BACK TO BASICS: A SCIENTIFIC AND ECONOMIC EVALUATION OF ADHERENT AND SUSPENSION PROCESSING FOR AAV AT SCALE

Background & Aim Adherent cultures demonstrate superior cell-specific productivity. The larger surface area available for cell attachment in adherent systems fosters higher cell densities and enhanced production of viral vectors. This heightened productivity is attributed to the optimal environment provided by the adherence, promoting efficient cell growth and viral vector expression. Adherent-based processes also exhibit greater control over cell behavior and phenotype, resulting in a more consistent and reliable production profile. The ability to manipulate cell attachment surfaces allows for fine-tuning of cellular processes, leading to improved vector quality and reduced production variability. Adherent cultures also offer advantages in downstream processing. Harvesting and purification of viral vectors from adherent systems are often more straightforward due to the reduced presence of cellular debris and contaminants. The cleaner harvest stream contributes to higher vector purity and facilitates subsequent purification steps. While scalability has historically been a challenge for adherent cultures, recent advancements such as innovative bioreactor designs have improved scalability. The aim of our research was to perform both a scientific and economic evaluation of both adherent and suspension viral vector production at scale to answer the age-old question: can all gene therapy indications be covered with a highly optimized adherent process, given advancements in fixed-bed bioreactors? Methods, Results & Conclusion A commonly used HEK293 suspension cell line, VPC 2.0, was compared to a clonally selected HEK293 adherent cell line, VintaCellTM. Upstream processing was assessed in both flatware and bioreactor systems. Several transfection reagents, were also evaluated to ensure fair comparison between adherent and suspension systems. Harvest material was tested for turbidity, host cell protein, and host cell DNA to compare downstrean purification burden resulting from each production approach. Results show that adherent processing offer advantages in terms of productivity (up to 20x vector produced per dollar), process control (demonstrated low variability between scientists), downstream processing (up to 30% reduction in host cell protein and host cell DNA), and potential improvements in vector quality. As technology continues to advance, the scalability challenges traditionally associated with adherent cultures are being addressed, making them increasingly attractive for viral vector production. Adherent cultures demonstrate superior cell-specific productivity. The larger surface area available for cell attachment in adherent systems fosters higher cell densities and enhanced production of viral vectors. This heightened productivity is attributed to the optimal environment provided by the adherence, promoting efficient cell growth and viral vector expression. Adherent-based processes also exhibit greater control over cell behavior and phenotype, resulting in a more consistent and reliable production profile. The ability to manipulate cell attachment surfaces allows for fine-tuning of cellular processes, leading to improved vector quality and reduced production variability. Adherent cultures also offer advantages in downstream processing. Harvesting and purification of viral vectors from adherent systems are often more straightforward due to the reduced presence of cellular debris and contaminants. The cleaner harvest stream contributes to higher vector purity and facilitates subsequent purification steps. While scalability has historically been a challenge for adherent cultures, recent advancements such as innovative bioreactor designs have improved scalability. The aim of our research was to perform both a scientific and economic evaluation of both adherent and suspension viral vector production at scale to answer the age-old question: can all gene therapy indications be covered with a highly optimized adherent process, given advancements in fixed-bed bioreactors? A commonly used HEK293 suspension cell line, VPC 2.0, was compared to a clonally selected HEK293 adherent cell line, VintaCellTM. Upstream processing was assessed in both flatware and bioreactor systems. Several transfection reagents, were also evaluated to ensure fair comparison between adherent and suspension systems. Harvest material was tested for turbidity, host cell protein, and host cell DNA to compare downstrean purification burden resulting from each production approach.