Analytical Development and Quality Control, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA.
J Immunol Methods. 2012 Nov 30;385(1-2):45-50. doi: 10.1016/j.jim.2012.08.004. Epub 2012 Aug 14.
Antibody dependent cell-mediated cytotoxicity (ADCC) is an important mechanism of action (MoA) for many monoclonal antibody (mAb) therapeutics. As such, quantitative measurement of ADCC activity is key to drug development. Traditional cell lysis based ADCC assays using PBMCs or NK cell lines can be challenging to develop and implement for routine testing. To provide an alternative to the cell lysis based ADCC assay, a non-cell based measure of ADCC activity was developed to determine the ADCC activity of an anti-CD20 mAb, which measures the ability of the mAb to bind to CD20 antigen and FcγRIIIa, simultaneously. The bridging of CD20 and FcγRIIIa is an essential interaction for the initiation of ADCC activity. This ADCC bridging method is simple, offers the ease of use of a standard ELISA, and shows reproducible dose-response curves in the concentration range of 50-1000 ng/mL. With interassay variability of 7-10% and recovery of 89-115%, the assay demonstrates acceptable precision and accuracy. The assay is able to detect degradative changes in anti-CD20 mAb samples subjected to light and acid exposure, suggesting that it is suitable for use as a stability-indicating method. The assay is also sensitive to mAb fucose levels. A linear relationship between the ADCC bridging assay and the cell lysis ADCC assay was demonstrated, strongly suggesting that the ADCC bridging assay can be used as a surrogate measure of ADCC.
抗体依赖的细胞介导的细胞毒性（ADCC）是许多单克隆抗体（mAb）治疗药物的重要作用机制（MoA）。因此，定量测量 ADCC 活性是药物开发的关键。使用 PBMC 或 NK 细胞系的传统基于细胞裂解的 ADCC 测定法在开发和实施常规测试方面可能具有挑战性。为了提供基于细胞裂解的 ADCC 测定法的替代方法，开发了一种非基于细胞的 ADCC 活性测量方法，以确定抗 CD20 mAb 的 ADCC 活性，该方法测量 mAb 与 CD20 抗原和 FcγRIIIa 结合的能力，同时。CD20 和 FcγRIIIa 的桥接是启动 ADCC 活性的必要相互作用。这种 ADCC 桥接方法简单，提供了标准 ELISA 的易用性，并在 50-1000ng/mL 的浓度范围内显示出可重复的剂量反应曲线。具有 7-10%的批间变异性和 89-115%的回收率，该测定法表现出可接受的精密度和准确性。该测定法能够检测到经受光照和酸暴露的抗 CD20 mAb 样品中的降解变化，表明它适合用作稳定性指示方法。该测定法也对 mAb 岩藻糖水平敏感。ADCC 桥接测定法和细胞裂解 ADCC 测定法之间显示出线性关系，强烈表明 ADCC 桥接测定法可用作 ADCC 的替代测量方法。
