Scientific Operations, Waters Corporation, Milford, MA, 01757, USA; Texas A&M University-San Antonio, One University Way, San Antonio, TX, 78224, USA.
National Institutes for Food and Drug Control (NIFDC), Tiantan Xili No. 2, 100050, Dongcheng District, Beijing, China.
Talanta. 2020 Dec 1;220:121335. doi: 10.1016/j.talanta.2020.121335. Epub 2020 Jul 6.
Recombinant human erythropoietin (rhEPO) is a glycoprotein that acts as the main hormone involved in regulating red blood cell production to treat anemia caused by chronic kidney disease or chemotherapy. Since the expiration of the patent of the innovator epoetin alfa, numerous rhEPO products have emerged in global markets. As described here, multiple complementary analytical approaches are utilized for the extensive characterization of rhEPO molecules, and more importantly for the structural comparison of the rhEPO analogues on the Chinese market. The focus of this study is placed on the overall glycosylation profiling, O-glycan profiling, and N-glycan mapping by UPLC-MS with an aim to develop an effective analytical methodology to monitor the product quality attributes of rhEPO analogues. Two rhEPO analogues manufactured in China were analyzed to demonstrate the principle of the developed methods. Each rhEPO product showed a characteristic glycoform profile with respect to the distribution of sialic acids across multi-antennary structures, the occurrence of O-glycosylation, O-acetylation on sialic acids, and the extension of N-glycan antennae with N-acetyllactosamine units. The study demonstrates that UPLC-MS is an effective analytical tool to characterize and monitor the glycosylation profiles among rhEPO analogues in order to detect and account for the divergence between rhEPO products, as well as the presence of unusual or unexpected glycans.
重组人促红细胞生成素（rhEPO）是一种糖蛋白，作为调节红细胞生成的主要激素，用于治疗慢性肾病或化疗引起的贫血。自从创新型促红细胞生成素阿尔法的专利到期后，全球市场上出现了许多 rhEPO 产品。如这里所述，多种互补的分析方法被用于广泛表征 rhEPO 分子，更重要的是，用于比较中国市场上 rhEPO 类似物的结构。本研究的重点是通过 UPLC-MS 进行全面的糖基化分析、O-聚糖分析和 N-聚糖图谱分析，旨在开发一种有效的分析方法来监测 rhEPO 类似物的产品质量属性。分析了两种中国制造的 rhEPO 类似物，以证明所开发方法的原理。每个 rhEPO 产品在唾液酸的多天线结构分布、O-糖基化、唾液酸化的 O-乙酰化以及 N-乙酰乳糖胺单元扩展 N-聚糖天线方面都表现出独特的糖型分布特征。该研究表明，UPLC-MS 是一种有效的分析工具，可用于表征和监测 rhEPO 类似物的糖基化谱，以检测和解释 rhEPO 产品之间的差异，以及异常或意外聚糖的存在。
