Key Laboratory of the Ministry of Health for Research on Quality and Standardization of Biotech Products, National Institutes for Food and Drug Control, No. 2, Tiantan Xili, Dongcheng District, Beijing 100050, China.
Key Laboratory of the Ministry of Health for Research on Quality and Standardization of Biotech Products, National Institutes for Food and Drug Control, No. 2, Tiantan Xili, Dongcheng District, Beijing 100050, China.
J Chromatogr A. 2021 Apr 26;1643:462043. doi: 10.1016/j.chroma.2021.462043. Epub 2021 Mar 15.
An imaged capillary isoelectric focusing (icIEF) - UV fluorescence imaging detection method is described for the direct charge heterogeneity characterization of recombinant human erythropoietin (rhEPO) drug products (DPs). rhEPO is one of the most important protein therapeutics for biopharmaceutical industry worldwide. As a heavily glycosylated protein therapeutic, its charge heterogeneity must be carefully monitored in each step of manufacturing and storage. Current charge characterization methods suffer from challenges to characterize rhEPO DPs, due to low sensitivity of the method and potential for interference from the DP's formulation. The method described herein leverages the separation power of imaged cIEF separation combined with the increased sensitivity afforded by UV fluorescence imaging detection and requires no pre-treatment of the DP sample prior to analysis. The method was evaluated initially using a simulated DP, and subsequently a mini method validation was performed using a commercial rhEPO DP sample according to the guideline set by the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH). The limit of quantitation (LOQ) of the method is validated to be 20.3 IU/mL (or 0.10 µg/mL), which is approximately 100 times more sensitive than CZE - UV absorption detection method. To demonstrate the applicability of the method for use, 8 different commercial rhEPO DPs with concentrations ranging from 2000 IU/mL - 10,000 IU/mL were successfully evaluated. This method allows for sensitive, rapid analysis of low concentration rhEPO drug products without sample pre-treatment to provide critical charge heterogeneity information.
一种基于成像毛细管等电聚焦（icIEF）-紫外荧光成像检测的方法，用于直接分析重组人红细胞生成素（rhEPO）药物产品（DP）的荷质比异质性。rhEPO 是全球生物制药行业最重要的蛋白治疗药物之一。作为一种高度糖基化的蛋白治疗药物，其荷质比异质性必须在生产和储存的每一步都得到仔细监测。由于方法灵敏度低，以及 DP 配方可能存在干扰，目前的荷质比分析方法难以用于 rhEPO DP 的分析。本文所述的方法利用了成像 cIEF 分离的分离能力，结合紫外荧光成像检测的高灵敏度，在分析前无需对 DP 样品进行预处理。该方法首先使用模拟 DP 进行了评估，随后根据人用药物技术要求国际协调理事会（ICH）制定的指南，使用商业 rhEPO DP 样品进行了小型方法验证。该方法的定量限（LOQ）验证为 20.3 IU/mL（或 0.10 µg/mL），比 CZE-紫外吸收检测方法灵敏 100 倍。为了证明该方法在使用中的适用性，成功评估了浓度范围为 2000 IU/mL-10,000 IU/mL 的 8 种不同的商业 rhEPO DP。该方法无需样品预处理，即可对低浓度 rhEPO 药物产品进行灵敏、快速分析，提供关键的荷质比异质性信息。
