High throughput camelid antibody screening as a drug discovery platform

 描述(申请人提供)：单抗用于治疗从癌症到传染病的多种疾病。然而，开发具有高亲和力和特异性的抗体仍然是耗时、费力和不可预测的，特别是针对低免疫原性或毒性靶点。迫切需要一种新的方法来快速开发针对各种疾病的抗体，特别是针对新出现的疾病的抗体。Abzyme Treateutics LLC正在积极开发无动物抗体发现平台，以加快诊断和治疗抗体的生成。第一阶段的目标已经圆满完成。具体地说，我们创建了一种所谓的酵母表面展示骆驼自我多样化VHH抗体库或SDALib，作为一种在体外提供多种抗体的单抗生成系统，而不使用体内免疫。该系统的主要特点是通过抗体编码基因的超突变诱导，自生成具有多种抗体的文库。作为原理证明，我们成功地利用该平台分离了一组抗流感H5N1病毒N1神经氨酸酶(NA)的单域抗体。获得的VHH抗体已被证明在细菌系统中高效表达，为加速抗体生产的治疗应用提供了机会。纯化了抗NA VHH抗体，并对其进行了体外鉴定。作为原理的证明，几种与H5N1和H1N1流感病毒的N1 Nas反应的抗NA VHH抗体中的一种(Flu27-8)已经人源化，保留了亲本分子的全部抗NA活性。在细菌系统中以线性融合形式产生的双价抗NA抗体在体外检测中显示抗体活性显著增加。第二阶段的后续工作将同时进行三个具体目标：(I)通过展示在体外和体内对H5N1和H1N1流感病毒的活性，推动在第一阶段分离的N1抗NA抗体走向临床应用；(Ii)扩大该平台的应用，以开发针对其他流感病毒亚型的血凝素和NNA的抗体，例如具有不寻常的人畜共患病潜力的H3N2和H7N9，并针对几种不同神经氨酸酶(H5N1、H1N1、H3N2和H7N9)的抗体；(Iii)利用抗体发现平台开发针对一组癌症靶点的VHH抗体，然后生产新的类似免疫球蛋白的VHH双特异性人源化抗体。第二阶段的工作是进一步扩大快速开发新抗体至治疗目标的技术应用，并将第一阶段分离的人源化抗流感抗体推进到临床研究中。

 DESCRIPTION (provided by applicant): Monoclonal antibodies are used for treatment of a wide range of diseases from cancer to infectious diseases. However, development of antibodies with high affinity and specificity is still time- consuming, labor-intensive and unpredictable, especially to low immunogenic or toxic targets. A new approach for rapid development of antibodies against a variety of diseases, especially against emerging diseases is much needed. Abzyme Therapeutics LLC is actively developing animal-free antibody discovery platforms to accelerate generation of diagnostic and therapeutic antibodies. Phase I objectives have been successfully completed. Specifically, we have created a so-called yeast surface display camelid Self-Diversifying VHH Antibody Library or SDALib as a monoclonal antibody generating system providing a diverse array of antibodies in vitro, without using in vivo immunization. The main feature of the system is the ability to self-generate a library with diversified antibodies by mean of hypermutation induction in antibody encoding genes. As proof-of-principle, we have successfully used the platform to isolate a set of single-domain antibodies against N1 neuraminidase (NA) derived from influenza H5N1 virus. Obtained VHH antibodies have been shown to express efficiently in a bacterial system providing an opportunity for accelerated scale-up of antibody production for therapeutic application. Anti-NA VHH antibodies have been purified and characterized in in vitro assays. As proof of principle one (Flu27-8) out of several anti-NA VHH antibodies, that is reactive with N1 NAs of both H5N1 and H1N1 influenza viruses, has been humanized preserving the full anti-NA activity of the parental molecule. Bi- valent anti-NA antibodies that have been produced in a bacterial system as linear fusions showed a significant increase in antibody activity in in vitro assays. Subsequent efforts of Phase II consis of three specific aims that will be performed simultaneously: (i) advance the N1 anti-NA antibodies isolated in Phase I toward clinical application by demonstrating both in vitro and in vivo activity against H5N1 and H1N1 influenza viruses; (ii) expand the application of the platform for developing antibodies targeting both hemagglutinin and NAs of other influenza virus subtypes, e.g. H3N2 and H7N9 which have unusual zoonotic potential and aiming for antibodies cross-reacting with several different neuraminidases (H5N1, H1N1, H3N2 and H7N9); (iii) utilize the antibody discovery platform for developing VHH antibodies against a set of cancer targets followed by production of novel IgG-like VHH-based bispecific humanized antibodies. Phase II efforts are to further extend the application of the technology for rapid development of novel antibodies to therapeutic targets and to advance humanized anti-influenza antibodies isolated in phase I into clinical studies.