Multivalent designed proteins neutralize SARS-CoV-2 variants of concern and confer protection against infection in mice.

New variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continue to arise and prolong the coronavirus disease 2019 (COVID-19) pandemic. Here we used a cell-free expression workflow to rapidly screen and optimize constructs containing multiple computationally designed miniprotein inhibitors of SARS-CoV-2. We found the broadest efficacy with a homo-trimeric version of the 75-residue angiotensin converting enzyme 2 (ACE2) mimic AHB2 (TRI2–2) designed to geometrically match the trimeric spike architecture. In the cryo-electron microscopy structure, TRI2 formed a tripod on top of the spike protein which engaged all three receptor binding domains (RBDs) simultaneously as in the design model. TRI2–2 neutralized Omicron (B.1.1.529), Delta (B.1.617.2), and all other variants tested with greater potency than that of monoclonal antibodies used clinically for the treatment of COVID-19. TRI2–2 also conferred prophylactic and therapeutic protection against SARS-CoV-2 challenge when administered intranasally in mice. Designed miniprotein receptor mimics geometrically arrayed to match pathogen receptor binding sites could be a widely applicable antiviral therapeutic strategy with advantages over antibodies and native receptor traps. By comparison, the designed proteins have resistance to viral escape and antigenic drift by construction, precisely tuned avidity, and greatly reduced chance of autoimmune responses. Computationally designed trivalent minibinders provide therapeutic protection in mice against emerging SARS-CoV-2 variants of concern.

严重急性呼吸综合征冠状病毒2（SARS - CoV - 2）的新变种不断出现，延长了2019冠状病毒病（COVID - 19）大流行。在此，我们使用无细胞表达流程来快速筛选和优化包含多种经计算机设计的SARS - CoV - 2微型蛋白质抑制剂的构建体。我们发现，一种由75个残基组成的血管紧张素转换酶2（ACE2）模拟物AHB2的同源三聚体版本（TRI2 - 2）具有最广泛的效力，其设计目的是在几何形状上与三聚体刺突结构相匹配。在冷冻电子显微镜结构中，TRI2在刺突蛋白顶部形成一个三脚架，如设计模型所示，它同时与所有三个受体结合域（RBDs）结合。TRI2 - 2对奥密克戎（B.1.1.529）、德尔塔（B.1.617.2）以及所有其他测试的变种的中和效力均高于临床上用于治疗COVID - 19的单克隆抗体。当在小鼠鼻内给药时，TRI2 - 2还对SARS - CoV - 2感染提供了预防和治疗保护。经计算机设计的微型蛋白质受体模拟物，其几何排列与病原体受体结合位点相匹配，可能是一种广泛适用的抗病毒治疗策略，具有优于抗体和天然受体陷阱的优势。相比之下，设计的蛋白质通过构建具有抵抗病毒逃逸和抗原漂移的能力、精确调整的亲和力以及大大降低自身免疫反应的可能性。 经计算机设计的三价微型结合物在小鼠中对新出现的令人担忧的SARS - CoV - 2变种提供治疗保护。