Raw Materials for Advanced Therapies: When the Process Is the Product, Ingredients Are Key

Photo courtesy of SAFC and MilliporeSigma (EMD Millipore outside the United States)

Scott Burger and Bill Janssen are both established, independent consultants specializing in gene and cell therapies. This past spring, we three discussed several aspects of raw material strategy for advanced therapies as well as the need for trained technicians in the industry.

With a bachelor of science degree in biology from Tulane University (New Orleans, LA, 1983) and a medical doctorate from the University of Pennsylvania (Philadelphia, PA, 1988), Scott Burger served his residency training and fellowship at Washington University (St. Louis, MO) from 1989 to 1994. From there, he joined the University of Minnesota (St. Paul, MN), where he was an assistant professor of pathology and laboratory medicine from 1994–2001. There, he directed a good manufacturing practice (GMP) laboratory making cell and gene therapy products for clinical trials and transplantation. He also provided regulatory assistance to other University of Minnesota investigators and collaborators.

After a year as vice president of research and development for Merix Bioscience, Burger stepped out on his own as founder and principal consultant for Advanced Cell & Gene Therapy in 2002 (Chapel Hill, NC). He consults on development and commercialization of cell/gene therapies and tissue-engineered products, focusing on due diligence and strategic partnering, GMP and good tissue practice (GTP) manufacturing, process development, characterization and comparability, and regulatory affairs. He also has served on the International Society for Cell and Gene Therapy’s (ISCT’s) global regulatory perspectives workshop steering committee since 2008.

Bill Janssen earned his bachelor of arts degree in biophysics from the University of California in Berkeley, CA (1977), then a master of science in microbiology and immunology from the University of Colorado Health Sciences Center in Denver, CO (1979). After earning his doctorate in biophysics from the State University of New York in Buffalo, NY (1984), he served a postdoctoral fellowship at the University of Florida College of Medicine in Gainesville, FL (1984–1989). There, he researched a methodology for purging neoplastic cells from bone marrow as preparation for autologous transplantation.

For over a decade, Janssen was a senior member and director of the cell therapy facility at Moffitt Cancer Center in Tampa, FL (1989–2014). He founded the blood- and bone-marrow processing laboratory there, developing it from a 600-ft2 space into an 8,000-ft² GMP-compliant cell therapy manufacturing facility employing two dozen technologists and supporting nearly 500 transplants and more than 100 immunotherapy products every year.

Next, as director of the human applications laboratory at St. Jude Children’s Research Hospital in Memphis, TN (2014–2019), Janssen oversaw all cellular processing activities (except for genetic modifications). Based on evidence-based best practices, he instituted a quality management system at the laboratory as it operated in compliance with GTP, GMP, and standards set by the Foundation for the Accreditation of Cellular Therapies.

As a consultant with William E. Janssen Cell and Gene Therapy Consulting Services since 2019, he assists academic and industry organizations with regulatory applications, facility layout and operations, bench-to-bedside translation, and informatics implementation for cell and gene therapy manufacturing operations.

Cells, Media, and Supplements
Gazaille: What starting materials attract the most attention in cell and gene therapy production, and why?

Janssen: Raw materials for cell therapy have advanced a lot in the past 10 years, especially from a regulatory standpoint. Some materials seem to garner more consideration than others, but the “elephant in the room” is the cells themselves. They probably should garner the most consideration, but they don’t.

On the side of purchased raw materials, the cell and gene therapy (CGT) industry has progressed from nothing being approved by the US Food and Drug Administration (FDA) and many questions over what we should or shouldn’t use and what qualifications to apply — to now having many options competing with each other. As a case in point, several manufacturers make culture media for expansion of T cells. (Editor’s Note: Examples include ExCellerate human T-cell expansion media from R&D Systems, ImmunoCult-XF T-cell expansion media from StemCell Technologies, and CTS OpTmizer T-cell serum-free media from Thermo Fisher Scientific.) And depending on which published paper you read, any one of those might prove to be the best, and any one of them might prove to be the worst. The biggest problem is that their formulas are proprietary. You can’t just buy the one with the highest concentration of methionine, alanine, and asparaginase, for example, if you know that works well with your cells. So the challenge comes in deciding which option will be best for your process without needing too much of the other raw material — the cells, which are in short supply — to test them all.

Burger: The number and quality of raw-material alternatives have improved as GMP-compliant formulations became critical raw materials. But the understandable tendency of vendors to rely on their drug master files rather than disclose the composition of these raw materials definitely makes things more challenging. There are benefits and drawbacks with the current situation.

Another change is the increasing availability of raw materials that are animal-origin free. However, there isn’t a standard definition of animal-origin free, or of primary, secondary, or tertiary levels of it. There’s a natural tendency to say, “Well, if it’s recombinant or plant-derived, then it must be animal-origin free.” But the details sometimes aren’t so clear as that. If a product is derived from plants, but those plants are grown out on a field where animals can interact with them, then the plants from which the raw material was purified might have been in direct contact with animals. That introduces complications.

Gazaille: Why should cells get more consideration as raw materials than they do now?

Janssen: The cells come from donors. Even with autologous products, with patients both donating and receiving the cells, there are potential complications. We can establish very tight qualification standards for the buffers or labware that we use (and even for those media with their proprietary formulations), and that gives us some assurance that every lot of such things will be alike and provide the same results.
Apheresis procedures give us what is probably the most common raw material for cell therapy. You can use the same program on the same instrument, and the same kind of components, with the same apheresis operator running the machine on the same donor five times on five different occasions — and you will end up with five different results. The cellular composition will be a function of the time of day (we all have circadian rhythms), what the donor ate recently, whether the person is developing a head cold, and so on. Expand out to different donors of different sexes and ages, with different lifestyles, and all those things will play a role.

What is arguably the most critical starting material for every cell-therapy process will be highly variable. Regulatory authorities want to treat cell-based therapeutics the same way they treat traditional pharmaceuticals: They want to see consistent end products. But it’s very difficult to produce that when you’re starting with highly inconsistent raw materials.

Burger: In addition to variability, the cellular starting material is inherently heterogeneous. Even the purest population of a given type of cells will contain subpopulations of cells, and we don’t know what all of those are. Autologous cell therapies add another level of complexity in that the cells can be affected by a patient’s previous treatments, such as chemotherapies or steroids. Such biological sources of variability and heterogeneity make process control all the more important. It is essential to reduce process-related variability, including cell collection.

Gazaille: Stem cells tend to become even more disparate over several passages of expansion. Wouldn’t that make process control even more difficult?

Janssen: True. There are different processes in the course of manufacturing, some of which will reduce the amount of variability. Many processes involve some kind of a cell selection using an instrument such as Miltenyi Biotech’s CliniMACS system, with cells either rejected or selected specifically based on surface markers and specific antibodies that bind to them as antigens. That step will reduce variability. But then culturing stem cells accentuates the differences.

Gazaille: How has the COVID-19 pandemic intensified sourcing concerns?

Janssen: I’m certain that it has done so — the principal challenge being just getting people to go to a collection center. Also, it may be difficult to maintain social distance within an apheresis facility. Scott, are you aware of any problems with donors having been exposed to SARS-CoV-2 or having disease effects, more than any other kind of viral infection would affect the product?

Burger: I haven’t seen data for SARS-CoV-2 specifically. Donor screening questions would exclude people with active symptoms of COVID-19 or known recent exposure, though.

Gazaille: How fresh must cellular materials be (or how well do they perform after cryopreservation)?

Burger: Cryopreserved leukapheresis products are used in many cell therapy manufacturing processes and perform well, if frozen and thawed properly. The problem is that manufacturing processes aren’t usually developed and validated to use fresh or cryopreserved cells interchangeably.

Janssen: Seasonal influenza changes donor patterns, of course. If somebody recently had a head cold, that would change the apheresis product from that person just because different cell subsets have been expanded as part of the immune system response. You would expect that with any kind of viral or bacterial infection.

Gazaille: How applicable are quality by design (QbD) tenets to addressing the wide variabilities in cell therapy starting material?

Burger: QbD is a useful approach for much cell therapy product development, but it doesn’t work for all aspects. It can’t be applied in the way it is used for small molecules or nonliving biologics such as peptides or monoclonal antibodies.

Janssen: The six-sigma approach also falls short in this context. Its focus is on reducing variability in supply streams, manufacturing methods, and so on. When your starting point has that much inherent variability, the rest of it just can’t hold together.

Gazaille: We’ve already talked a little bit about culture media and the cells themselves. Are there other raw materials that need more or less consideration?

Burger: Fetal calf serum is commonly used as a culture medium supplement in research laboratories. But it’s usually one of the first things we want to eliminate or at least minimize in cell therapy manufacturing. Because of its origin, serum is inherently variable and not fully defined. Sourcing it is complex and introduces infectious-disease considerations. If we can’t eliminate it during process development, it needs to be sourced and controlled rigorously.

Cytokines are essential for manufacturing many cell therapies and present challenges of their own. Many cytokines mediate multiple functions in different cell types. The function measured with a standard assay for a particular cytokine may not be the function that it mediates in your manufacturing process. That can make changing suppliers difficult because even if cytokines from two suppliers have the same name and behave the same way in the standard assay, they won’t necessarily work the same way in your manufacturing process.

There are not many suppliers of GMP-grade cytokines, and some cytokines are available from only one or two companies. Although growing numbers of cytokine manufacturers are stepping up to the challenge of offering GMP formulations, some cytokines still are available as research-use only (RUO). In those cases it’s necessary to “qualify up” the RUO formulation by testing, and that can be an expensive proposition.

Janssen: A lot of other reagents are sold as relatively pure, but when you dig into how they’re sourced, you discover some animal or bacterial product in their making. Some years ago, I was involved in cellular vaccine work, and one of our starting materials came from explanted tumors. We needed buckets of collagenase to break down the tumors into single-cell suspension. One vendor claimed its collagenase was “made to GMP specifications.” So we started qualifying that product as equivalent to the research-grade collagenase we’d been buying from a regular chemical provider. But it turned out that the new collagenase wasn’t synthesized; it came from several different species of bacteria grown in culture. One key ingredient of the broth came from cows, including from the dura mater of those cows. We did not realize this until we received a letter from the FDA (which was sent to many facilities) informing us about it. So even when something seems to be innocuous, it has the potential not to be. You have to look up the production chain.

Human Resources
Janssen: I want to make a pitch for considering the human resources in CGT. There is a serious shortage of technical staffing, people who can do the necessary work to produce these therapies. It’s more complex than assembling an automobile, for example. You need staff with enough scientific training to understand the role of weights and measures and the analytical processes used to make sure that biomanufacturing goes as specified. We don’t see enough ramped-up training programs that could help. I haven’t noticed community colleges starting up training programs for cell- and gene-therapy technologists.

Burger: Exactly. The biotechnology training programs offered by some community colleges are a good model for training cell- and gene-therapy technical operators. Until recently, this was such a small field that there wasn’t enough interest in training people for it, but that’s beginning to change. For now, though, most training that is specific for cell and gene therapy happens on the job. That can require investing six months in training a new employee. This will be one of our biggest challenges over the next five to 10 years: establishing training programs to meet the need for trained personnel.

Janssen: This is where automated systems can help. You can’t completely replace human workers — nor would we want to — but some cell manipulations can be automated with human operators (who then need appropriate training to run the machines). Further development of automated systems — particularly those with robust interfaces and an ability to communicate with each other — will be a key part of moving this from a rapidly growing but nascent field into a prime-time therapeutic option.

Gazaille: What skill sets do cell-therapy technicians need but aren’t likely to get from a traditional biomanufacturing program?

Burger: The biggest difference is that in biomanufacturing, you’re making the products of cells. So the cellular starting materials are consistent and relatively uniform cell lines. You can go to a cell bank, get a vial of cells, and use those to make the product. But when living cells are the product, keeping them alive and functional is our main goal. Every aspect of our manufacturing and testing reflects that, so we have different processing methods and technologies — including our automated processing devices.

Gazaille: What kinds of incentives could training programs use to attract people into CGT work?

Burger: We need to convey that this is a career with a good future, for one thing. Cell and gene therapy has decades of innovation ahead. More tangible incentives are needed too, of course. This has to be a rewarding career in every respect — both personally and financially fulfilling.

Janssen: I would like to have seen more laboratory work featured in the news reporting on the pandemic. With the influenza pandemic of 1918, nobody really knew what a virus was in the way that we do now. Nobody understood DNA or RNA or ribosomes. The reason our current vaccines could be created in record time is that we got the genetic sequencing done so fast. And I wish the general public were more aware of that. Things would be much worse than they are without the work going on in biotechnology laboratories.

Burger: The pandemic vaccines and treatments are built on scientific and technical knowledge that’s been hard won over the course of many years. This is the payoff from decades of research in biomedical science, decades of investment in the National Institutes of Health (NIH) and other research institutions, and a robust biotechnology/biopharmaceutical industry. And it’s the payoff from teaching high school students cell biology so that some would pursue careers in biomedical science. That’s what will get us out of this pandemic.

Janssen: And the next one.

Burger: There’s always another one!

Janssen: That reminds me of the tee-shirts that say, “Not all super heroes wear capes. A lot of us wear lab coats.” We need to get that message out better than we do. Yes, you want people to understand that there’s job security and a financial benefit for going into this field, but they also need to feel that what they’re doing is interesting and important.

Bill Janssen is principal consultant of WEJ Cell and Gene Therapy Consulting Services, LLC, 2640 Ivy Road, Eads, TN 38028; https://www.linkedin.com/in/william-janssen-37015b14/. Scott Burger is principal consultant of Advanced Cell & Gene Therapy, 105 Highgrove Drive, Chapel Hill, NC 27516; 1-984-444-4641; [email protected]; https://www.ac-gt.com. Corresponding author Brian Gazaille is associate editor of BioProcess International, part of Informa Connect; [email protected].