Producing MSC-derived cell therapies: workflows, technologies, and case studies

The following content was originally published by Cell & Gene Therapy Insights on March 28, 2025

Irana Coletti Malaspina, Shirley Mei, Tony Ting, and Whitney Cary Wilson

SM: MSCs became prominent because of their ability to differentiate into bones, cartilage, and fat, as well as their use in various regenerative purposes. For example, there is a lot of potential in orthopedics and other fields. Over time, people have realized that MSCs can not only regenerate tissue but also modulate the host immune system. When MSCs are used in different disease environments, such as sepsis or acute respiratory distress syndrome (ARDS), they can interact with the immune cells and secrete molecules such as microRNAs, exosomes, and proteins, which can influence the immune system’s response.

For instance, in sepsis, it is not just that patients have hyperinflammation—many also develop immune suppression. This means that if you only give a drug to suppress inflammation, it will not be beneficial for patients who later enter the immune-suppressive stage. Fortunately, MSCs can adapt to the conditions and influence the body either to boost or dampen the immune system.

The key factor that attracts developers, academics, as well as big companies in the industry to MSCs is their ability to be used in an allogeneic manner, meaning it is not necessary to match the donor and recipient. This opens the potential for MSC-based cell therapies to be developed into commercially viable products. For example, you can isolate cells from a single donor or a pool of donors and produce large quantities of MSCs. These cells can then be packaged in various doses and given to unrelated recipients, potentially reducing the cost of goods (COGs).

TT: Given their properties and strong safety record, there have over a thousand studies using MSCs, covering many different indications. As mentioned earlier, orthopedics was one of the initial areas where MSCs were explored. Graft-versus-host disease (GVHD) is another area, and we have recently witnessed the first approval of an MSC product in the US for the treatment of pediatric steroid-resistant GVHD. Additionally, MSCs are being used for various respiratory indications, such as COVID-19-related ARDS. MSCs are also being studied for neurological conditions, with research in stroke and other CNS disorders. Autoimmune diseases, such as multiple sclerosis, are yet another area that has been explored.

SM: As mentioned earlier, MSCs can be used for sepsis and septic shock, which currently do not have an effective treatment—we have been developing a modified MSC product aimed at solving these issues. ICM There are many neurodegenerative applications, such as Huntington’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis.

WCW: There are also many topical applications for MSCs, such as for conditions like rosacea and basic wound healing. There is also a very interesting study going on at UC Davis, using an MSC-based patch to help infants with the spina bifida condition [1]. The surgery is performed in utero by applying the patch to the fetus. 

ICM: From my perspective, the major challenge in using MSCs as a medicinal product is establishing a standard protocol. As the production is scaled up and the developers move from small batches in academic settings to larger batches, it is crucial to be cautious with the isolation step. The isolation process can introduce contamination from other cell types or even microbiological contaminants, which can depend on the tissue source.

Another challenge is related to phenotype and potency, particularly with genetic markers, which makes it difficult to establish a standard quality control. In the beginning, we may have many MSCs, but we need to select a small number of batches to move forward into production. Additionally, MSCs can undergo senescence during extended culture periods, leading to more challenges.

We also need to consider good proliferation rates when working with MSCs. It is essential to limit the number of passages the cells undergo during cultivation and be mindful of their phenotype and differentiation potency. Strong quality control and quality processes throughout production are crucial. Scalability is another significant challenge. Therefore, when working with MSCs, we must carefully plan how to scale up the process and ensure that quality control is in place to support production.

SM: The tissue source and regulatory compliance are also crucial considerations. For example, if the cells are isolated from bone marrow, adipose tissue, or cord blood, you not only have to go through an ethics board but also ensure proper donor screening. Furthermore, the tissue source must be tested, and once it is made into a master cell bank, that also needs to be tested.

Different regions have varying regulatory requirements that must be satisfied. As an academic, you might primarily focus on isolating cells and scaling them up for a clinical trial. However, if you are in a company, you also need to consider different types of regulatory requirements, especially if you plan to market or sell the product across different jurisdictions.

From a more scientific point of view, potency is another critical factor. The type of potency assay you develop must be indication-specific. Many researchers use a T cell inhibition assay because MSCs naturally inhibit T cell proliferation, making this approach commonly used for conditions like GVHD. However, for diseases such as sepsis, the potency assay must be carefully designed. During product manufacturing and process development, it is essential to ensure that the cells remain potent as they are scaled up, and regulatory agencies increasingly require this data. In the beginning, they may accept early-stage assay data, but later on, these assays could become part of the release specifications. Therefore, any MSC developer must carefully consider potency assays when developing a scale-up strategy to produce clinical-grade cells.

TT: It is also important to understand the mechanism of action and ensure that you have control over the manufacturing process. Developing a robust potency assay is critical, and it should be one that can be used routinely. Another important point to mention is the media. In the early days of MSCs, fetal bovine serum (FBS) was commonly used, but it is not an ideal ingredient for MSC manufacturing. Most commercial companies have now moved to xeno-free media formulations, either completely chemically defined or containing human platelet lysate. However, even with human platelet lysate, there are concerns about batch-to-batch variability, which requires careful management.

SM: In my laboratory, we work on different ways to scale up the production of these cells for clinical use, which is very different from working at the research laboratory level. For instance, when working with mice, you only need around 1 million MSCs per animal. But when thinking about using MSCs to treat patients, the required dose can range from 30 million to 1 billion cells per dose. How do you scale up to produce so many MSCs while also complying with clinical-grade and regulatory requirements?

We also work on optimizing expansion protocols by collaborating with vendors and exploring strategies such as microcarriers and bioreactors. The tricky aspect with MSCs is that they are adherent cells, meaning they need a surface to grow on. MSCs need something to adhere to, which is why microcarriers are used. However, there are many different types of microcarriers, and some are good for MSC growth but may also prevent the cells from coming off, which is another challenge we actively work on solving.

Additionally, there are many media choices available, and each of them can grow MSCs differently in 3D or bioreactor-based cultures. We also carry out 2D expansion because it has a lower barrier to technology transfer. Moving a protocol from an R&D product development setting to a CDMO, which will manufacture GMP-grade MSCs, provides more surface area for the cells to grow, but it also introduces new challenges. Typically, it is required to have a larger facility, incubators, and more people to manage the cell cultures and harvest the cells. You also need a large amount of media, which will impact the COGs. For early trials or cell therapies that do not require large doses, 2D expansion is usually easier. However, after scaling up the cells, we must also consider whether they can still maintain their potency. Genetic stability, which I mentioned earlier, is also very important.

TT: One of the biggest challenges is the variability of the starting material. As we discussed, MSCs can be isolated from a variety of tissue sources. While there have been over 1,000 clinical studies using MSCs, very few products have been approved. One of the considerations is that while MSCs are effective, they may not always be potent enough. This has led to various techniques aimed at enhancing the potency of the cells. This can include preconditioning MSCs with different cocktails of cytokines. There are also people working on genetically engineered MSCs to enhance their properties. One of the more interesting technologies I have seen is the development of iPSC-derived MSCs. These offer several advantages, such as a consistent donor source (the original iPSC line), as opposed to standard MSCs, which require multiple donors. Overall, I think these approaches will help overcome some of the challenges.

Furthermore, it is important to think about the commercialization plan. If your program is successful, how many cells will you need for your product? The sooner you think about a scalable manufacturing process, the easier it will be down the manufacturing path. I would even suggest that, before starting a clinical trial, if you can move to a bioreactor system, you will be set for the long term. It is not the cheapest approach, but if you have the resources and capabilities, it will make further development much easier.

WCW: It is crucial to identify the critical quality attributes of the MSC therapeutic early in the process, as well as develop potency assays. Developers must also account for how they will scale the cells and develop potency assays that work at scale. The earlier one can start thinking about these factors, the better off they will be in the long run.

ICM: From my perspective, it is not just the MSC scale-up we need to be preoccupied with. We also must figure out how to produce the large amounts of media required for large-scale production. We must also be mindful of the downstream process, ensuring it is gentle on the cells, as well as the fill-and-finish steps. With MSCs, there are a lot of steps in the process that we need to handle carefully.

WCW: There are many ways to expand MSCs. As mentioned earlier, MSCs are adherent cells, meaning they prefer to attach to a substrate. Some of the earlier technologies, which are still widely used, include stackable cell culture vessels. There have been improvements in closing these systems, such as adding closed system caps with tubing for the fill and harvest process steps. There are also hollow fiber bioreactors that have been used for expanding MSCs, as well as some fixed-bed bioreactors on the market, which are part of the early adoption of MSC scale-up. Additionally, as already mentioned, a lot of work has been done with microcarriers in what we would call a pseudo-suspension, which allows for much more efficient use of media and reagents when scaling up.

TT: The technologies for cell culture and bioreactors are improving. In particular, more companies are developing technologies that allow us to monitor the process in real time. For example, now we can measure lactate and glucose in real time, which was impossible 10 years ago. It is encouraging to see that companies are developing these tools for large-scale manufacturing of MSCs.

WCW: To emphasize again, it is crucial to ensure that the media and reagents are all GMP-compliant. Historically, MSCs have been grown with FBS. However, FBS is not the optimal product for manufacturing, as there is significant lot-to-lot inconsistency with it. Therefore, using more defined media and products can help reduce batch-to-batch variability as you scale up manufacturing, which is very important.

SM: FBS also poses regulatory concerns: for example, if the MSCs grown in these media are given to the patient, you will have to justify the source of the serum. Some sources of FBS cannot even be used because regulatory agencies will reject them due to safety concerns, such as those related to mad cow disease. For this reason, many media companies are developing serum-free or chemically defined media.

From a developer’s point of view, you do have to test these media because MSCs are isolated from different tissue sources and have different isolation protocols. They might not work well in all media types, so you must test which one suits your needs. Additionally, you also want to work with a reliable vendor that has experience producing a certificate of analysis and can justify the sourcing of the raw materials used to produce the media. Working with good vendors who can help ensure the media are GMP-grade, or at least closer to GMP-grade for early-phase trials, is crucial. They should have a track record and specifications that can address regulatory concerns. These are important considerations when moving into clinical applications because you will need to address each of them in your CMC documents when submitting them to regulatory agencies.

Another factor to consider is cost. Some of these media and reagents can be quite expensive, which will factor into the COGs, so you will need to address this and find ways to minimize it for successful commercialization. This is especially important if you aim to treat a widespread disease, like COVID-19, where many people would need access to the treatment.

TT: As you move toward commercialization, it is crucial to ensure that you have multiple vendors for each key ingredient in the media, if possible. It is also important to think about how the product will be stored, whether in a bag or a vial.

ICM: During MSC production, we do not only have the MSCs themselves but also the media for culture and preservation. We must be careful about residual products in the final product. Using GMP-compliant, specialized products will be better in terms of residual considerations. This is important for ensuring the safety of the product during clinical trials.

TT: Given all the different indications that have been explored with MSCs, one can imagine there have been many different delivery processes. In essence, it is about understanding the biology of the MSCs in relation to the specific disease indication. For example, in a variety of CNS indications, researchers have tried direct injection into the brain, intrathecal delivery, or intraspinal delivery. However, intravenous delivery is most commonly used.

Regarding dosage, it is a huge challenge in the MSC space. Most studies have been done on small animals, such as mice or rats, and it is very difficult to scale dosing to humans. I was fortunate enough to conduct cardiac studies in pigs, whose hearts are roughly the same size as a human heart, which made it much easier to work on the dosing strategy. However, if you are working with mice or rats and then transitioning to humans, your first human clinical studies must evaluate a range of doses to establish safety and determine the optimal dose for efficacy.

ICM: Since MSCs are a treatment, we need to determine the optimal dose for each patient. At the beginning of your clinical trials, it is important to consider various doses to find the best one for treating the specific disease.

SM: Cell therapy developers must communicate with the people who will eventually deliver the therapy because it will affect how the final cell product is packaged and delivered. It is not just about injecting the treatment into the patient. In fact, there are many questions to address: Is it in a bag? Is it in a vial that needs to be washed or diluted? Will it be gravity-fed, or will it go through an infusion pump?

Due to these complexities, there has been a lot of movement away from using fresh cells, as was common in the past. If you are thinking about commercialization, treating more patients, and addressing urgent diseases such as sepsis, ARDS, or COVID-19, you typically do not have time to grow, process, cryopreserve, and thaw the cells for weeks. Instead, you need a standardized protocol that allows for readiness.

In the hospital setting, there are different clinicians or coordinators you can work with, thereby the protocol must be adaptable to most of them. When running a clinical trial, you do not want issues such as non-compliance or loss of cell viability or potency. These factors could affect trial outcomes and, ultimately, the product’s progression to the next stage. We have encountered this challenge both in academic and industry trials, and it is important to approach the problem from different angles and work closely with the clinical teams.

WCW: Going back a little bit to the biology of the MSC itself and its mechanism of action in eliciting a therapeutic effect, when we consider how to deliver the MSC, we also need to understand whether we are aiming for a transient effect or a longer-term therapeutic effect.

For example, if you have engineered the MSC to secrete a growth factor, such as brain-derived neurotrophic factor for Huntington’s disease, you might want the MSC to survive longer in the location where you are injecting it. In this case, you could think of the MSC as a biofactory. On the other hand, it is different if you are using the cell to provide a transient effect, such as an immunomodulatory effect. This is an important aspect to consider when deciding between an IV injection or a direct injection.

Additionally, it is crucial to consider scaffolds. If you are seeding the MSC onto a scaffold and want the cell to persist for a while, but not necessarily proliferate, you need to choose a scaffold that supports the cell and allows it to receive nutrients from the body.

Lastly, another consideration is the environment into which you are injecting the cells. Will they be exposed to a hypoxic environment, such as the brain, or will they have access to nutrients and oxygen?

Fundamentally, it all comes back to understanding the mechanism of action and how you expect the MSC to elicit the therapeutic effect.

ICM: MSC products are fresh, meaning we must be careful not only in the fill/ finish model but in all the steps—both upstream and downstream—to ensure a sterile process. We must maintain the sterility of the entire process, and the fill/finish is the last step. The best option, of course, is utilizing closed systems, but there are currently limited machines available that can perform the fill/finish in these systems. Time is also critical in these steps because we are using dimethylsulfoxide, a cryoprotectant, to preserve the cells.

Consistency between different bags or vials, especially regarding the volume concentrate, is another important consideration during fill/finish. For this, a proper formulation for cryopreservation media, as well as a well-defined standard protocol and standard operating procedure for this step, is crucial. After fill/finish, cryopreservation is the next critical step because the cells must be cryopreserved in nitrogen. Ice formation during this process is one of the most challenging aspects. All things considered, from my perspective, the fill/finish process is the major challenge in the final production of MSCs.

For storage, we need the MSCs to be a long-term product, so we must ensure the stability of the process. Stability studies are essential to ensure consistency in cell viability and recovery after cryopreservation. It is important to be mindful of the cryopreservation solution and the temperature ratio, as these factors are critical during the fill/finish steps.

WCW: Another significant consideration in the final fill/finish is handling volume. Depending on the vessel used for scaling the product, the physical volume that comes out of the technology will vary. It is crucial to consider this aspect, especially when planning for volume reduction while maintaining an aseptic environment and performing the process in a closed system.

SM: It is crucial to establish a stability program when developing therapies because regulators will ask about it. The specifications and functional, identity, and other (FIO) assays to evaluate the stability of your product are crucial. In the laboratory, you might think about using a complicated co-culture assay to measure various factors, but it can be very difficult and expensive to tech transfer that assay to a CDMO or CRO.

The key is developing assays that provide quantitative measurements that can give outputs equivalent to the potency or specifications of the product. You want to satisfy the regulatory requirements, but you also want to ensure that the assay is highly reproducible and standardized. This assay will likely be required not only for your product release but also for the stability program.

TT: It is also important to retain enough product so that, if additional testing is needed, you have enough on hand. I would recommend keeping more than you think you need.

TT: I am thrilled about the clinical studies that are currently ongoing with iPSC-derived MSCs. I think it is going to be a very exciting space to watch. Gene-engineered MSCs will also play a crucial role in improving therapeutic potency.

WCW: I am very excited to see these therapies becoming more available. From an accessibility perspective, it is important to drive down the cost of manufacturing. I see a lot of efforts in that space, aiming to make cell manufacturing more affordable. It is truly amazing to develop these different therapeutics and see the biology behind them, but if we can only administer them to a small group of people who can afford them, that is not a good solution for humanity.

ICM: I am also very excited about the potential of having a product based on exosomes. Often, exosomes were considered a residual byproduct, but now we can use both MSC products and exosomes from a single production process. The key point about using MSCs and exosomes is the possibility of treating diseases that currently have no treatments available, giving hope to thousands of patients.

SM: With the approval of the first MSC therapy in the USA, there is a lot to be excited about. When I first started in the MSC field 20 years ago, there were barely any MSC-specific media. Now, there is a repertoire of options for various purposes, and many companies are willing to invest in different systems to scale up MSC production and support clinical translation. The monoclonal bioreactor developments will help MSC technologies address challenges and further our understanding.

1. Farmer DL. Cellular therapy for in utero repair of myelomeningocele—the CuRe trial (CuRe). Oct 1, 2024; ClinicalTrials.gov. https://clinicaltrials.gov/study/NCT04652908.

Irana Coletti Malaspina is currently a Field Application Scientist—Latin America for Corning Life Sciences. She has more than 9 years of experience in end-to-end MSC manufacturing for advanced therapy medicinal products.

Irana Coletti Malaspina, Field Application Scientist—Latin America, Corning Life Sciences, São Paulo, Brazil

Shirley Mei has over 20 years of experience in both industry and academia. She is currently a Scientific Investigator in the Regenerative Medicine Program at the Ottawa Hospital Research Institute, where her lab develops and optimizes MSC expansion protocols and investigates how MSCs interact with immune cells.

Shirley Mei, Scientific Investigator, Regenerative Medicine Program, Ottawa Hospital Research Institute, Ottowa, ON, CAN

Tony Ting is the CSO for Kiji Therapeutics, which is developing off-the-shelf engineered cell therapies. He has over 30 years of academic and industry experience in translational science and global regulatory filing including more than 20 years in the cell therapy field, at companies including Takeda, Bone Therapeutics, and Athersys.

Tony Ting, CSO, Kiji Therapeutics, Cleveland, OH, USA

Whitney Cary Wilson is a Field Application Scientist at Corning Life Sciences. Previously, Whitney spent 12 years at the UC Davis Institute for Regenerative Cures and was Director of the UC Davis Stem Cell Core. Now, she works with process development groups to optimize production capabilities and cellular scale-up conditions.

Whitney Cary Wilson, Field Application Scientist, Corning Life Sciences, Sacramento, CA, USA

Contributions: The named authors take responsibility for the integrity of the work as a whole, and have given their approval for this version to be published.

Acknowledgements: None.

Disclosure and potential conflicts of interest: The authors have no conflicts of interest.

Funding declaration: Tony Ting has received consulting fees from Healios KK and Steminent BioTherapeutics.

Copyright: Published by Cell & Gene Therapy Insights under Creative Commons License Deed CC BY NC ND 4.0 which allows anyone to copy, distribute, and transmit the article provided it is properly attributed in the manner specified below. No commercial use without permission.

Attribution: Copyright © 2025 Corning Incorporated. Published by Cell & Gene Therapy Insights under Creative Commons License Deed CC BY NC ND 4.0.

Article source: This article was based on an expert roundtable.

Expert Roundtable recorded: Feb 13, 2025.

Revised manuscript received: Mar 19, 2025.

Publication date: Mar 28, 2025.