With an increasing number of approved and phase 3 clinical trial gene therapies using adeno-associated virus (AAV) as vectors, facilities manufacturing these vectors can barely keep up with the current demand.
Scaling the viral vector production is inevitable to increase the capacity focusing on higher yields and lower costs of goods. Even more important is safe handling of the liquids from small to large volumes to safeguard each production step, avoid batch loss and reliably stick to manufacturing plans. Not all GMP vector manufacturing organizations are prepared yet to handle especially large volumes of liquids, as this poses big challenges. The good news: a seamless transition to commercial AAV manufacturing is possible.
What are AAVs currently used for?
AAVs are currently one of the most frequently used vectors for gene therapy. (1) For both in vivo and ex vivo gene therapy, viral vectors are being used to transport genetic material into cells for gene correction. With the help of viral vectors gene therapies can target cells in order to potentially cure different genetic disorders and a wide range of diseases. Among different viral vectors, AAV has proven to be one of the most suitable vehicles to target cells for this purpose. It outshines other vectors due to its safety, its persistence of the virus, the lack of pathogenicity, its low immunogenic disposition, but also its ability to infect cells that are nondividing and dividing. (1)
The many advantages have made AAV an increasingly common vector for human clinical trials. And undoubtedly the demand will continue to grow.
Challenges in AAV manufacturing
The increased demand of AAVs drives GMP manufacturing of vectors to increase batch sizes. Upscaling of AAVs is a major challenge for most companies, especially the tech transfer from laboratory to commercial production.
Production of larger volumes poses new challenges compared to small- or lab-scale production. In development and early clinical stages, R&D scientists perform mixing or transfection of liquids below 1L within a few seconds and mainly manually. The same applies to liquid transfer and storage. Scaling-up to 1000L batch sizes and more, however, adds more complexities and hence the need for standardization. Safe liquid handling during filling, cold chain logistics and shipment require a GMP compliant standardized process in order to ensure reproducibility and control. Lack of control threatens the effectiveness of the product at the end and increases batch-to-batch variability. (2)
AAVs also bear challenges, e.g., preexisting or generation of antibodies in the patient’s organism, harsh and potentially life-threatening side effects when increased doses are needed or non-permanent expression documented in some cases. A promising solution to overcome these hurdles is the usage of engineered commensal viruses, e.g., from the large group of anelloviruses.
No matter which type of viral vector will be used to develop gene therapies and will hopefully bring cures to many patients, increasing number of batches and large volumes of viral vectors for clinical trials and gene therapy will be needed with increasing numbers of gene therapies addressing so far non-treatable and often lethal medical conditions. Supplying large volumes of AAVs and other viral vectors entails a controlled and scalable process to manifest growth.
Ensuring safe handling of AAVs for large volumes
Gradual expansion of volumes from 1L up to 2000L cannot be coped with at the initial lab facilities. Transition from small volume production to commercial production requires numerous different platform systems and facilities. This includes aseptic filling and freezing for long-term storage to -80°C.
Smart platform solutions cover applicability throughout different scales whilst maintaining the same automation software. Adhering to one continuous cGMP control software throughout all scales facilitates consistent, comparable and uniform deployment into each manufacturing execution system (MES).
Modular and automated filling solutions and cryogenic applications from Single Use Support respond to the challenge of securing safe handling for different scenarios and volumes: The control unit for GMP-compliant dispensing into single-use bags and bottles remains. Different filling racks can be attached that differ in number and size of single-use bags. Even hybrid filling of both primary packaging, single-use bioprocess containers and bottles, is possible. Plug & play modularity enables controlled fluid management for different scenarios in filling process. Implementing automation for aliquoting large volumes into single-use bioprocess containers contributes to accuracy, quality and cost-efficiency. It is the entry ticket to reproducible cGMP vector manufacturing.
Freezing AAVs to -80°C is advised for shipment and long-term storage. The adeno-associated virus is a vector with minimal loss of activity during freezing, however, due to its large protein surface it is more likely to aggregate. Fast and controlled freezing counteracts the phenomenon of cryoconcentration in order to facilitate homogeneous and reproducible freezing results that reduces protein activity to a minimum. Single Use Support’s freeze and thaw platform solutions come with different sizes. Their unified method of freezing and thawing through plates and its CFR part 11 compliant control software does not only secure automation and therefore standardized cGMP for freezing. Its recipes are transferable to secure identical freezing curves throughout different-sized platform options.
Digital Transformation to foster growth of AAV manufacturing
Automation and the interconnectedness of platform solutions through digitalization are the way forward to establish safe handling throughout AAV manufacturing. Averting from siloed platforms, it is the holistic connection of process steps that facilitates dealing with liquid transfer at different scales. With standardization in cold chain logistics comes control of safe handling – even for large volumes.
(1) Gene Therapy Using Adeno-Associated Virus Vectors - PMC (nih.gov)