven A. Benner Bharat Gawande Abstract While vaccines and antivirals may be long term solutions to the current coronavirus pandemic, it is now widely appreciated that the pandemic can be managed before these emerge by rapid, point-of-sampling "public space entry tests" (PSETs), given appropriate FDA regulatory relief. PSETs will be even more important if a good vaccine never emerges, a possibility given experience with other coronaviruses. In the 2003 SARS crisis, we delivered a "best in class" PCR kit for SARS that targeted coronaviral RNA. This was possible due to our first generation nucleic acid innovations, which were also used in respiratory pathogen panels and cystic fibrosis tests, inter alia. Luminex acquired EraGen in 2011 for $34 million. Since then, the PI generated 2nd and 3rd generation innovations that supported tests for MERS, arboviruses, norovirus and, this year, CoV-19 itself. These include a PSET that gives 10 minute results using proprietary displaceable probe loop amplification. Intrinsic chemistry suggests, however, this is the fastest any RNA-targeted test can be. Here, we will use our 3rd-generation DNA innovations not to target coronaviral RNA, but the coronavirus itself. Firebird will create reagents (AEGISZymes) from an artificially expanded genetic information system (AEGIS) that catalyze covalent acylation of lysines on the surface of ~300 spike proteins per virus. Each acylation will generate ~300 redox active moieties on an electrode surface per virion. The test is conceptually advanced over antibody tests, as it uses stable covalent bond formation, rather than non-covalent antibody: antigen interactions, and exploits electrochemical detection rather than a lateral flow. Our Phase 2 partner, MightyGate, has shown with standard aptamers detection in less than a minute with such architectures. With >100x sensitivity over Abbott BinaxNOWTM, the assay should detect virus in any contagious individual at entrances to arenas, schools, and other spaces as fast has handbags and are checked. Specifically, we will: Aim 1. Under IRB 2020001, we will use AEGIS-LIVE to evolve AEGISZymes in saliva that acylate lysines in spike proteins and peptide loops of CoV-19, SARS, and MERS. Aim 2. Metric the AEGISZymes that emerge, measuring their binding affinities, acylation rates, and specificity among the 3 homologous surface loops. The metrics are: M1. Acylation with kcat ≈ 1 sec-1. This should be readily achieved based on pmolar affinities that are now routine with AEGIS-LIVE; similar rate constants are now routine in analogous catalysts. M2. Specificity with kcat/Kdiss ratios >103, to ensure that CoV-19 is distinguished from other coronaviruses. If the metrics are met, MightyGate has committed itself to participate in Phase 2 work that will incorporate AEGISZymes onto its kiosk-style instrument. The kiosk has been proven with standard aptamers to give electrochemical readouts in less than 1 minute. Firebird's AEGISZymes will immediately improve the sensitivity of that kiosk, and immediate commercial value. However, our approach is general to any virus target.

化学标记特异性冠状病毒刺突蛋白的试剂 火鸟生物分子科学有限责任公司。 Steven A. Benner 巴拉特·加万德 摘要 虽然疫苗和抗病毒药物可能是目前冠状病毒大流行的长期解决方案，但现在 人们普遍认识到，通过快速、定点采样的“公众”， 空间进入试验（PSET），给予适当的FDA监管救济。PSET将更加重要，如果 好的疫苗永远不会出现，考虑到其他冠状病毒的经验，这是一种可能性。 在2003年的SARS危机中，我们推出了针对冠状病毒RNA的SARS“一流”PCR试剂盒。这 由于我们的第一代核酸创新，这也被用于呼吸道病原体 面板和囊性纤维化测试，阿利亚是。Luminex于2011年以3400万美元收购了EraGen。从那时起，PI 产生了第二代和第三代创新，支持对MERS、虫媒病毒、诺如病毒的检测， 今年，CoV-19本身。其中包括PSET，使用专有的可移位探针循环提供10分钟结果 放大然而，内在化学表明，这是任何RNA靶向测试可以达到的最快速度。 在这里，我们将使用我们的第三代DNA创新，而不是针对冠状病毒RNA，而是冠状病毒 本身Firebird将从人工扩展的遗传信息系统中创建试剂（AEGISZymes） （AEGIS），其催化每个病毒约300个刺突蛋白表面上的赖氨酸的共价酰化。每个 酰化将在每个病毒体的电极表面上产生约300个氧化还原活性部分。该测试在概念上 先进的抗体测试，因为它使用稳定的共价键形成，而不是非共价抗体： 抗原相互作用，并利用电化学检测，而不是横向流动。我们的第二阶段合作伙伴， MightyGate已经展示了使用这种架构在不到一分钟的时间内进行标准适体检测。 该检测试剂盒的灵敏度超过Abbott BinaxNOWTM 100倍， 进入体育场，学校和其他空间一样快有手提包，并检查。具体而言，我们将： 目标1。根据IRB 2020001，我们将使用AEGIS-LIVE在唾液中进化AEGISZymes， CoV-19、SARS和MERS的刺突蛋白和肽环。 目标二。测量出现的AEGISZymes，测量其结合亲和力、酰化速率和特异性 在3个同源表面环中。这些指标是： M1.用kcat进行酰化反应，反应时间为1 sec-1。这应该很容易实现的基础上pmolar亲和力，现在是 常规与AEGIS-LIVE;类似的速率常数，现在常规在类似的催化剂。 M2.特异性：kcat/Kdiss比值>103，以确保CoV-19与其他冠状病毒区分开来。 如果指标得到满足，MightyGate承诺参与第二阶段的工作， AEGISZymes在其亭式乐器上。该信息亭已被证明与标准适体， 电化学读数在不到1分钟。Firebird的AEGISZymes将立即提高灵敏度 和即时的商业价值。然而，我们的方法对任何病毒目标都是通用的。