Choosing between adherent cell culture and suspension cell culture is not always an obvious choice. Each production method can offer specific advantages to viral vector or other gene therapy manufacturing. However, as there are so many different considerations for each gene therapy product, we recommend performing a risk analysis. Read on to discover which factors to consider.
If you have a starting cell type, it can affect speed to market
New gene therapies frequently start their development journey being produced in adherent cell lines, usually for good reasons. All you really need at this stage is an incubator, and as the amounts required for early-stage development are very small, cell maintenance and production are relatively straightforward. It’s no wonder that many academic and research facilities prefer adherent cells. However, once feasibility is established and you decide to move into the next phase of the development process to produce larger quantities for analysis or preclinical material, you may consider adapting your cells to a suspension platform.
Speed to market is key when developing a new gene therapy. If you already have your cell line, sticking with the cell type you have is one way to keep pace. You don’t have to do the work to convert adherent to suspension or vice versa. Luckily, bioreactors for both adherent and suspension cells are available. Choose a system that is designed with scale up in mind, and you’ll be able to move from process development to commercial manufacturing smoothly.
But what if you don’t yet have a starting cell type? The choice between adherent and suspension cells is less straightforward, and you will need to weigh pros and cons against your goals. Licensed gene therapies are already manufactured in adherent and suspension bioreactors, so from a regulatory perspective, either can be acceptable.
Advantages of adherent cell culture
Because early-stage development usually involves adherent cells, they are often seen as the fastest route to market. Provided the lab-scale process is delivering in terms of desired yields and quality, why introduce a change that could delay getting your treatment to the patients who need it? The case for this approach is strengthened when you consider that there is proven technology that will get you there. There are also aspects of process development that can take advantage of having your cells fixed to a surface. For example, it’s easier to fully change the medium in the bioreactor, providing the opportunity to optimize growth and production phases. Also, perfusion is a more accessible option and can be employed for processes producing less stable products such as lentivirus.
Another advantage is that downstream clarification is easier with adherent cells. The presence of a fixed bed and constant recirculation of media through it can reduce turbidity and the number of particulates in the crude harvest. As a result, fewer clarification steps are needed.
Fig 1. Adherent bioreactors from Cytiva
Challenges of adherent cell culture
Yet while there may be benefits in speed to market, adherent manufacture may not be the best option for all processes. The current adherent bioreactors on the market can provide up to 600 m2 of surface area for cell growth, which still may not be sufficient to produce the titers needed for gene therapies requiring a high dose per patient, or for indications where the target population is high. The obvious solution is to scale up the process.
But scaling up comes with a variety of costs. Adherent cells require more operator intervention and processing time than suspension cells. The seed trains become more difficult and labor intensive to maintain, and they require more space in incubators because they must remain in flatware or scale bioreactors. You’ll need more labor, space, and equipment for scale up.
Another complication of adherent cells is that it is difficult to get accurate cell counts because the cells must remain attached to the surface. Furthermore, fixed bed bioreactors are a newer technology, and you may not have the in-house knowledge for using these systems.
Advantages of suspension cell culture
A suspension process has the advantage of being easier to operate than an adherent one. Adherent processes require cells to be dissociated from their growth surface for routine maintenance, scale-up, and counting. Dispensing of this step greatly simplifies the process. Once your cells are no longer dependent on a surface for growth, scale-up becomes much easier and the range of technology to support manufacture increases. Suspension cells require less hands-on maintenance, less processing time, and less space in incubators. The cells can be easily removed or added to cell culture media via pumps or syringes for sampling and seeding.
If your therapeutic can be produced in a stirred-tank bioreactor, then your options for manufacturing capacity open, as this technology is widespread at many contract development manufacturing organizations (CDMOs).
Fig 2. Suspension bioreactors from Cytiva
Challenges of suspension cell culture
Processes of up to 2000 liters in single-use systems are common, though this does introduce other potential issues such as performing transfection at large-scale. As your culture volume increases, so does your transfection mix volume. Plasmid DNA and transfection reagents must be mixed and added to the bioreactor in a consistent way in each production run. Variation at this step could lead to changes in complex formation, and this in turn could affect transfection efficiency and yields.
Because the cells are free floating in their media, media exchanges are difficult, and perfusion processes require additional equipment. Also, downstream clarification of suspension cells is more complicated than clarification of adherent cells. And if you’re harvesting adeno-associated viruses (AAV) or adeno viruses (AV), you need to perform cell lysis, which results in a mix of virus, hcDNA, host cell proteins, and other impurities.
Comparable production costs
These points lead to the cost per dose of each production method. A comparison of installed capital, labor, and footprint across biomanufacturing platforms shows that, while scale up from a multi-tray platform is time-consuming and labor-intensive, both adherent and suspension single-use bioreactors have lower labor costs and require less space. In fact, the costs associated with adherent and suspension single-use bioreactors are comparable.
Conclusion
To summarize, each production method can offer specific advantages to gene therapy manufacturing. However, as there are so many different considerations for each gene therapy product, we recommend performing a risk analysis considering some of the factors discussed in this blog. Going through this process will help define your development plan and contingencies if you don’t see the results you desire.
Whether you choose adherent or suspension cells, Cytiva has you covered. Our family of iCELLIS™ bioreactors, is designed for adherent cells, and our Xcellerex™ bioreactors are used for suspension cells.