Therapeutic Antibodies for Rabies

描述（由申请人提供）：Trellis Bioscience已将其专利CellSpot技术发展成为一个强大的平台，用于表征单个人类淋巴细胞分泌的抗体对多种抗原的特异性和亲和力。例如，该平台被用于分离一种罕见的、高质量的呼吸道合胞病毒（RSV）人抗体，该抗体目前正准备由Trellis的许可方MedImmune进行人体临床试验。在这个项目中，我们建议使用CellSpot分离抗狂犬病毒感染的治疗性抗体。接种过RV疫苗的成年人的血液淋巴细胞将被分层到抗体捕获表面。孵育5小时后，每个细胞分泌的抗体在细胞附近形成一个“足迹”（直径约150 <m）。然后将这些足迹与抗原靶标结合的荧光珠孵育，在本项目中抗原靶标将是RV的G蛋白。G蛋白与狂犬病疫苗中使用的抗原相同，已被证明可引发保护性抗体。不同菌株的G蛋白存在差异，不同RV菌株的G蛋白既有同源的保守区，也有异源的可变区。最有用的单克隆抗体是那些对最广泛的变异G蛋白起反应的抗体。因此，抗体足迹将通过与不同菌株的g抗原结合的一组可区分的荧光珠来筛选。在自动显微镜检查下检测含有多种G蛋白抗体的斑点，分离相关细胞，然后对抗体进行cDNA克隆，在转染到表达系统后进行进一步表征。这些抗体将通过体外中和试验和体内动物感染模型来筛选生理活性，以测量对感染的直接保护。在该项目的第一阶段，我们预计将分离出约15种针对G蛋白的人抗体，其中至少一种抗体将在动物模型中显示出保护作用，从而证实单克隆抗体的实用性，可与免疫γ球蛋白（目前的护理标准）相媲美。基于这一I期目标的成功，我们将提交II期申请，将该抗体的开发推进到临床试验，特别关注降低制造成本，以与免疫γ球蛋白竞争。该项目的总体目标是一种抗RV的治疗性抗体，这种抗体具有良好的耐受性，并且对多种RV菌株具有高效的保护作用。

DESCRIPTION (provided by applicant): Trellis Bioscience has developed its patented CellSpot" technology into a robust platform for characterizing the secreted antibody from single human lymphocytes with regard to specificity and affinity against multiple antigens. For example, the platform was used to isolate a rare, high quality human antibody to Respiratory Syncytial Virus (RSV) that is now being prepared for human clinical testing by Trellis' licensee, MedImmune. In this project, we propose to use CellSpot to isolate therapeutic antibodies to combat rabies virus (RV) infections. Blood lymphocytes from adults who have been vaccinated against RV will be layered onto an antibody capture surface. After 5 hours of incubation, each cell's secreted antibody forms a "footprint" in the vicinity of the cell (~150 <m diameter). These footprints will then be incubated with fluorescent beads conjugated to the antigen target, which in this project will be the G protein of RV. The G protein is the same antigen used in rabies vaccines and has been shown to elicit protective antibodies. There are differences between G proteins of different strains, and there are both homologous, conserved regions and heterologous, variable regions on the G proteins of different RV strains. The most useful monoclonal antibodies are those which react against the broadest set of variant G proteins. Therefore, the antibody footprints will be screened with a panel of distinguishable fluorescent beads conjugated to the G-antigens from different strains. Spots that contain antibody to multiple G proteins will be detected under automated microscopic examination and the responsible cells will be isolated, followed by cDNA cloning of the antibody for further characterization following transfection into an expression system. These antibodies will be screened for physiological activity both by an in vitro neutralization assay and by an in vivo animal infection model to measure direct protection from infection. In Phase I of this project, we anticipate isolating ~15 human antibodies to the G protein, of which at least one will be shown to be protective in the animal model, confirming utility by a monoclonal comparable to that provided by immune gamma globulin, the current standard of care. Based on success in this Phase I objective, we will submit a Phase II application to advance the development of this antibody to clinical trial with a particular focus on reducing the cost of manufacturing to be competitive with immune gamma globulin. The overall objective of this project is a therapeutic antibody to RV which is well tolerated and highly effective in protecting against a broad range of RV strains.