EVOLVING VIRUS-SPECIFIC sACE2 MIMICS FOR COMPETITIVE INHIBITION OF SARS-CoV-2

项目摘要 高致病性的新型SARS冠状病毒2（SARS-CoV-2）的快速传播引起了全球范围的关注， 健康紧急情况。因此，迫切需要有效的抗病毒治疗剂来对抗这种病毒。 SARS-CoV-2病毒通过结合病毒刺突蛋白2的ACE 2受体1进入细胞。在其 可溶性形式的ACE 2（sACE 2）具有用作稳定的和非免疫原性的竞争性ACE抑制剂的潜力。 SARS-CoV-2的抑制剂，目前正在临床试验中探索3。由于潜在的负面影响 抗尖峰单克隆抗体的作用18，以及ACE 2表现出其他生物学作用的事实4 -6，包括整合素 信号调节7，8，刺突特异性受体模拟物将产生新的治疗SARS-CoV-2和 其他潜在的高传染性疾病。 该提案寻求使用机器学习和定向进化来开发高亲和力， 内源性无活性的sACE 2模拟物，以创造快速可实施的治疗方法， SARS-CoV-2和潜在的冠状病毒。这种方法将允许生成可扩展的 可翻译的生物制剂，并提供一个平台，以快速纠正可能出现的潜在突变 在未来利用UniRep 49的深度学习，将设计和生成紧密结合的sACE 2变体 SARS-CoV 2 -2刺突蛋白，但不与内源性靶点如整联蛋白交叉相互作用[目的1]。 同时，我们将进行定向进化以优化尖峰结合，并选择 结合内源性蛋白[目的2]。最后，我们将确定主要候选人，并评估耐受性， 小鼠中工程化sACE 2变体的免疫原性[目的3]。总的来说，这一建议将发展 高度特异性ACE 2受体模拟物，以便及时创造具有最低免疫原性的新型抗病毒药物 以挽救生命并防止未来的疫情爆发。 10

PROJECT SUMMARY The rapid spread of the highly-pathogenic, novel SARS-coronavirus 2 (SARS-CoV-2) has caused a global health emergency. Thus, there is a desperate need for effective antiviral therapeutics to counteract this virus. The SARS-CoV-2 virus enters cells using the ACE2 receptor1 which binds the viral spike protein2. In its soluble form, ACE2 (sACE2) has the potential to be used as a stable and non-immunogenic competitive inhibitor to SARS-CoV-2 and is presently being explored in clinical trials3. Due to the potential negative side effects of anti-spike mAbs18, and the fact that ACE2 exhibits other biological roles4–6 including integrin signaling regulation7,8, spike-specific receptor mimics would yield novel therapeutics for SARS-CoV-2 and potentially other highly infectious diseases. This proposal seeks to use machine learning and directed evolution to develop high affinity, yet endogenously-inactive mimics of sACE2 in order to create rapidly implementable therapeutics to combat SARS-CoV-2 and potential corona-like viruses. This approach would allow for the generation of scalable and translatable biologics, and provide a platform to rapidly course-correct for potential mutations that may arise in the future. Utilizing deep-learning with UniRep49, will design and generate sACE2 variants that tightly bind the SARS-CoV2-2 spike protein but do not cross-interact with endogenous targets such as integrins [Aim 1]. Simultaneously, we will perform directed evolution to optimize spike-binding and select against variants that bind endogenous proteins [Aim 2]. Finally, we will identify lead candidates and evaluate the tolerance and immunogenicity of engineered sACE2 variants in mice [Aim 3]. Collectively, this proposal will develop highly-specific ACE2 receptor mimics in order to create novel antivirals with minimal immunogenicity in time to save lives and prevent future outbreaks. 10