Production of pharmaceutical-grade plasmids at high concentration and high supercoiled percentage

Abstract The increased use of plasmid-based vaccines to replace their more challenging viral counterparts has increased the demand for high purity and high concentration plasmids. Here we report the production of plasmids encoding different transgenes for DNA vaccine candidates at gram scale with an integrated process consisting of batch fermentation and limited steps of purification. Plasmid products encoding for eight smallpox antigens that were combined into a bioterrorism DNA vaccine exhibited high purity with undetectable RNA, protein and endotoxin, concentration of up to 13.6 mg/mL and supercoiled percentage of 94.5 ± 1.1% after storage at −80 °C for over 1 year. The process has been scaled up for the cGMP manufacture of pharmaceutical-grade human papillomavirus and influenza DNA vaccines up to a 50 g scale, also demonstrating high purity and high concentration.