574. Early Stage Product Development to Scale-Up: Use of Mini-Fermentation System for Screening and Process Development of Plasmid DNA Products

Abstract

Top of pageAbstract Introduction: As the number of gene therapy studies and trials has increased in the past few years, an ever-increasing demand for plasmid DNA-based products exists. In this expanding field of plasmid production, emphasis has always been placed on the quality and safety of the plasmid product. Much of the focus has been placed on increasing the expression of these products, while few focus on the early-stage screening and development of efficient plasmid production. The typical transition of a plasmid from research to the manufacturing group begins with shaker flask culture productions, which often yield little information on the cell growth and yield characteristics. Scale-up of these shaker flask-derived products has been difficult to achieve, and there is a need to translate the results from these cultures to lab scale and pilot or production scale. Adequate information of growth and yield characteristics for various plasmid products is important in the early developmental stage of any product. Method: We have successfully demonstrated the use of the Sixfors|[trade]| mini-fermentation system for the efficient screening of early-stage development products along with process development of plasmid DNA production and the ability to scale up the process to more than 100L. The Sixfors|[trade]| mini-fermenters, with a maximum working volume of 500mL, have been configured to mimic a large-scale fermenter with efficient control of dissolved oxygen. These fermenters were used for process development of plasmid DNA production. Their use has been established in the screening of bacterial strains and the optimization of fermentation media and process parameters using statistical experimental design techniques. Results: Using the mini-fermenter system, we were able to achieve early-stage predictability of the use of bacterial strains and growth conditions for growing a particular plasmid along with the scalability of fermentation conditions. The use of statistical design techniques for optimizing media formulations and process conditions resulted in the optimum process conditions in a limited amount of time with excellent reproducibility. The studies conducted to demonstrate the successful scale-up of the fermentation process indicated that the data obtained from the mini-fermenters are reproducible even at larger scales of operation. The use of this mini-fermenter system resulted in overall reduction of the time required for the completion of plasmid DNA production. Conclusions: The use of Sixfors|[trade]| has been shown as an excellent tool for early stage process development, including plasmid DNA screening studies. This method is far superior to performing early experiments with shaker flasks and is an ideal replacement for using these cultures for screening studies. The process using this Sixfors|[trade]| system has been tested in the lab scale (10L) and the pilot scale (100L) fermenters and has been shown to scale-up with excellent reproducibility.