Pilot Purification Procedure And Tentative Quality Control Of Foxm1-based Recombinant Adenovirus Vector

Gene therapy technology has been accepted as one kind of therapy on genetics diseases. For more than twenty years, with the development of gene therapy technology, it has been widely applied in many kinds of disease treatment, especially for cancer gene therapy. Data from Worldwide Gene Therapy Clinical Trials Database show that adenoviruses now are the most commonly used vector in global cancer gene therapy technology. The deeper development of gene therapy, the more quantity and the higher quality we need for adenovirus vector, so large-scale clinical grade manufacturing of adenovirus vector is a challenge. Purification of adenovirus vector using two rounds of cesium chloride (CsCl) density gradient ultracentrifugation was not scalable, so liquid chromatography is emerging as the most powerful mode of purifier adenovirus vector. Recombinant adenovirus vector AdFoxMlshRNA has been structured based on FoxMl as a target gene that can suppress tumor growth. In this study we use AdFoxMl shRNA for the study of pilot purification procedure and tentative quality control of adenovirus vector. We established our cell banks of293A cell to ensure the stability and identity of our next resrarch. Then we use roller bottle system to culture293A cells, then infect cells, and collect supernatant fluid. In the downstream,we utilize ultrafiltration concentration、the anion exchange chromatography and gel filtration chromatography method to obtain a large-scale AdFoxMlshRNA.Finally, we detect viral purity determination、viral titer determination、replicative competent adenovirus testing comparison with the production gain from traditional method, and then make sure AdFoxM1siRNA from liquid chromatography has the superiority on the quality.