February 1, 2008 (Vol. 28, No. 3)

Field Is Evolving Rapidly, and Many Firms Have Products Ready for the Clinic

Last month investment banker John Scully and his wife, Regina, donated $20 million to the Stanford University School of Medicine and Stanford Hospital & Clinics to fund work on stem cells.

The gift will help build a new facility to house research programs in stem cells and regenerative medicine at the medical school as well as build space at the new Stanford Hospital, where the results of this research may one day be applied to patients, according to Stanford officials.

The news from Stanford represents the latest example of the continuing support for such a promising potential source of novel therapeutics. “The field has moved to a point where there are clinical proof-of-concept applications for the science,” says Robert Deans, Ph.D., svp at Athersys (www.athersys.com).

Dr. Deans is scheduled to be one of the speakers at Cambridge Healthtech’s “Stem Cells Congress,” which be held next month in San Francisco. The meeting will address stem cell use in multiple applications from halting vision loss, to vascular regeneration, to treating graft vs. host disease.

The nature of this emerging field is such that many of the presenters are examining the applications of stem cells along multiple fronts. “For example, if you make new stem cell transplant tissues in the lab, the organ environment might have an inhibitory influence on the transplant,” says Irina Conboy, Ph.D., assistant professor of bioengineering at the University of California, Berkeley.

“The negative influences of aged or pathological tissues need to be neutralized to harness the regenerative potential of stem cells optimally, and that’s one of the problems we are looking at in my lab.”

Dr. Conboy says that she uses several approaches to explore how the process of tissue repair is controlled, why the injured tissues are not productively repaired as we age, and what approaches restore the regenerative potential to the aged organ stem cells. “Currently, we use the injury regeneration of skeletal muscle as our experimental system,” she says. “But we hope to also identify fundamental mechanisms of aging within stem cell niches, which apply to a variety of organs and tissues.”

Dr. Conboy’s presentation will focus on the effect of aging on tissues. The lack of tissue repair, leading to degeneration and loss of organ function, is an undeniable and devastating trait of aging, she notes. The development of stem-cell based therapies for Parkinson’s, Alzheimer’s, muscular atrophy and other degenerative diseases that accompany human aging requires an improved understanding of why stem cells in older tissue do not engage in repair, even though they have the capacity to do so.

“We found that stem cells residing in aged organs retain their intrinsic ability to regenerate,” Dr. Conboy says, “and that aged, differentiated tissues actually inhibit the responses of endogenous stem cells dedicated to repair and maintenance.”

This same research uncovered the fact that while adult stem cells are unable to deter the inhibitory affects of aged niches, human embryonic stem cells (hESCs) can neutralize the aged environment and robustly restore the regenerative responses of muscle stem cells in the old niche, in vivo and in vitro. “Both the age-specific inhibition and the hESC-specific rejuvenation are conserved between mouse and human, allowing the use of an animal model for identifying therapeutically relevant hESC-derived factors.”

Stemming Vision Loss

There is growing evidence that stem cells can play a key role in ameliorating such eye conditions as age-related macular degeneration (AMD), diabetic retinopathy, and retinopathy of prematurity (ROP).

“Recent advances in the field of vascular biology strongly suggest that specific molecules, already identified as critical to normal angiogenesis, will have utility in preventing the abnormal growth of new blood vessels in the eye,” notes Martin Friedlander, Ph.D., professor of cell biology at the Scripps Research Institute and chief of retina section, division of ophthalmology, Scripps Clinic.

Dr. Friedlander’s team recently demonstrated that combining anti-angiogenic molecules is far more effective than monotherapy in preventing abnormal growth of blood vessels in the eye and tumors and is already moving into the clinics. “We are hopeful that a new therapeutic paradigm, one in which it may be possible to mature or stabilize immature, abnormal vessels, will be of far greater benefit to patients suffering from ischemic retinopathies.”

This may be possible through the use of autologous bone marrow or cord blood-derived hematopoietic stem cells that selectively target sites of neovascularization and gliosis where they provide vasculo- and neurotrophic effects.

“Such a therapeutic approach would obviate the need to employ destructive treatment modalities and would facilitate vascularization of ischemic and otherwise damaged retinal tissue,” says Dr. Friedlander. “Consequently, such treatments would have application to ischemic retinopathies such as ROP and diabetes as well as degenerative retinopathies such as AMD and retinitis pigmentosa.”

In addition to using these cells for their inherent trophic activity, it is possible to transfect them ex vivo, prior to injection into diseased eyes, with plasmids encoding neurotrophic and/or angiostatic molecules in a form of selectively targeted, cell-based therapy.

Dr. Friedlander is also investigating the use of these cells as therapy for a variety of neurodegenerative diseases. “It’s now a matter of engineering a process to move this technology into the clinics.”

Heart Repair

One of the problems that early cell-transplantation research ran into, says Joseph Gold, Ph.D., senior director of stem cell biology and research operations, Geron (www.geron.com), was that researchers would attempt to either implant stem cells in an undeveloped state and hope that they were triggered to differentiate into the appropriate tissue, or transplant fully differentiated cells and hope they would turn into something else.

“One trial attempted to put in skeletal myoblasts hoping they would change, and that didn’t work. Other trials have used mesenchymal stem cells; they neither formed new cardiomyocytes nor survived long term,” notes Dr. Gold. “What we are doing is generating large quantities of bona-fide human cardiomyocytes for transplantation.” Human embryonic stem cells present a potential source for the generation of unlimited quantities of differentiated cells for regenerative medicine.

Geron is investigating the utility of hESC-derived cardiomyocytes for in vitro and therapeutic applications. During the conference, Dr. Gold will describe the efficient, scalable system the company has developed that generates cells with the immunocytochemical and electrophysiological properties of human cardiomyocytes. Dr. Gold will also detail preclinical results that suggest these cells can be used to improve the function of injured hearts. “The goal is to ultimately use these cardiomyocytes to arrest and reverse heart damage,” he says.

This has already been done in small animal models, and results have shown that the animals in which the cardiomyocytes have been transplanted do not show the further typical deterioration seen with heart disease. “We can freeze and thaw them, and they still work,” reports Dr. Gold. “As an off-the-shelf therapy, this is key.”

The crucial step in the company’s research will be when it moves from mouse to pig studies. “A pig’s heart rate and size is comparable to that of a human,” notes Dr. Gold. The large animal study will be an important milestone in this research.

Discovery is driven forward by meeting a previously unmet clinical need. One area that demands attention, notes Dr. Deans, is adjunct therapy in allogeneic bone marrow transplants. “If you look at the research, clinically significant graft versus host disease occurs in greater than 50 percent of matched, and over 70 percent of mismatched allogeneic bone marrow transplants,” he says, “and even with current immunosuppression regimens, it’s a major cause of post transplant mortality.”

Dr. Deans’ team is working with adherent adult stem cells which, he notes, can be a potent source of paracrine factors with significant clinical benefit. These include factors influencing inflammation, homing, angiogenesis, and cytoprotection.

Treatment of Graft vs Host Disease

“We have developed an isolation system for an adherent bone marrow-derived stem cell that possesses strong in vitro immunomodulatory properties and shows significant benefit in graft vs. host disease animal models,” reports Dr. Deans. “The immunology and cell-expansion properties of these pluripotent stem cells support their use as a universal cell product, well suited as an adjunct cell therapy in allogeneic bone marrow transplant.”

The applications for these cells can be manifold, he notes. “We are finding that you can use these cells to stimulate new blood vessel growth in stroke or acute myocardial infarct models. In diseases like stroke, using these cells to replace vascularization in the short term may help cell survival. The models we are developing show that our cell product mediates multiple recovery paths at the same time.”

The result, Dr. Deans says, can be a powerful drug-like product to treat acute injury or inflammation. “The goal remains to create a stem cell bank to produce products that can be used off the shelf, in the short term, with no immunological downsides. Academic clinicians are already treating patients this way on a case-by-case basis.”

According to Dr. Deans, it is possible to make hundreds of thousands of multistem product doses from the adult stem cell bank without any decrease in benefit. “We can make uniform clinical doses to cover large Phase III trial requirements without losing any bioproperties,” confirms Dr. Deans. “The FDA has already formally approved this single, healthy donor bank for use in GVHD and acute myocardial infarct Phase I trials.”

Key Pathways and Biomarkers

There are a number of different ways to approach stem cell research. “We are studying stem cells and other systems using our next-generation ChIP-on-chip technology, ChIP-DSL (chromatin immunoprecipitation DNA selection and ligation), which facilitates genome-wide identification of transcription factor/promoter interactions, epigenetic modifications, and DNA methylation sites,” says Jeffrey Falk, Ph.D., director of technology and business applications, Aviva Systems Biology (www.avivasysbio.com).

ChIP-on-chip technology has been around for eight years but until now has had limited application to stem cell research due to the large number of cells required by traditional methods, says Dr. Falk. “We are identifying key pathways and biomarkers involved in human stem cell differentiation using our ChIP-DSL promoter array technology to map hES cell promoter-methylation patterns in native hES cells and in hES cells at various stages of differentiation.

“We are also using ChIP-DSL to rapidly obtain a genome-wide overview of promoter-methylation patterns in stem cells at various differentiation stages. When cells differentiate, we are able to map the changes in methylation and expression of genes. By knowing and correlating these changes it gives us insight about what has caused the changes in the cell required for differentiation, and what is going to happen next.”

Differentiation of stem cells is an application that is taking off, and Aviva is talking to a number of stem cell companies about some of the growing needs in the field. “One advantage is that, in addition to being a ChIP-on-chip company, we also have a high-throughput antibody production facility that has manufactured antibodies to the entire human and mouse transcription factor families,” notes Dr. Falk. “Consequently, if you have a need for antibodies, we’ll have one that works, which gives us a big advantage in ChIP-on-chip studies that are driven by antibodies.”

At present the biggest downside to the stem cell research is the cost. “Getting a lot of stem cells is costly,” notes Dr. Falk. Also, every time you get into a new area of research people are hesitant to get involved, and directed differentiation of stem cells into specific tissue types such as pancreas beta cells and hepatocytes is really in its infancy.

“It is similar to gene therapy in some respects with some of the same problems. It’s a pretty big jump to get something that differentiates in the lab to actually work in humans. We have a long way to go.”

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