Optimizing seed train processes

Much of the focus on manufacturing challenges in the CGT space is looking at opportunities to improve processes related to cell expansion or supply chain inefficiencies. By comparison, viral vectors are often overlooked, but they remain a key component for CGTs ripe for streamlining.

Vectors are typically produced in HEK293 cells using adherent cell culture platforms. The process is well-established at this point, and as a result, most manufacturers focus on elements that seem to have a more direct impact on the therapeutic effect, like transfection and vector development. However, adherent culturing is a highly manual process, limiting its scalability. There is particular room for improvement in technology for the seed train, where the cell lines that produce viral vectors are scaled up.

Seed train processes are underdeveloped, relying on hundreds of highly inefficient roller bottles or multiple stacked culture systems to produce enough vectors for a single therapeutic dose. The inefficiency of working with ineffective flasks adds expense and waste. These outdated approaches are a starting place for further development.

Improving seed train efficiency

Roller bottles are easy to use and good for mixing, but are designed with small growth surface for cells. They are typically loaded onto incubators, where they are rotated to expose cells to both the culture medium and gases. However, their size limits scalability, and pooling becomes time intensive.

The traditional alternative is multi-stack trays, which have larger growing surfaces but are static systems with poor media and gas distribution. They also come with notable physical challenges, such as the need for a lot of incubator or facility space. They are difficult to handle, both due to the weight and the need to maneuver them in three axes to introduce and remove media.

Recent alternatives to roller bottles have the potential to improve efficiency on several fronts. These include flasks utilizing more complex internal shapes, like the Archimedes screw, which allows up to 12 times more surface area than a comparably sized roller bottle. Such devices can potentially be used as one-to-one replacements for interoperability with existing technology, but with 12 times the efficiency, reducing the need for 120-bottle racks to a single small rack of 10 screw-based flasks.

This efficiency allows for easier process intensification. Everything is more condensed, with less need for cell culture media and less trypsin required to detach the vectors following expansion. It can also enable the use of closed systems, ideal for an industry that is still largely reliant on manual handling, but which is progressing rapidly toward automation.

Future of the seed train

Soon to be available for GMP systems, these more sophisticated flasks are showing great appeal for developers that are moving toward clinical manufacturing. Developers are implementing early to avoid the need for expensive, late-stage comparability testing in the future. Early implementation is also allowing companies to begin seeing gains in efficiency earlier, positioning themselves to control long-term costs related to labor, materials and footprint.

The manufacture of these complex flasks is similarly forward-looking. They are produced via 3D printing, allowing for up to 80% less material for the same growth surface area and leading to less waste. Further, they are made from plant-based polylactic acid (PLA) bioplastic, helping labs’ carbon reduction goals. PLA is also more hydrophilic than polystyrene used in traditional single use plastic rollers, making it better suited for cell culturing applications.

As the CGT space grows, making these therapies available to more patients will require an all-of-the-above approach to improving efficiency to lower manufacturing costs. New seed train technologies can not only improve viral vector manufacturing but has applications for other types of adherent production that may be leveraged for a broad range of next-generation therapeutics, like induced pluripotent stem cells or mesenchymal stem cells.