With their amazing ability to develop into any cell type in the human body, pluripotent stem cells (PSCs) hold great promise for medicine – although it’s only recently that they have begun to live up to their potential. Once associated exclusively with the need to harvest cells from embryos — a controversial matter, to say the least — the discovery of induced pluripotent stem cells (IPSCs) in 2006 significantly altered their public perception and medical prospects. IPSCs can be generated from adult tissue and cells — even harvested from our own skin — and turned into stem cells largely indistinguishable from those derived from an embryo. This technology earned researchers John B. Gurdon and Shinya Yamanaka the Nobel Prize in Physiology or Medicine 2012. It also opened up the possibility that each of us could be our own best source of new cells to replace or repair parts of our bodies.
As much as that possibility may smack of science fiction, it is being explored in earnest. According to University of Toronto chemical engineer Peter Zandstra, who holds the Canada Research Chair in Stem Cell Bioengineering, in North America alone there are hundreds of early-stage clinical trials being conducted to evaluate the potential of adult and pluripotent stem cells in treating disease. If even a fraction of those efforts reveal some new approach to a lingering medical challenge, such as eliminating congenital disabilities or growing tissues for transplantation, interest in this field will spike, along with the public demand for access to the resulting treatment.
Peter Zandstra, a Canada Research Chair in Stem Cell Bioengineering and professor at the University of Toronto’s Institute of Biomaterials and Biomedical Engineering, manipulates stem cells with the aim of generating functional tissues. Photo credit: Kathryn Boon
Long before such milestones can be celebrated, however, Zandstra notes that these trials will have to pass subsequent stages leading up to the use of stem cells in human beings. Along the way, he says, researchers will also have to learn a great deal more about how stem cells work, including what it takes to produce them and their functional derivatives in amounts large enough for practical medical purposes. Zandstra’s laboratory at the U of T’s Institute of Biomaterials and Biomedical Engineering is poised to do just that. “Where we’re moving to is the next stage of manufacturing high-value biologics, where the cells are actually the products,” he says. “This is very exciting. If you can manufacture cells under controlled conditions, there are a number of diseases that you might be able to treat. Pluripotent cells represent a nice input population for the generation of many types of cells.”
Zandstra admits this was nothing he could have envisioned 12 years ago, when his doctoral work focused simply on how to grow stems cells found in the blood of umbilical cords. Such cells were assembled in simple layers in petri dishes or floated around in modest flasks, surrounded by essential nutrients. Success there was measured merely in terms of keeping populations and properties at numbers similar to their starting points, never mind coping with the logistics of generating much larger numbers of cells.
Since then, Zandstra and his colleagues have broadened their horizons, growing much larger numbers of PSCs in bioreactors, which can handle one- or two-litre batches of cells. From there, he says, turning out the tens of litres that would be needed for widespread biomedical applications begins to look much more feasible. In fact, there are solid precedents for this kind of activity. “Chemical engineers have been involved in producing biological products at scale for a number of years,” Zandstra says, pointing to the Novartis drug imatinib as an outstanding example. Marketed as Gleevec, this drug set a new standard in chemotherapy by killing only cancer cells and leaving healthy cells alone. Such targeted treatment has been made possible through the creation of cells that combine the properties of tumour cells and mammalian cells to yield antibodies tailored to fight this disease.
The development of imatinib in the 1990s was paralleled by the development of equipment to nurture and maintain large numbers of these manufactured cells. Zandstra anticipates that much of what has been learned about the mechanics of this kind of cell production will eventually be applied to a similar scale-up for PSCs and some other types of stem cells. The next phase of this activity should take on the trappings of an engineering venture, dwelling on details such as the shear forces generated by the stirring impellers in a bioreactor, the cost of production and the development of technologies to control the cell environment. “People are taking the manufacturing systems that we’ve used for 20 years for antibodies and bacterial-based protein production and trying to adapt them to stem cell uses,” Zandstra says. “As these technologies start to come more into industrial and clinical use, there are further opportunities for the customization of those devices, to deal with aspects such as shear or how you harvest cells.”
Zandstra says that this work could be logistically daunting, since the same feature that makes stem cells so attractive — their ability to differentiate into functional cell types — also make them difficult to maintain in their native, unaltered state. For example, within a large aggregate of such cells, slight variations in the concentration of oxygen, metabolites or signaling proteins (cytokines) could lead a particular group of cells to change. That change could quickly propagate through the cell population.
Zandstra and his colleagues have been considering ways in which stem cells might be isolated and grown singly, which would allow for better control of the ambient conditions. Laboratories at the U of T and the University of Calgary last year published the results of work that demonstrated the feasibility of this process using mouse cells, which subsequently grew in the suspended culture of a bioreactor.
Life in the bioreactor can still get complicated for these single cells, says Michael Kallos, a professor at the U of C’s Department of Chemical and Petroleum Engineering. Kallos remembers working as a graduate student to solve some of the mechanical problems around growing recently discovered neural stem cells. That was in 1995, when his background in biology was skimpy enough that he would confess to not being all that sure what a stem cell was. Nevertheless, he could see that the project was being held back by limitations of static culture methods such as petri dishes, which did not allow large numbers of these cells to flourish. He proposed placing the cells in a dynamic culture, specifically a stirred tank bioreactor. “It provides them with a well-mixed environment, so they’re getting a lot of nutrients and other things,” Kallos says. These “other things” turned out to be most interesting. While the stirring action of the reactor had to be moderated to prevent shear forces from tearing the cells apart, the steady movement of fluids around the cells appeared to keep them in an undifferentiated state much longer that if they were in a static setting. “Something about the shear affects the pathways and the signalling inside the cell to turn on some of these pluripotency genes,” he says. “So we can use that to our advantage when we’re making IPSCs or growing lots of pluripotent stem cells.”
This bioreactor, in contrast to petri dishes or flasks, has impellers that circulate nutrients, enhancing stem cell growth rates to nurture the numbers needed for medical research. Photo credit: Kallos lab: Kathryn Boon
For Kallos, who now knows far more about stem cells than he ever could have imagined in 1995, this observation raises tantalizing questions about what happens to them in a bioreactor. If a Goldilocks-like flow regime can be established for these cells — not too fast, not too slow — their pluripotency could be optimized and even larger batches grown. Kallos is now working with other engineers who specialize in oil and gas reservoir dynamics to obtain specific calculations that could set the stage for this kind of optimization.
In the meantime, other researchers continue to refine the process of obtaining stem cells from adult tissue. Freda Miller, a cell and molecular development neurobiologist at The Hospital for Sick Children Research Institute in Toronto, has pioneered a method of extracting skin-derived precursor (SKP) cells that can assist with growing new skin on burn victims. Those cells are at the heart of Kallos’ latest undertaking, which would find ways of growing enough of these SKP cells to assist with sizeable skin grafts. Eventually, such grafts might be able to replace patches of scar tissue long after a burn has healed, thereby restoring the function of the skin.
Despite the tantalizing promise of such therapies, the allure of stem cells will fade if they turn out to be too cumbersome or expensive to produce. Whether any of us can take advantage of this technology will depend very much on work that makes the production of these cells more efficient, and above all cost effective. Zandstra offers an outstanding example: a collaboration between United Kingdom laboratory equipment maker TAP Biosystems and Canada’s Centre for Commercialization of Regenerative Medicine (CCRM), a not-for-profit network that includes universities and private companies.
Zandstra, who is CCRM’s Chief Scientific Officer, describes the partnership’s primary objective as making stem cells more affordable as medicine. “TAP and CCRM are working together to develop suspension culture processes in bioreactors that can be translated for commercial manufacture of stem cell therapies,” he says. “The goal is to reduce costs without compromising cell quality.”
Zandstra adds that it is crucial for Canadians to remain engaged in this field, which means continuing to put people and money into stem cell technology even as its clinical applications are still being explored. It may look like the future for the time being, but once its value is established within the field of medicine, Zandstra believes that stem cell production will be stampeded by eager competitors. “Canada is at the forefront of this right now,” he says. “But we really need to continue investing in the area to stay at the forefront, or else it’s going to go somewhere else.”