摘要
目的 用Monte Carlo方法确立IgG4抗体的复合型阴离子交换层析填料NM 90-Agarose TM HAM纯化工艺的设计空间,并进行验证。方法 利用全因子实验设计对影响复合阴离子交换层析的3个因素(上样pH值、上样电导、上样载量)进行筛选,建立模型。用Monte Carlo方法确定工艺的设计空间,并模拟设计空间内工艺稳健性,将确定的NM 90-Agarose TM HAM纯化工艺与亲和层析相结合,纯化中国仓鼠卵巢细胞(Chinese hamster ovary cells,CHO)表达的IgG4单抗样品,验证工艺参数的稳健性。结果 Monte Carlo方法确定了复合阴离子层析纯化的3个关键工艺参数的设计空间,其中上样液pH值设计空间为4.6±0.1,上样液电导值设计空间为(50±0.03)ms·cm^(-1),上样量≤60 mg·mL^(-1)填料;设计空间内参数的过程能力指数(capability index of process,CPK)≥1.22,且预测值与实际值的偏差<5%。Monte Carlo所建模型有较好的预测性能。设计空间内试验点达标,可以保证复合阴离子层析步骤单元关键质量属性指标的动态稳定。而设计空间范围外,运行1批次,部分指标不达标,表明此法建模可满足一定的指标预测,对工艺过程控制起到预警作用。同时与传统叠加法确定设计空间相比,在处理多个变量时,更为有效,简便,且提高了设计空间边缘的可靠程度。结论 用Monte Carlo模型评价IgG4抗体的复合型阴离子层析纯化工艺,是有效、简便、可行的。
OBJECTIVE To establish design space of the NM 90-AgaroseTM HAM purification process of IgG 4 antibody by Monte Carlo method and verified.METHODS Using the design of experiment(DOE)all-factor experimental design,the three factors(pH,conductivity,loading capacity)were screened,and the model was established.The design space of the process was determined by Monte Carlo and simulated process robustness in the design space,combining the defined NM90-AgaroseTM HAM purification process with affinity chromatography to purify IgG 4 mAb expressed by Chinese hamster ovary cells(CHO)to verify the robustness of the process parameters.RESULTS Monte Carlo determined the design space of three key process parameters purified by composite anion chromatography,in which the pH was 4.6±0.1,the conductivity was(50±0.03)ms·cm^(−1),the loading capacity was≤60 mg·mL^(−1);the complex process capability index(CPK)was≥1.22,and the deviationhe between the predicted value and the actual value was<5%.Monte Carlo had a relatively good prediction performance.The design space could ensure the dynamic stability of the critical quality attributes(CQA)of the process.However,outside the design space,one batch was run,and some of the indicators were unqualified,indicating that this method could meet certain index prediction and played an early warning role in process control.Compared with the traditional superposition method,it was more effective and simple to deal with multiple variables,and improved the reliability of the design space.CONCLUSION It is effective,simple and feasible to evaluate the purification process of composite anion chromatography by Monte Carlo model.
作者
白利利
靳征
张波
冯宇
王彩艳
BAI Lili;JIN Zheng;ZHANG Bo;FENG Yu;WANG Caiyan(Shenyang Sunshine Pharmaceutical Co.,Ltd.,Shenyang 110027,China)
出处
《中国现代应用药学》
CAS
CSCD
北大核心
2024年第18期2510-2519,共10页
Chinese Journal of Modern Applied Pharmacy
