重组质粒pUDK-HGF 的中试纯化工艺

Pilot-scale production of recombinant plasmid pUDK-HGF

摘要:

pUDK-HGF 是携带人肝细胞生长因子的裸质粒，目前已进入I 期临床试验，因此需要大量符合药学规格的质粒DNA。文中建立了pUDK-HGF 中试规模纯化制备的新工艺。流程包括：发酵、离心收获菌体、碱裂解、超滤浓缩碱裂解液、Sephacryl S-1000 层析除去RNA 并更换缓冲液、plasmidselect 捕获超螺旋质粒DNA、琼脂糖凝胶6BFF 除盐。新工艺可获得浓度为2.0 mg/mL、纯度在1.70 以上的裸质粒原液，符合相关质量标准，并避免使用动物源性的酶及有毒试剂。

Abstract:

pUDK-HGF, the recombinant plasmid DNA encoding human hepatocyte growth factor (HGF), can treat ischaemic disease. A great quantity of pharmaceutical pUDK-HGF is needed. A pilot-scale production process of pUDK-HGF was established based on a new chromatographic media (plasmidselect), including fermentation, cell harvesting, alkaline lysis, ultrafiltration, RNA removing and buffer exchanging on Sephacryl S-1000, capturing supercoiled plasmid DNA with plasmidselect, and removing the salt with Sepharose 6BFF. The process does not use RNase enzyme and toxic solvents.