Antibody Engineering Program

虽然基于抗体的治疗方法已经成为癌症治疗的主要组成部分，但产生针对细胞表面受体和离子通道等重要靶点的抗体仍然很困难。这些蛋白质含有隐藏的功能位点，这些位点通常是传统的小鼠或人类基于igg的抗体无法到达的。单域抗体已显示出针对困难抗原和隐藏表位的有希望的能力。美国国家癌症研究所的Mitchell Ho博士已经证明，单域抗体能够靶向癌症信号复合物中隐藏的功能位点[Feng等人]。PNAS, 2013;Gao et al . Nature Communications, 2015；Li等人。PNAS, 2017;Li等人。肝脏病学,2019]。Ho实验室已经构建了大型鲨鱼和骆驼单域（“纳米体”）文库，并分离了多种抗原的结合物[Feng等人]。Antibody Therapeutics, 2019]，表明噬菌体显示的单域抗体文库可以成为分离治疗性抗体的有价值来源。AEP正在进行两个领域的研究，合作者使用何氏实验室创建的鲨鱼和骆驼单域噬菌体文库。一种是通过纳米体与MHC相关肽复合物的结合来靶向细胞内癌症靶点。另一种方法是将纳米体与拉沙病毒和SARS-CoV-2等重要和新出现的病毒分离开来。在21财年，AEP发表了两篇文章和其他几篇正在准备的手稿。其中一篇文章发表在Molecular Cancer Therapeutics [Duan and Ho Mol Cancer Ther 2021]。基于抗体的免疫疗法在各种癌症类型中显示出临床有效性。然而，靶标库仅限于表面或可溶性抗原，这是癌症蛋白质组中相对较小的百分比。人类蛋白质组的大多数蛋白质都是细胞内的。来自细胞内靶点的短肽可以通过细胞表面的主要组织相容性复合体I类（MHC-I）分子呈现，使其成为癌症免疫治疗的潜在靶点。可以开发针对这些肽/MHC复合物的抗体，类似于T细胞受体（TCR）对这些复合物的识别。这些抗体被称为T细胞受体模拟物（TCRm）或tcr样抗体。正在进行的临床前和临床研究将有助于了解它们的作用机制和免疫治疗目标表位的选择。我们的综述总结和讨论了细胞内抗原的选择、肽/MHC复合物的产生、用于治疗的TCRm抗体的分离、TCRm抗体的局限性以及在临床中推进基于TCRm抗体的方法的可能途径。在《Antibody Therapeutics》上发表的另一篇文章[Sun and Ho， Antib Ther 2020]中，我们总结了针对SARS-CoV-2的中和抗体的发展，重点讨论了各种抗体发现策略（动物免疫、噬菌体展示和B细胞克隆），描述了结合表位并比较了中和活性。针对SARS-CoV-2和SARS-CoV刺突蛋白的广谱中和抗体可能有助于治疗COVID-19和未来的感染。基于S309的VIR-7831/7832是临床后期开发的唯一抗体，虽然不能直接阻断病毒受体结合，但可以中和SARS-CoV-2和SARS-CoV。到目前为止，唯一的交叉中和抗体也是受体结合阻滞剂是纳米体VHH-72。开发纳米体作为治疗COVID-19和其他呼吸系统疾病的吸入药物的可行性是一个值得探索和测试的有吸引力的想法。鸡尾酒策略，如REGN-COV2，或工程多价和多特异性分子，结合两种或更多抗体可能提高功效，并防止病毒逃逸突变体产生耐药性。除了受体结合结构域，其他病毒抗原，如刺突蛋白的S2亚基和宿主细胞上的病毒附着位点，如硫酸肝素蛋白聚糖，也值得研究。

While antibody-based therapeutics have emerged as a major component in cancer treatment, the generation of antibodies to important targets such as cell surface receptors and ion channels remains difficult. These proteins contain buried functional sites that are often unreachable by classical mouse or human IgG-based antibodies. Single domain antibodies have shown a promising ability to target difficult antigens and hidden epitopes. Dr. Mitchell Ho at the NCI has demonstrated that single domain antibodies are capable of targeting buried functional sites in cancer signaling complexes [Feng et al. PNAS, 2013; Gao et al Nature Communications, 2015; Li et al. PNAS, 2017; Li et al. Hepatology, 2019]. The Ho lab has constructed large shark and camel single-domain ('nanobody') libraries and isolated binders to a wide range of antigens [Feng et al. Antibody Therapeutics, 2019], indicating that the phage-displayed single domain antibody libraries can be a valuable source to isolate therapeutic antibodies. Two areas of research are being pursued at the AEP with collaborators using the shark and camel single domain phage libraries created by the Ho lab. One is focused on targeting intracellular cancer targets via binding of nanobodies to the MHC associated peptide complexes. The other is to isolate nanobodies to important and emerging viruses such as Lassa virus and SARS-CoV-2. In FY21, the AEP published two articles and several other manuscripts in preparation. One article was published in Molecular Cancer Therapeutics [Duan and Ho Mol Cancer Ther 2021]. Antibody-based immunotherapies show clinical effectiveness in various cancer types. However, the target repertoire is limited to surface or soluble antigens which are a relatively small percentage of the cancer proteome. Most proteins of the human proteome are intracellular. Short peptides from intracellular targets can be presented by major histocompatibility complex class I (MHC-I) molecules on cell surface, making them potential targets for cancer immunotherapy. Antibodies can be developed to target these peptide/MHC complexes, similar to the recognition of such complexes by the T cell receptor (TCR). These antibodies are referred to as T cell receptor mimic (TCRm) or TCR-like antibodies. Ongoing preclinical and clinical studies will help understand their mechanisms of action and selection of target epitopes for immunotherapy. Our review summarized and discuss the selection of intracellular antigens, production of the peptide/MHC complexes, isolation of TCRm antibodies for therapeutic applications, limitations of TCRm antibodies, and possible ways to advance TCRm antibody-based approaches in the clinic. In the other article published in Antibody Therapeutics [Sun and Ho, Antib Ther 2020], we summarized the development of neutralizing antibodies against SARS-CoV-2, with a focus on discussing various antibody discovery strategies (animal immunization, phage display and B cell cloning), describing binding epitopes and comparing neutralizing activities. Broad-neutralizing antibodies targeting the spike proteins of SARS-CoV-2 and SARS-CoV might be helpful for treating COVID-19 and future infections. VIR-7831/7832 based on S309 is the only antibody in late clinical development, which can neutralize both SARS-CoV-2 and SARS-CoV although it does not directly block virus receptor binding. Thus far, the only cross-neutralizing antibody that is also a receptor binding blocker is nanobody VHH-72. The feasibility of developing nanobodies as inhaled drugs for treating COVID-19 and other respiratory diseases is an attractive idea that is worth exploring and testing. A cocktail strategy such as REGN-COV2, or engineered multivalent and multispecific molecules, combining two or more antibodies might improve the efficacy and protect against resistance due to virus escape mutants. Besides the receptor-binding domain, other viral antigens such as the S2 subunit of the spike protein and the viral attachment sites such as heparan sulfate proteoglycans that are on the host cells are worth investigating.