Gene Therapy Manufacturing 2.0: Manufacturing solutions that Work

Gene therapy and cell therapy commercialization is entering an exciting phase of expansion towards new indications that also brings development and manufacturing challenges. This presentation describes an overview of AAV and lentiviral manufacturing demands across both vector and virus production. Potential improvements in upstream, downstream, analytics and vector design will be explored.

In a joint collaboration, Oxford Biomedica and Repligen increased the yield of viral vectors from suspension cultured bioreactors several fold using tangential flow depth filtration (TFDF). The yield for a single clarification step was 95% as compared to 70% by standard depth filtration. The TFDF tubular format and low shear enabled further yield increases through multiple harvests from the same seeding. A single seeded bioreactor produced two harvest of 95% and 85%. The 180% yield represents a 2.5-fold yield increase over a single depth filtration harvest at 70%. The process was shown to scale from 5 to 50 L with an average flux of 750 LMH and three different lentiviral model systems, demonstrating broad applicability. Logistics and execution required a single filter, 3 AseptiQuik connections, typically consuming 15-20 minutes for set-up and breakdown and a four-fold reduction in required footprint as compared to conventional depth filtration

Demand for viral vector process development and manufacturing has soared in the last few years.  Adeno-associated virus (AAV), lentivirus, and related technologies are providing platform solutions for thousands of diseases for which there is no treatment or cure. Though upstream technologies for producing these critical vector-based cures may vary significantly, all methodologies have common downstream filter-based process elements including clarification, volume reduction, buffer exchange, formulation and sterilizing filtration. This discussion will focus on scale-down, scale-up and process characterization using Repligen KrosFlo® KR2i and KTF platforms employing tangential flow filtration (TFF) for clinical and commercial production of high-quality AAV-based viral vectors.

Towards increasing capacity, efficiency and process flexibility, hollow fiber and flat sheet cassettes were evaluated for three different gene therapy manufacturing unit operations. TFF based microfiltration using a hollow fiber filter was evaluated as an alternative to centrifugation for harvest. Both hollow fiber and flat sheet cassettes were evaluated for two distinct downstream UF/DF steps. Establishment of UF/DF methods for both hollow fiber and flat sheet platforms enables processes to be executed using the technology best suited for each product. Experimental design, example run data and the larger implications of process efficiency and flexibility will be discussed.

Microorganism, such as bacteria like E. coli, have several applications in the biopharma, food, and agriculture industry. Process monitoring and control are of the highest importance in the manufacturing of such microorganisms to allow good product quality when monitoring the growth kinetics of the cells. This limited linear range of traditional UV brings a challenge when adapting optical density measurements as an in-line or at-line monitoring tool for cell growth in bioreactors. The variable path-length technology from C Technologies, a Repligen company, overcomes this challenge by increasing the linear range between the absorbance and cell concentration.