Sun Ruoxuan, Qian Mark G, Zhang Xiaobin
Global Drug Metabolism, Pharmacokinetics & Modeling, Preclinical & Translational Sciences, Takeda Development Center Americas, Inc., Cambridge, Massachusetts 02139, United States.
Anal Chem. 2025 Jul 15;97(27):14273-14280. doi: 10.1021/acs.analchem.5c01017. Epub 2025 Jul 2.
Biotherapeutics have demonstrated remarkable therapeutic efficacy across a wide range of diseases; however, the desired clinical outcomes of biotherapeutics are sometimes hindered by the development of antidrug antibodies (ADAs). Traditionally, ADA monitoring relies heavily on ligand-binding assays (LBAs) such as enzyme-linked immunosorbent assays (ELISA) and electrochemiluminescence assays (ECLIA). Although effective, these methods require multiple washing and incubation steps, extending the data generation time to several hours or even days. In contrast, homogeneous immunoassays offer a promising alternative due to their "mix-and-read" feature, which significantly reduces operational complexity, time, and cost compared to conventional LBAs. Luciferase complementation represents an invaluable tool for visualizing protein-protein interactions in vitro, in cellulo, and in vivo. In this study, we explored the feasibility of rapid ADA detection using a wash-free homogeneous assay based on a split nanoluciferase (SplitNluc) biosensor system. Through case studies involving diverse therapeutic modalities, including antibody-drug conjugate (ADC), Fc-fusion protein, Fc-less multidomain biotherapeutic (MDB), and single-domain antibody (SdAb), we demonstrated that the SplitNluc platform is a versatile and efficient tool for ADA assay, offering ideal sensitivity, high precision and broad adaptability to various modalities. In summary, the SplitNluc assay represents an innovative and valuable platform for expedited and robust ADA detection. Moreover, our study highlighted the broader potential of luminescent biosensor-based approaches in advancing bioana-lytical workflows.
生物疗法已在多种疾病中展现出显著的治疗效果；然而，抗药抗体（ADA）的产生有时会阻碍生物疗法达到预期的临床疗效。传统上，ADA监测严重依赖配体结合分析（LBA），如酶联免疫吸附测定（ELISA）和电化学发光测定（ECLIA）。尽管这些方法有效，但需要多次洗涤和孵育步骤，将数据生成时间延长至数小时甚至数天。相比之下，均相免疫测定因其“混合即读”特性提供了一种有前景的替代方法，与传统LBA相比，显著降低了操作复杂性、时间和成本。荧光素酶互补是一种在体外、细胞内和体内可视化蛋白质 - 蛋白质相互作用的宝贵工具。在本研究中，我们探索了基于分裂纳米荧光素酶（SplitNluc）生物传感器系统的免洗均相测定法快速检测ADA的可行性。通过涉及多种治疗方式的案例研究，包括抗体药物偶联物（ADC）、Fc融合蛋白、无Fc多结构域生物疗法（MDB）和单结构域抗体（SdAb），我们证明SplitNluc平台是一种用于ADA测定的通用且高效的工具，具有理想的灵敏度、高精度以及对各种治疗方式的广泛适应性。总之，SplitNluc测定法代表了一个用于快速且可靠的ADA检测的创新且有价值的平台。此外，我们的研究突出了基于发光生物传感器的方法在推进生物分析工作流程方面的更广泛潜力。
