Broad Spectrum Neutralizing Human Abs to SARS and Related Coronaviruses

描述（由申请人提供）：人单克隆抗体（mAb）疗法在病毒爆发环境中的预防、预防性和急性治疗方面具有相当大的优势，但由于长期存在的障碍，如循环病毒株的抗原变异性和中和逃逸，特别是对于RNA呼吸道病原体，如冠状病毒（cov）和流感，它作为治疗新出现的病毒病原体的药物的使用一直被拖延。重要的是，制造成本，一个历史上被认为是基于单克隆抗体的免疫疗法的限制，近年来急剧下降。在这里，Marasco、Baric和Liddington实验室建议开发平台，以SARS-CoV为模型，快速识别对新出现的流行病和人畜共患病池菌株有效的广谱中和性人类单克隆抗体（BnAbs）。该模型的主要优势包括强大的流行病和人畜共患病分离物序列数据库、合成基因组学和反向遗传学以及再现严重人类终末期肺病和急性呼吸窘迫综合征的年轻和老年小鼠体内发病模型；SARS-CoV受体ACE2及其与Spike蛋白结合的人bnab的原子结构我们的合作团队开发了先进的方法和新型试剂。我们的方法是基于我们发表的数据，即BnAbs可以被设计成防止病毒中和逃逸和/或减弱病毒进化。同时，从体外研究中获得的信息将用于制定新的战略，以最大限度地提高疫苗对更广泛的异质动物毒株的效力。这些研究的影响将是很高的，为其他重要的人类新兴病原体提供类似设计平台的模板。具体来说，我们将进一步建立我们的发现，即针对Spike蛋白上的受体结合基元（RBM）的强效人类抗sars单抗80R，以及80R的BnAb衍生物，这些衍生物随后被设计成对所有从人类和动物（蝙蝠除外）分离的sars冠状病毒具有活性。现在已经知道，许多人类致病性冠状病毒的天然宿主是蝙蝠，其中的祖先冠状病毒，包括蝙蝠- sars冠状病毒，可以复制、重组并重新出现为新的人类病原体。蝙蝠的sars冠状病毒库已经牢固建立，从公共卫生的角度来看，新出现和再出现的sars冠状病毒对人类健康构成了现实威胁。我们建议同时开发针对sars - cov保守的S2结构域的BnAbs，这可能具有更大的广度（包括针对未来可能从蝙蝠中出现的新型sars -样冠状病毒和其他致病性冠状病毒的活性）、中和效力和抵抗中和逃逸。在4个具体目标中，我们将检验12个假设，这些假设涉及定义针对sars - cov的新抗体、它们的中和表位、逃逸机制和防止中和逃逸的策略。在这些研究中解决的bnab - spike复合物的共晶结构将指导我们的病毒学研究和抗体工程工作。从体外中和逃逸研究中获得的信息也将用于设计RBM和s2导向的原型疫苗研究，以确定我们是否可以控制诱导的nab的类型、它们的中和广度和它们对中和逃逸的抗性，目的是诱导能够阻止病毒进化的BnAb反应。

DESCRIPTION (provided by applicant): Human monoclonal antibody (mAb) therapy offers considerable advantages for prophylaxis, preemptive and acute treatment in viral outbreak settings, yet its' use as a therapeutic agent against emerging viral pathogens has been protracted because of longstanding barriers such as antigenic variability of circulating viral strains and neutralization escape, particularly for RNA respiratory pathogens such as coronaviruses (CoVs) and influenza. Importantly, cost of manufacturing, a historically perceived limitation of mAb-based immunotherapy has dramatically decreased in recent years. Here, the Marasco, Baric and Liddington labs propose to develop platforms for rapidly identifying broad-spectrum neutralizing human mAbs (BnAbs) that are effective against emerging epidemic and zoonotic pool strains, using SARS-CoV as a model. Major strengths of this model include robust sequence database of epidemic and zoonotic isolates, synthetic genomics and reverse genetics and in vivo pathogenesis models in young and aged mice that recapitulate severe human end-stage lung disease and ARDS; atomic structures of SARS-CoV receptor ACE2 and human BnAbs bound to its Spike protein; and well developed methodologies and novel reagents developed by our collaborative team. Our approach is based on our published data that BnAbs can be designed to either prevent virus neutralization escape and/or attenuate virus evolution. Information gained from in vitro studies will be used to develop, in parallel, new strategies for maximizing vaccine efficacy against the broader heterogeneous pool of animal strains. The impact of these studies will be high, providing a template for similar design platforms for other important human emerging pathogens. Specifically, we will further build on our discovery of the potent human anti-SARS mAb 80R that is directed to the receptor binding motif (RBM) on Spike protein, and BnAb derivatives of 80R that have been subsequently engineered to be active against all SARS-CoV isolates from humans and animals (except bats). It is now known that the natural reservoir of many human pathogenic CoVs is the bat, where ancestral CoVs, including the bat-SARS-CoV, can replicate, recombine and reemerge as new human pathogens. The bat reservoir of SARS-CoVs is firmly established, and from the public health standpoint, emerging and re-emerging SARS-CoVs represent a real threat to human health. We propose to simultaneously develop BnAbs against the conserved S2 domain of SARS-CoVs, which may have even greater breadth (including activity against new SARS-like-CoVs and other pathogenic CoVs that might emerge from bats in the future), potency of neutralization and resistance to neutralization escape. In 4 specific aims, we will test 12 hypotheses that relate to defining new nAbs against SARS-CoVs, their neutralizing epitopes, escape mechanisms and strategies to prevent neutralization escape. Co-crystal structures of BnAbs-Spike complexes solved during these studies will guide our virologic studies and antibody engineering efforts. The information gained from in vitro neutralization escape studies will also be used to design prototypic RBM- and S2-directed vaccine studies to determine if we can control the type of nAbs that are elicited, their breadth of neutralization and their resistance to neutralization escape, with the goal of eliciting BnAb responses that can block virus evolution. PUBLIC HEALTH RELEVANCE: Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) is a novel virus that caused the global outbreak of SARS in 2002/03 with 10% mortality and a cluster of cases in 2003/04. Recently, it was discovered that bats harbor the ancestors of SARS-CoV and many if not all human CoV respiratory pathogens. Currently, there are no approved therapies for prevention and treatment of pathogenic CoV infections, which could reemerge at anytime and become a serious threat to human health. This project will develop a human antibody cocktail for this unmet need.