Improved system for helper-dependent adenoviral vector production

摘要: Helper-dependent adenoviral vectors (HDAds) are devoid of all viral coding sequences and have demonstrated tremendous potential for gene therapy by providing increased cloning capacity (up to 37 kb) long-term, high-level transgene expression in vivo with negligible toxicity. Currently, the most widely used method producing HDAds is Cre/loxP system developed Graham co-workers. However, two major obstacles currently hinder progress this promising technology: (1) difficulty large-scale vector production (2) helper virus (HV) contamination. We an improved producer cell line, HV, protocols that successfully addressed these problems. With system, >1 x 10(13) particles (vp) can be easily produced from 3 liters cells within 2 weeks rescue, specific yields >10,000 vp/cell exceedingly low HV contamination 0.4-0.1% without relying on density-based purification 0.02-0.01% following CsCl purification. This new represents a improvement over original terms simplicity, speed, yield, purity, it will significantly improve our ability assess technology, especially large animal models and, ultimately, clinical applications.