药用重组人IL-15蛋白及氧化还原副产物的HPLC及LC-MS分析

HPLC and LC-MS Analysis of Therapeutic rh IL-15 and Its Oxidation and Reduction By-products

重组人白细胞介素-15(rh IL-15)具有显著的T细胞及NK细胞增殖和活化功能,在肿瘤免疫治疗、疫苗佐剂等领域的临床应用前景良好,已有多个临床试验在进行中。为提高rh IL-15的质量控制水平,须开发快速、精准的检测方法。本研究针对rh IL-15纯化过程中的氧化还原副产物,开发了基于反相液相色谱(RP-HPLC)及液相色谱-质谱联用(LC-MS)技术的方法,通过分析色谱峰及相应的相对分子质量,以准确判断产品及其氧化还原状态。利用本方法能够分析rh IL-15生产工艺中的中间产品和终产品的质量,指导生产工艺的改进,提高rh IL-15产品的质量水平。

Interleukin-15 is a strong immune stimulant for T lymphocytes and NK ceils, and the recombinant human interleukin-15 （rhIL-15） has been a promising clinical candidate for tumor immunotherapy as well as being an effective vaccine adjuvant. To assist the pre-clinical and clinical development of rhIL-15, its preparation has been extensively studied in recent years. However, rhIL-15 is unstable and easy to be oxidized or reduced during manufacturing processes. Therefore, the quality control system with high sensitivity and precision is needed urgently. In this study, we adopted advanced RP-HPLC and LC-MS methods to analyze the oxidation and reduction by-products of rhIL-15. The results illustrated the significant differences between rhIL-15 and its oxidative or reductive derivatives. At most 3 oxygen or 4 hydrogen atoms were successfully determined by the intact molecular weights in LC-MS, which could precisely annotate the peaks separated by RP-HPLC. The method reported here is promising to be applied in the quality control of rhIL-15 manufacturing, helping to optimize the processes, thus improving the quality of rhIL-15 products.