Depth filtration for clarification of intensified lentiviral vector suspension cell culture

Depth filtration significantly impacts efficiency of lentiviral (LV) vector purification process. However, it is often deprioritized in the overall scope of viral vector manufacturing process optimization. The demand for LV vectors has increased with the rise in disease indications, making it crucial to improve current manufacturing processes. Upstream bioreactor process intensification has enabled cell densities of over 107 viable cells/mL, creating challenges for harvest unit operations. The larger size of LV vectors and their physiochemical similarity to host cell-DNA (HC-DNA) and poor clarification performance causes significant challenges for the subsequent chromatography-based purifications. As a result, a robust and scalable harvest of LV process is needed, especially for LV in vivo therapeutic quality needs. In this study, we systematically evaluated the overlooked yet important issue of depth filtration systems to improve enveloped LV functional vector recovery. We found that an established depth filtration system in process A that provided 94% (n = 6) LV functional recovery could not be translated to intensified Process B cell culture. Hence, the depth filtration process became a bottleneck for the purification performance in an intensified process. We demonstrated an improvement in LV functional vector recovery from 34% to 82% via filter train optimization for an intensified suspension cell culture system (>107 cells/mL with higher titer), while still maintaining a loading throughput of ≥82 L/m2 and turbidity ≤20 NTU. It was demonstrated that the two or three-stage depth filtration scheme is scalable and more suitable for high cell density culture for large scale for LV manufacturing process.