Repligen Corp.

Rachel Legmann

Platform approach for intensify virus-based therapeutics process manufacturing

Expanding the disease indications of gene therapies beyond rare diseases as well as developing more complex but also more fragile target viral vectors require not only expanding existing manufacturing facilities but also developing new capacities. The manufacturing processes that enabled the first landmark gene therapies to successfully complete clinical trials and regulatory approvals lack the efficiency and productivity required to meet current and future demand. Special requirements on the viral vector manufacturing processes, such as low cell density, low production and instability create challenges in their scale-up that cannot be solve by traditional platform approaches. Advanced technologies are needed to meet the vector demand and provide the required reliability and robustness for manufacturing enabling gene therapies to meet their full potential.

Challenges across the vector process workflow

A key challenge in the field of therapeutic viral vector manufacturing is maximizing vector yield during the entire process at all scales. Current upstream viral vector manufacturing processes primarily are based on transient expression, using multiple plasmid DNA, typically suffer from low productivity and a complexity that can challenge manufacturing reproducibility. Current FDA approved Adenoassociated Virus (AAV)-based gene therapies are facing scalability challenge since they are mainly generated by adherent cell culture using either flatware’s or fixed-bed bioreactors.

Operation with non-fit platform during purification leads to low recovery yield of viral vectors. Translation of filtration and chromatography platforms used for biological drug processes to viral vector processes does in fact lead to product loss and low recovery yield due to hold up volume, unfit pump, aggregation, or shear stress. Affinity resins efficiently remove host cell proteins (HCPs) and DNA impurities from AAV but are challenged by the diversity of AAV serotypes.

Because the product itself is a virus or viral vector, therapeutic protein-based contaminant (bacteria or adventitious viruses) removal techniques such low pH viral inactivation and sterile/ virus filtrations are not compatible. This drives a strong need for contamination prevention strategy over removal. Furthermore, analytical technologies for in-line monitoring of the product critical quality attributes (CQAs) are limited for gene therapy. Most gene therapy analytical methods are conducted off-line with high turnaround time eliminating the ability to make smart decisions during the process development, leading to more development cycles and therefore the cost and slow time to market.

Traditional upstream, downstream, and analytic platforms are not able to overcome those challenges.

Dare to Innovate – Overcome the main manufacturing challenges

One must integrate an advanced manufacturing platform approach to significantly enhance viral vector overall yield leading to AAV and lentivirus (LV) process cost reduction and therefore, affordable virus based therapeutic drug. To meet the high vector demand, the market is moving into suspension cells or producer cell line, even though their current specific productivity is lower, the process can be scalable. To enhance the vector yield for the suspension cells it is critical to integrate a better fit perfusion system into the vector production bioreactor enabling both higher growth and continuous harvest during production. The perfusion system, such as the tangential flow depth filtration (TFDF) is a great fit to vector production since it does both. This perfusion system currently benefits mostly envelope viruses that are secreted outside the cells like lentivirus, retrovirus, VSV and many current AAV serotypes that are secreted outside the cells.

Higher recovery yield of the functional vector and process performance consistency, at all manufacturing scales and during each of the downstream step, can be achieved by using systems that were designed by having the needs for advanced therapy medicinal products (ATMP) in mind. The main design features of concentration and purification systems should have over-molded tubing connections, compact XO valve designs, combining filter and bubble trap, and advanced gradient control enabling consistent and better separation of AAV during the polishing step functions primarily to separate capsid lacking DNA (empty) from capsid containing DNA (full). Systems that have those features such as RS TFF system and KRM chromatography system enable increased process efficiency and overall process step yield, protect potency and product integrity, reduce the overall risk of deviations through design, and enhance user experience.

Another major challenge in the vector production process is the lack of on-line rapid and accurate analytical tools. The implementation of the FlowVPX System technology with variable pathlength technology (VPT) can offer a quick and direct total viral vector analysis during development to enhance throughput and improve decision-making.

Willing to build an agnostic single use and closed automated platform by adopting advance technologies that are better fit to large fragile viruses, should enhance vector yield through process efficiency as well as protecting vector potency and integrity, reduce deviation risks through process control and meet vector demand at affordable cost.