Konry Tania, Sulllivan Matthew, Rozzo Aldo, Ward Antonio, Rao Patricia, Soler-Ferran Dulce, Greenberg Steven
Northeastern University.
Abcuro, Inc.
Res Sq. 2024 Sep 8:rs.3.rs-4745216. doi: 10.21203/rs.3.rs-4745216/v1.
Progress in developing improvements in the treatment of autoimmune disease has been gradual, due to challenges presented by the nature of these conditions. Namely, the need to suppress a patient's immune response while maintaining the essential activity of the immune system in controlling disease. Targeted treatments to eliminate the autoreactive immune cells driving disease symptoms present a promising new option for major improvements in treatment efficacy and side effect management. Monoclonal antibody therapies can be applied to target autoreactive immune cells if the cells possess unique surface marker expression patterns. Killer cell lectin like receptor G1 (KLRG1) expression on autoreactive T cells presents an optimal target for this type of cell depleting antibody therapy. In this study, we apply a variety of in vitro screening methods to determine the efficacy of a novel anti-KLRG1 antibody at mediating specific natural killer (NK) cell mediated antibody-dependent cellular cytotoxicity (ADCC). The methods include single-cell droplet microfluidic techniques, allowing timelapse imaging and sorting based on cellular interactions. Included in this study is the development of a novel method of sorting cells using a droplet-sorting platform and a fluorescent calcium dye to separate cells based on CD16 recognition of cell-bound antibody. We applied this novel sorting method to visualize transcriptomic variation between NK cells that are or are not activated by binding the anti-KLRG1 antibody using RNA sequencing. The data in this study reveals a reliable and target-specific cytotoxicity of the cell depleting anti-KLRG1 antibody, and supports our droplet-sorting calcium assay as a novel method of sorting cells based on receptor activation.
由于自身免疫性疾病的本质所带来的挑战，在开发改善其治疗方法方面的进展一直很缓慢。也就是说，需要在抑制患者免疫反应的同时，维持免疫系统在控制疾病方面的基本活性。针对消除引发疾病症状的自身反应性免疫细胞的靶向治疗，为大幅提高治疗效果和管理副作用提供了一个有前景的新选择。如果自身反应性免疫细胞具有独特的表面标志物表达模式，单克隆抗体疗法就可以用于靶向这些细胞。自身反应性T细胞上的杀伤细胞凝集素样受体G1（KLRG1）表达，是这类细胞耗竭性抗体疗法的一个理想靶点。在本研究中，我们应用了多种体外筛选方法，以确定一种新型抗KLRG1抗体介导特异性自然杀伤（NK）细胞介导的抗体依赖性细胞毒性（ADCC）的效果。这些方法包括单细胞液滴微流控技术，可实现延时成像并基于细胞间相互作用进行分选。本研究还包括开发一种使用液滴分选平台和荧光钙染料基于细胞对结合抗体的CD16识别来分选细胞的新方法。我们应用这种新型分选方法，通过RNA测序来可视化结合抗KLRG1抗体后被激活或未被激活的NK细胞之间的转录组差异。本研究中的数据揭示了细胞耗竭性抗KLRG1抗体具有可靠且靶向特异性的细胞毒性，并支持我们的液滴分选钙测定法作为一种基于受体激活来分选细胞的新方法。
