July 1, 2005 (Vol. 25, No. 13)
Appraising the State of the Technology
At a recent conference in Greece, a German researcher described his experience of being shouted down in his home country when he attempted to deliver an invited presentation on embryonic stem cells. On the other side of the world, Korean researchers are talking about human cloning.
Attitudes on this controversial topic are widely divergent, and emotions are running high. The promiseor perceived threatof stem cell work is discussed in the highest circles, but what’s rarely mentioned are the cold, hard facts. The fact is this research is still in its infancy and the technology too often is oversold.
“There is far more to learn than we know now, before the full potential of stem cell therapy can be realized,” notes Ian Wilmut, Ph.D., professor, University of Edinburgh, formerly with the Roslin Institute, and the creator of Dolly the sheep, the world’s first mammal to be cloned from the cell of an adult animal.
“Let me mention a few examples. We need to know which stage of differentiation is best able to integrate with existing cells and function normally and how to differentiate to that stage. In addition, it may be important to obtain pure populations of those cells.
“At present, differentiation of some cells to the desired point is obtained, but rarely are all cells at the same stage of differentiation. We will have to learn how best to deliver the cells and what number are required.”
Above all, he continues, “we will have to demonstrate, perhaps in animal models, that the cells we propose to use are able to function normally for a considerable period of time. It would be ideal to know that they functioned for the length of a human lifespan, but this may not be practicable as the animals that are used are not likely to have sufficiently long lives says Dr. Wilmut, who is also Editor-in-Chief of Cloning and Stem Cells (www.liebertpub.com/clo), which is published by Mary Ann Liebert, Inc., publishers.”
To reach commercialization, “the challenge for both embryonic and adult stem cell researchers is to establish cells as therapeutic agents and to get regulatory and scientific bodies to believe this can be done,” maintains Doug Armstrong, Ph.D., CEO, Aastrom Biosciences (Ann Arbor, MI).
In terms of efficacy, production speed, and expense, “there’s room for improvement,” according to Tom Okarma, M.D., Ph.D., president and CEO, Geron (Menlo Park, CA). In the next several years, “we’ll learn a lot about how to make this better.” And, he says, “We need a battery of safety tests.”
Knowledge of Mechanisms
Much has to be done in terms of understanding growth factors and culture conditions, too. Additionally, researchers need to understand the mechanisms involved in transplant rejection and identify the problems in tissue compatibility in a normal environment. The latter are key issues in allogeneic research.
Beyond that, Dr. Armstrong notes, companies need “cell products that function, and the ability to make them as products. Many companies have had cells with potential, but couldn’t make them as products.”
A host of companies in the 1990s developed business plans based upon autologous stem cells, but found “it was virtually impossible to purify and grow them,” Dr. Armstrong says.
Much of the research is still in preclinical stages, with only a few companies in clinical trials.
“Early clinical trials for specific crippling conditions may begin in a few years,” adds Dr. Wilmut, but “it will then take far longer to reach a point when large numbers of patients are being treated with cells.”
Despite the talk of therapeutic applications bandied about in the press and before legislative bodies, “it’s way too early” to know for certain what’s realistically possible, according to Stephen Chang, Ph.D., president, MultiCell Technologies (Lincoln, RI).
The problem, Dr. Chang says, is the lack of convincing data. “Much information is hype or anecdotal, and anecdotal data are hard to evaluate.”
The situation is akin to the early days of bone marrow transfers or monoclonal antibody development. The promise was great, but achieving that promise took decades.
The approaches to stem cell applications are so different, that some biotech executives bristle at the mention of a stem cell industry. There is even controversy over the term “stem cell.”
Linguistic purists are adamant that stem cells are those that self renew and also make other cell types. Under that definition, many adult stem cells aren’t really stem cells at all, even though they do differentiate into multiple tissue types.
Despite the semantic differences, the term “stem cells” is applied both to pluripotent embryonic stem cells and to adult stem cells, which differentiate but may not be pluripotent.
Adult Stem Cells
According to Dr. Wilmut, “It is very likely that basic research into the molecular mechanisms that regulate development will provide means of changing some cell types to a different cell lineage, but this is a major challenge and could take many years.”
In the meantime, one of the benefits that MultiCell and others working with adult stem cells are exploiting is the cells’ immuno-privileged status. Adult stem cells lack the surface proteins that trigger an immune response.
“They may express them later, but that may be okay,” Dr. Fraser hypothesizes. Adult stem cells are involved only in tissue repair and, therefore, will either repair damaged tissue or be absorbed into the body.
Importantly, “Adult stem cells work by more than just differentiation,” Dr. Fraser says. They make growth factors, including vascular endothelial growth factor (VEGF) and the anti-apoptotic hepatocyte growth factor (HGF). Because of this, when they are injected at an injury site, they trigger the re-growth of injured tissues as needed.
How these cells work is, so far, a mystery, although there are several theories. Solving that conundrum is one of the steps in advancing stem cell work toward practical clinical usage. Although, it must be said, when lives are at stake, knowing that it does work can be more important than knowing how it works.
“At the present time,” Dr. Wilmut adds, “only embryo stem cells have been shown to have the full developmental potential and the lifespan of an adult. However, further research may devise protocols to enhance the developmental potential and lifespan of other cell types. As cells derived from the patient would be immunological matched to them, this would be a major advantage.”
Many of the challenges of using adult stem cells and autologous cell transfers are being solved. Aastrom’s tissue repair cells (TRCs), for example, are derived from bone marrow, Dr. Armstrong says. By not purifying the cells and by growing them outside the body as a mixture with the other bone marrow cells, TRCs don’t have the peripheral growth factor signals that doomed other efforts.
The company’s patented AastromReplicell System is an identical 12-day procedure for each patient treated, that increases the number of progenitor cell populations as much as 250-fold.
“The TRCs have multiline capabilitiesthey can make bone, blood cells, vascular tissue, adipose tissue. We give them to patients and let the site of injury provide the instructions to become a given tissue. They should be an alternative to embryonic stem cells for certain clinical applications,” Dr. Armstrong says.
In a clinical trial involving long bone nonunion fractures, all patients treated with Aastrom’s TRCs exhibited both clinical and functional healing and, at six months, five of the six showed bone regeneration at the fracture site.
As Dr. Armstrong elaborates, “The cells show reliable activity and functionality. Since they’re patient derived, they have no compatibility issues and offer guaranteed delivery to the therapeutic site.” Importantly, these cells can differentiate into vasculature, as well as bone, providing a more complete solution to the injury site, based upon the body’s individual needs at that site.
Next, Aastrom is expanding its trials for bone regeneration. A Phase II trial for severe fractures will be completed in December and another Phase II trial using TRCs to make vascular tissue for diabetic patients with severe limb ischemia is being prepared, along with studies to regenerate spinal bone tissue.
By removing compatibility issues, autologous stem cell research has an advantage over allogeneic therapies. The challenge for those researchers is in proving that autologous therapies are effective. Research so far is promising.
“For persons with a heart attack, if your own cells can provide a benefit, why go elsewhere?” asks John K. Fraser, Ph.D., vp, research and technology, Cytori Therapeutics (San Diego). Autologous stem cell therapy takes from one part of the body to give to another, “so why not take it from fat, which is an excellent source of stem cells,” he continues.
The ratio of total cells to stem cells in bone marrow cells is 100,000:1, so stem cells must be grown up and purified. With adipose cells, however, “fat floats and is easily removed. What’s left has a ratio of total cells to stem cells of 100:1,” Dr. Fraser says. There is no need to grow the cells or to purify them, so the entire process from removal to re-injection takes about one hour.
Cytori and other independent groups have published data showing the benefits of adipose stem cells. Cytori is working with large animals at several academic centers in the U.S. and abroad and, in collaboration with UCLA, with small animals.
MultiCell Technologies also bases its work in adult stem cells. It is developing its line of liver cells in two ways: in a medical device for kidney dialysis and in a drug screening device. The latter is in preclinical trials now and is expected to be commercialized in about three years, Dr. Chang says. It will be used for toxicity testing.
The company was granted a patent this spring that incorporates hepatocyte cells with stem cells that can differentiate into mature, functioning, hepatocyte or bile duct cells that eventually may treat degenerative liver diseases or inherited functional deficiencies in the liver.
It is embryonic stem cells, however, that are at the heart of ethical debates throughout the world. Typically, they are extracted from blastocysts. “Embryonic stem cells are developmental cells that come from a place where their role was to make an organisma heart here, two lungs there, and so on,” notes Cytori’s Dr. Fraser.
Because of this, when you inject an embryonic stem cell into an animal, it forms a teratoma. Extensive differentiation is needed to prevent that malignancy from forming.
To put the challenge in perspective, to inject stem cells into an individual for cardiac therapy, “first, you must make an embryonic stem cell that’s not rejected by the recipient, which involves therapeutic cloning, growing up those cells, differentiating them into heart cells (for example), injecting them, and relying on the differentiation to prevent them from forming a teratoma,” Dr. Fraser says.
Embryonic stem cells do offer some advantages, however. They appear to be the only cells that can differentiate into all cells and tissues in the body, and can renew themselves indefinitely in the undifferentiated state.
Dr. Okarma notes that “human embryonic cells express huge quantities of telomerase (key to cellular immortality), are scalable, and are a renewable source,” making embryonic stem cells amenable to scale-up and manufacturing.
Geron has shown proof of concept injecting oligodendrocytes and dopaminergic neurons derived from human embryonic stem cells directly into the injury site, in this case, into the spinal cord, and “we’re seeing dramatic motor improvement,” notes Dr. Okarma.
An early study to repopulate heart tissue was also successful. Currently, Geron is conducting IND-enabling studies to demonstrate safety in animals and to determine the tumor-causing threshold. “We’re doing spiking studies now,” Dr. Okarma says.
Geron currently has programs under way for cardiac disease, diabetes, osteoarthritis, osteoporosis, and hematology and is working with 8 of the 212 possible cell types. “This is a product-based model, just like [business models] for pills, he says, “made from frozen aliquot.”
ReNeuron (Guildford, U.K.) takes cells from human fetal organs, “which have a reasonable proportion of undifferentiated cells,” according to John Sinden, Ph.D., CSO. The company plans to enter clinical trials for stroke and Huntington’s disease in 2006.
The approach uses what Dr. Sinden calls “a manufactured stem cell therapy.” ReNeuron adds the c-MycER fusion protein to conditionally immortalize the cells. “Take away a chemical constituent of the growth media and the cells will mature into functional nerve cells,” he says. “We have a number of different cell line products already partially attuned to their final cell fate,” Dr. Sinden says.
Its most advanced program is in late preclinical development for stroke-disabled patients. “We’re making a GMP master/working cell bank, following a conventional biopharma development route,” Dr. Sinden says. The goal is to develop allogeneic products (cells in vials) that are safe, scalable, stable, and efficacious.
“We have a number of different cell line products, made to cGMP standards, and many already are partially attuned to their fate,” Dr. Sinden says.
A Third Option
BioE (St. Paul, MN) is positioned neatly between embryonic and adult stem cells, with its Multi-Lineage Progenitor Cells (MLPCs) derived from umbilical cord blood.
“MLPCs are rare, early-stage adult stem cells that are comparable to embryonic stem cells in their ability to differentiate and replicate, in a controlled fashion, into tissues representative of the three germinal layersincluding neural stem cells, nerve cells, liver/pancreas precursors, skeletal muscle, fat cells, bone cells, and blood vessels,” according to Michael Haider, president and CEO.
“We have developed the MLPCs into single-cell clones,” Haider says, “that can expand multiple times without differentiation.” Unlike populations of stem cells, these cells do not differ from one another. Nor do they have the tumorigenic or tetraogenic characteristics that mark embryonic stem cells, he says.
“Compared to adult stem cells, they are more flexible and expressive when it comes to supporting multiple differentiation paths.” These highly characterized stem cells are available for research and can be shipped within 24 hours.
The promise of stem cell research, in many ways, is less about practical therapies and more about sparking the imagination of a generation of biologists. Careers are changing, as researchers’ begin to focus more on cell signaling rather than on molecular biology.
The first commercialized usage of stem cells is likely to be in drug screening. “We believe it is important to use cells from cloned human embryos for research because of concerns that the present cloning procedures may introduce epigenetic abnormalities into the cells,” Dr. Wilmut explains. The research using these cells will itself be useful.
For example, this summer, Dr. Wilmut plans to begin a project to generate stem cells that carry the gene defects that cause motor neurone disease, to learn about the earliest events that lead to cell death. The results will provide new opportunities to study human genetic disease, he says, but comparing the differences between healthy cells and those that inherited the disease.
“At present,” Dr. Wilmut says, “a small number of drugs can be tested in animal models. A high throughput laboratory test would be able to screen hundreds of drugs in the same time and for less cost. The aim is to identify drugs that are able to stabilize the patient so further degeneration does not occur.”
That and other projects will also provide information about the lifespan of stem cells whether they function normally, and other information that, Dr. Wilmut says, “is required before we can think of using cells from cloned embryos to treat disease.”