重组人尿激酶原纯化工艺病毒去除效果验证

Verification of Purification Procedure of Recombinant Human Prourokinase for Virus Removal

目的验证重组人尿激酶原(Prourokinase,Pro-uk)纯化工艺的病毒去除效果。方法采用模拟Pro-uk纯化工艺中阴离子和阳离子交换层析的scale-down模型,以水泡性口炎病毒(Vesivular stomatitis virus,VSV)、脑心肌炎病毒(Encephalo-myocarditis virus,EMCV)、人单纯疱疹病毒1型(Herpes simplex virus type-1,HSV-1)为指示病毒,采用细胞病变法评价离子交换层析的病毒去除效果。结果阴离子交换层析对脂包膜病毒VSV和HSV-1的去除能力强,对无脂包膜病毒EMCV的去除能力较弱;阳离子交换层析对VSV和HSV-1的去除能力较弱,对EMCV的去除能力较强。经离子交换层析纯化后,3种指示病毒的累计去除效率均>99.99%(大于4 log10)。结论 Pro-uk纯化工艺中的离子交换层析是控制病毒安全性的有效步骤,其去除效率达到了《生物组织提取制品和真核细胞表达制品的病毒安全性评价技术审评一般原则》规定的安全标准。

Objective To verify the purification procedure of recombinant human prourokinase（Pro-uk） for virus removal.Methods The effect of viral removal by ion exchange chromatography was evaluated by scale-down model simulating the anion and cation exchange chromatography in purification procedure of Pro-uk,using esivular stomatitis virus（VSV）,encephalomyocarditis virus（EMCV）,herpes simplex virus type-1（HSV-1） as indicators.Results Anion exchange chromatography was effective in removal of lipid-enveloped viruses such as VSV and HSV-1,while was less effective in non-lipid-enveloped EMCV.However,cation exchange chromatography was less effective in removal of VSV and HSV-1 while was effective in EMCV.All the cumulative removal efficacies of the three kinds of indicator viruses by ion exchange chromatography were more than 99.99%（more than 4 log10）.Conclusion The ion exchange chromatography step in purification procedure of Pro-uk was effective in control of safety of viruses,by which the removal efficacy of viruses met the General Requirements for Technical Evaluation on Virus Safety in Products Extracted from Organic Tissues and Expressed in Eukaryotic Cells.