Reagents to Chemically Tag Specific Coronavirus Spike Proteins

化学标记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。 史蒂文·A·本纳 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，但现在是 人们普遍认识到，大流行可以在快速、抽样点的公众出现之前得到控制 太空进入试验“(PSETS)，考虑到FDA适当的监管放松。如果一个 好的疫苗永远不会出现，考虑到其他冠状病毒的经验，这种可能性是存在的。 在2003年的SARS危机中，我们提供了一种针对冠状病毒RNA的针对SARS的“同类最佳”聚合酶链式反应试剂盒。这 这要归功于我们的第一代核酸创新，它也被用于呼吸道病原体 小组和囊性纤维化测试，等等。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MERS、虫媒病毒、诺沃克病毒等测试 年，CoV-19本身。其中包括一个PSET，它使用专有的可移位探头回路提供10分钟的结果 放大。然而，内在化学表明，这是任何以RNA为目标的测试所能达到的最快速度。 在这里，我们将使用我们的第三代DNA创新不是针对冠状病毒RNA，而是针对冠状病毒 它本身。火鸟将从人工扩展的遗传信息系统中创造试剂(AEGISZymes) (宙斯盾)，催化每种病毒约300个刺突蛋白表面赖氨酸的共价酰化。每个人 在每个病毒粒子上，酰化反应将在电极表面产生约300个氧化还原活性部分。这项测试是概念性的 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，而不是侧向流动。我们的第二阶段合作伙伴， MightyGate已经在不到一分钟的时间内展示了使用这种架构进行标准适配子检测的情况。 由于&gt；对雅培BinaxNOWTM的灵敏度是100倍，该方法应该可以在以下位置检测到任何有传染性的个人中的病毒 场馆、学校和其他场所的入口处都有手提包，并进行了检查。具体来说，我们会： 目标1.根据IRB 2020001，我们将使用宙斯盾-LIVE进化唾液中使赖氨酸酰化的AEGISZyme CoV-19、SARS和MERS的刺突蛋白和肽环。 目的2.测量出现的AEGISZyme，测量它们的结合亲和力、酰化率和特异性 在3个同源表面环中。这些指标包括： M1。与KCAT≈的酰化反应1秒-1。这应该很容易实现，基于现在的磨牙亲和力 Aegis-live的例程；类似的速率常数现在类似的催化剂中是例行公事。 M2。KCAT/Kdiss比率&103的特异性，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已承诺参与第二阶段的工作，该阶段将包括 AEGISZymes安装到它的Kiosk式乐器上。Kiosk已经被证明具有标准的适配子来提供 不到1分钟的电化学读数。火鸟的AEGISZymes将立即提高灵敏度 以及直接的商业价值。然而，我们的方法对任何病毒目标都是通用的。

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.