Abstract:
The superior safety profile and transduction efficiency make AAV an excellent viral vector for human gene therapy. However, current AAV manufacturing suffers from low productivity and scalability, high raw material cost and supply chain complexity, and high batch-to-batch variation. To overcome these limitations and industrialize AAV manufacturing, we have generated a high-performing, helper virus-free, Xcite® AAV stable producer cell line (PCL). The cell line is built upon our proprietary high-producer HEK293 cells used for AAV transient transfection platform, grown in chemically defined, serum-free, ADCF medium in suspension. The DNA expression vectors are designed to tightly control viral gene expression via engineered inducible promoters, and stably integrated into the cell genome in one step by our VS piggyBacTM transposon system, which markedly increase the efficiency of cell line construction. Upon small chemical doxycycline induction at regular cell density, our selected PCL clones outperform transient transfection method with even higher AAV titer (>1E12 vg/mL) and comparable AAV genome packaging (>30% full capsids) at harvest. Moreover, these cells maintain the stability of AAV productivity and packaging over 30 cell passages corresponding the scale far beyond 2000 liters. Taken together, our Xcite® AAV stable PCL represents a next-generation platform that can enable high-quality commercial AAV manufacturing at large scale with significantly reduced cost and process complexity.
Author: Gang Li, PhD, R&D Scientist II