Wednesday 4th December – Day 2 Main ConferenceWednesday, 4 December 2024 - GMT (Greenwich Mean Time, GMTZ)

AAVrh74 has historically been chosen as the vector for gene therapies that treat muscular dystrophy. Due to the amount of muscle cells in the human body, it is critical to optimize the amount of vector generated in the manufacturing process, while minimizing COGs. To meet these clinical and commercial demands, the industry has moved away from adherent HEK293 cells in cell stacks and towards suspension HEK293 cells in closed bioreactor systems. In this presentation, we discuss upstream scaling challenges seen with our lead candidate, EPI-321, and some of the work executed to investigate this phenomenon.

For upstream process development engineers, scaling bioreactors presents a large challenge. It can be difficult to decide whether the best strategy is to scale up or to scale down. Both have their advantages, but scaling down is often preferred due to the ability to predict product quality in a faster and cheaper manner. A major hurdle in scaling is understanding how volume-dependent and volume-independent parameters in the process affect our cell culture consistency. To ensure consistent productivity, scaling strategies must be defined as early as possible in the development process. Upon the establishment of scale-down models, studies were executed to assess the impact of tip speed, transfection turbulence, and general optimization. After implementing changes to some critical parameters, we have shown successful scale-up of the upstream manufacturing process from bench-scale to 200 L production-scale.

Host Cell Proteins (HCPs) are critical process-related impurities in drug products that must be rigorously measured and monitored due to potential safety concerns. Unlike traditional biologics, gene therapy products present significant challenges in HCP quantitation and control because of their highly complex and variable manufacturing processes. Traditional methods for measuring HCPs, such as commercially available ELISA assays, may not accurately represent specific processes. Moreover, HCPs encompass both low and high molecular weight species, necessitating comprehensive detection methods to achieve sufficient coverage. Various techniques, including Antibody Affinity Extraction (AAE), 2D-Western analysis, and LC-MS-based approaches, can be employed for this purpose. This presentation will explore a case study on the roadmap for HCP analysis in an AAV-based gene therapy product, along with considerations for selecting specific methods at different stages of clinical development.