Zuo Chengyi, Zhou Jingwei, Bian Sumin, Zhang Qing, Lei Yutian, Shen Yuan, Chen Zhiwei, Ye Peijun, Shi Leying, Mu Mao, Qu Jia-Huan, Jiang Zhengjin, Wang Qiqin
Institute of Pharmaceutical Analysis, College of Pharmacy/State Key Laboratory of Bioactive Molecules and Druggability Assessment/Guangdong Province Key Laboratory of Pharmacodynamic Constituents of TCM and New Drugs Research of China, Jinan University, Guangzhou, 510632, China.
School of Engineering, Westlake University, Hangzhou, 310024, China.
J Pharm Anal. 2024 Nov;14(11):101015. doi: 10.1016/j.jpha.2024.101015. Epub 2024 Jun 4.
Dynamic tracking analysis of monoclonal antibodies (mAbs) biotransformation is crucial, as certain modifications could inactivate the protein and reduce drug efficacy. However, a particular challenge (i.e. immune recognition deficiencies) in biotransformation studies may arise when modifications occur at the paratope recognized by the antigen. To address this limitation, a multi-epitope affinity technology utilizing the metal organic framework (MOF)@Au@peptide@aptamer composite material was proposed and developed by simultaneously immobilizing complementarity determining region (CDR) mimotope peptide (HH24) and non-CDR mimotope aptamer (CH1S-6T) onto the surface of MOF@Au nanocomposite. Comparative studies demonstrated that MOF@Au@peptide@aptamer exhibited significantly enhanced enrichment capabilities for trastuzumab variants in comparison to mono-epitope affinity technology. Moreover, the higher deamidation ratio for LC-Asn-30 and isomerization ratio for HC-Asn-55 can only be monitored by the novel bioanalytical platform based on MOF@Au@peptide@aptamer and liquid chromatography-quadrupole time of flight-mass spectrometry (LC-QTOF-MS). Therefore, multi-epitope affinity technology could effectively overcome the biases of traditional affinity materials for key sites modification analysis of mAb. Particularly, the novel bioanalytical platform can be successfully used for the tracking analysis of trastuzumab modifications in different biological fluids. Compared to the spiked phosphate buffer (PB) model, faster modification trends were monitored in the spiked serum and patients' sera due to the catalytic effect of plasma proteins and relevant proteases. Differences in peptide modification levels of trastuzumab in patients' sera were also monitored. In summary, the novel bioanalytical platform based on the multi-epitope affinity technology holds great potentials for in vivo biotransformation analysis of mAb, contributing to improved understanding and paving the way for future research and clinical applications.
单克隆抗体（mAb）生物转化的动态跟踪分析至关重要，因为某些修饰可能会使蛋白质失活并降低药物疗效。然而，当修饰发生在抗原识别的互补决定区时，生物转化研究中可能会出现一个特殊的挑战（即免疫识别缺陷）。为了解决这一局限性，提出并开发了一种利用金属有机框架（MOF）@Au@肽@适配体复合材料的多表位亲和技术，该技术通过将互补决定区（CDR）模拟表位肽（HH24）和非CDR模拟表位适配体（CH1S-6T）同时固定在MOF@Au纳米复合材料表面。比较研究表明，与单表位亲和技术相比，MOF@Au@肽@适配体对曲妥珠单抗变体的富集能力显著增强。此外，只有基于MOF@Au@肽@适配体和液相色谱-四极杆飞行时间质谱（LC-QTOF-MS）的新型生物分析平台才能监测到LC-Asn-30较高的脱酰胺率和HC-Asn-55较高的异构化率。因此，多表位亲和技术可以有效克服传统亲和材料在mAb关键位点修饰分析中的偏差。特别是，该新型生物分析平台可成功用于不同生物流体中曲妥珠单抗修饰的跟踪分析。与加标的磷酸盐缓冲液（PB）模型相比，由于血浆蛋白和相关蛋白酶的催化作用，在加标的血清和患者血清中监测到了更快的修饰趋势。还监测了患者血清中曲妥珠单抗肽修饰水平的差异。总之，基于多表位亲和技术的新型生物分析平台在mAb体内生物转化分析方面具有巨大潜力，有助于增进理解，并为未来的研究和临床应用铺平道路。
