BioScience Biz

The brave new world of stem cells and human cloning

One of the great promises of embryonic stem cell research is being able to use human cloning to derive stem cells that carry genetic defects associated with myriad maladies. These cells can be used to study the development of tissues that are affected by genetic abnormalities and used as tools for testing new therapies for intractable genetic diseases.

The way that this works is that a researcher derives an embryonic stem cell line from someone with, say Parkinson’s disease. These stem cells can be coaxed into developing into the dopamine-producing neurons that are defective in patients with the disease. Then, a number of different things can be done. For instance, the development of these diseased neurons can be compared to the development of normal neurons in well controlled environments and, hopefully, yield new information on the origins and progression of the disease. Alternatively, the Parkinsonian neurons can be used to test new approaches for treating the disease.

Thus, cloning and derivation of disease-specific stem cells promises to be a powerful and novel tool for studying certain types of cardiovascular disease, certain cancers such as neuroblastoma, Alzheimer’s and Parkinson’s disease, amyotrophic lateral sclerosis (ALS or Lou Gehrig’s disease), metabolic problems such as diabetes, and so on.   

Ethical concerns of cloning human embryos notwithstanding (I am working on a column on this topic that will be posted at a later date), a confounding technical problem is where will researchers find the eggs necessary for the nuclear transfer cloning procedure (the procedure used to clone Dolly, the sheep)? Obtaining human eggs is done routinely at in vitro fertilization clinics, but it does involve hormonal manipulation of young women and a somewhat invasive procedure to harvest the eggs. Who would volunteer for this just so a scientist can do lab research? How many eggs will we need to all the research scientists want to do and are there enough women donors to supply the research needs?

Researchers in England are taking a new approach to deal with the problem of egg supply. They propose to undertake nuclear transfer cloning using eggs from pigs and chromosomes from a human with the desired disease in order to create animal-human hybrid stem cells. A UK regulatory agency recently licensed a laboratory to create human-pig embryos in order to study heart disease.

In fact this is the third animal-human hybrid embryo license to be issued by the British Human Fertilisation and Embryology Authority. In an article just published in the British newspaper, The Telegraph, an HFEA spokesman said it had just approved an application from the Clinical Sciences Research Institute, University of Warwick, for the creation of hybrid embryos. This effort at the University of Warwick is led by Professor Justin St John. "This new license allows us to attempt to make human pig clones to produce embryonic stem cells," he said.

"We will take skin cells from patients who have a mutation for certain kinds of heart disease (cardiomyopathy, which makes the heart lose its pumping strength) and put them into pig eggs after their chromosomes have been removed. We will then make embryos so that we can attempt to derive embryonic stem cells which will allow us to study some of the molecular mechanisms associated with these heart diseases.

"Ultimately they will help us to understand where some of the problems associated with these diseases arise and they could also provide models for the pharmaceutical industry to test new drugs. We will effectively be creating and studying these diseases in a dish.

"But it's important to say that we're at the very early stages of this research and it will take a considerable amount of time. There is still a great deal to learn about these techniques and much of our early work will involve understanding how we can make the hybrid cloning process as efficient as possible."

The study is aimed at understanding the way the cell’s power-producing structures, called mitochondria, are passed from egg to embryo. Mitochondria contain their own small genetic program that produces many of the proteins these organelles need to power cells. Therefore, in the hybrid stem cell, the mitochondria will mostly come from the pig egg, and the researchers will do experiments in order to ensure that the trace of human mitochondria takes over, not least because it is designed to work with human nuclear DNA.

"The key thing we are doing is trying to create stem cells without any animal mitochondria in them. So even though these hybrid embryos normally have…animal mitochondria, we are hoping to create hybrid embryo cells that would have human chromosomes as well human mitochondrial DNA." The reason is that, as the team puts it, "mixing of these two diverse populations of mitochondria can be detrimental to cellular function."

Other research teams in Newcastle and London are also creating human-animal hybrid stem cells. The former have already created hybrids with cow eggs to study genetic regulation in early development, the latter made hybrids with a range of species to generate stem cells from people with neurodegenerative disorders.  Meanwhile, Chinese researchers in Shanghai have reported success in creating human-rabbit hybrid stem cells.

Such research is not allowed in the US, at least in federally-funded labs. But, this does not seem to stop this field from going forward world-wide. Are we in a brave new world, or are we making a Faustian bargain?

Read more on human-hybrid stem cells:

Hybrids: separating hope from the hype

Questions answered on animal-human embryos

Embryo research: a source of hope or horror?

Treating stroke, Parkinson's and other brain diseases with stem cells

Brain repair using genetically engineered embryonic stem cells could offer novel treatments for stroke, Alzheimer's, Parkinson's and other neurological conditions, after encouraging preliminary tests.

Scientists at the Burnham Institute for Medical Research in La Jolla, California, have, for the first time, genetically programmed embryonic stem cells, which have the potential to turn into any type, to become nerve cells when transplanted into the brain, according to a study in The Journal of Neuroscience.

The research showed that mice afflicted by stroke showed "tangible therapeutic improvement" following transplantation and none developed tumors, which had been a major setback in prior transplants.

The team was led by Prof Stuart Lipton, who treats patients with these disorders. "We found that we could create new nerve cells from stem cells, transplant them effectively and make a positive difference in the behavior of the mice," said Prof Lipton.

"These findings could potentially lead to new treatments for stroke and neurodegenerative diseases such as Parkinson's disease."

Prior to this research, creating nerve cells from embryonic cells in a reliable way had been problematic and sometimes cells would seed tumors. Prof Lipton tackled these problems by inducing the stem cells to make a protein called myocyte enhancer factor 2C (MEF2C), which turns on specific genes that drive stem cells to develop into nerve cells.

"We need to have a reliable source of nerve cells that can be easily grown, differentiate in the way that we want them to and remain viable after transplantation," said Prof Lipton.

"MEF2C helps this process first by turning on the genes that, when expressed, make stem cells into nerve cells. It then turns on other genes that keep those new nerve cells from dying. As a result, we were able to produce neuronal progenitor cells that differentiate into a virtually pure population of neurons and survive inside the brain."

The next step was to show whether the transplanted "progenitor" cells became nerve cells that integrated into the existing network of nerve cells in the brain. Performing intricate electrical studies, the team showed that the new nerve cells, derived from the stem cells, could send and receive proper electrical signals to the rest of the brain. The team found that mice which received the transplants showed significant behavioral improvements, although their performance did not reach that of normal control mice.
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Excerpted from an online article by Roger Highfield in the Telegraph on 6/24/08.

New stem cell database is launched to steer scientists through the stem cell maze

Have you ever wondered where a cardiac muscle stem cell comes from and how it is related to a skeletal muscle stem cell?  Or, can a neural stem cell also differentiate into a glial cell?

If you were to drive from San Francisco to New York without any street signs or a road map, you would spend a lot of time going down wrong paths and may never get to the Big Apple. Similarly, as a stem cell is going from its initial, basic level to a complex ear cell, for example, it needs to be steered through a specific path. This map gives scientists not only the knowledge of where they're going, but key "markers" that need to be crossed along the way. Ultimately this will save stem cell companies time and money. 

Read the news article from the San Jose Mercury News, here.  The new stem cell database can be accessed here, www.embryome.com.

FDA delays embryonic stem cell clinical trial

 Geron Corporation, the Menlo Park, California-based company had sought permission from the U.S. Food and Drug administration to begin a human trial to test its GRNOPC1 stem-cell compound in patients with spinal cord injuries. In a press release dated yesterday, Geron anounced that they received oral notice from the FDA had delayed the Investigational New Drug application the company filed in order to begin clinical trials.  This delay does not mean that the IND was rejected, but until Geron receives the official letter, it will not known why the application was delayed or how long it will take to rectify the issues the FDA has.

Geron worked with the FDA over the last four years leading up the filing of a 21,000-page IND application.  Read the full press release here.

Madison's stem cell frontier, By Steve Clark

In an earlier column about the recent Stem Cell Symposium held on the Promega Campus, I extolled the exciting frontier of stem cell basic science that was on display; however, it was just as interesting to catch up with local stem cell researchers who attended the Symposium. I caught a glimpse of the current status of stem cell science in the Madison area.

Other health benefits of embryonic stem cells

For instance, I ran into Tim Kamp, an MD in the UW-Madison Department of Cardiology who, along with Professor Jamie Thomson, recently developed a reliable way to derive human heart cells from embryonic stem cells (ESCs).

About four years ago I first met Kamp in his UW-Madison office to learn about his research. At that time, researchers knew that when given the chance, human ESCs haphazardly differentiate in tissue culture into all the different tissue types and Kamp, using a microscope, had been able to find among the clutter of different cells a few well developed heart cells that actually were beating! You can see a short video clip of one of the beating heart cells here.

Using a steady-handed robot, Kamp inserted a very fine probe into a beating heart cell and measured its depolarization or the exchange of ions across its membrane, which constitutes the electric current that causes heart muscle to beat. With this, he recorded an “EKG” on a single human heart cell that changed as expected when he added to the culture, a drug often given to heart patients.

Currently, animal models are the best way to measure pharmacological effects of drugs on the heart—an important but insufficient model since 30% of drug failures are due to cardiotoxicity. Clearly, we need ways to test drugs on human heart cells, but until the advent of ESCs, there was no reliable way to obtain and grow them in the lab. Now, being able to derive functional heart muscle cells from ESCs provides a important option for testing drugs on real human heart tissue, thereby improving the safety and efficacy of new drugs. At least this was Kamp’s goal four years ago when I talked with him in his office.

Things seem to be progressing well. A couple of years ago, Kamp and his co-workers launched the local biotechnology company, Cellular Dynamics International, in order to bring this technology to fruition. In early March, Roche Palo Alto reached an agreement with CDI to begin using their ESC-derived heart cells for testing the cardiotoxicity of candidate drug compounds.

Using ESCs to derive fully functional mature cell types for testing potential drugs and toxins directly on human tissues is an under-appreciated and poorly communicated application for ESCs, but one that will soon be widely employed in the pharmaceutical industry. Thus, human ESCs will likely play an important role in human health, even if they are never used to directly treat human disease.

Kamp indicated that similar screening methods are being developed for other tissue cell types derived from human ESCs.

Treating neurological diseases

A few years ago, I attended a seminar by UW-Madison neuroscientist, Clive Svendson, who showed a video clip of patients with Parkinson’s disease before and after treatment with a nerve cell factor known as GDNF. The result was a dramatic slowing of disease progression in treated patients.

As encouraging as this therapy was, it remains highly experimental since GDNF cannot cross the blood-brain barrier and must be delivered by cannula—a thin tube inserted deep into the brain area affected by Parkinson’s disease—not an attractive long term option.

Furthermore, GDNF therapy only retards the progressive loss of dopamine producing neurons that is characteristic of Parkinson’s disease; it does not reverse the process. Therefore, this will not likely benefit patients with advanced disease who have lost too many of these critical cells. This is where the hope of stem cell therapy merges with the other great therapeutic hope—gene therapy.

For instance, ESCs alone are not likely to be much of a benefit for patients with Parkinson’s, because stem cell-derived dopamine-producing neurons transplanted in the brains of Parkinsonian patients likely will suffer the same fatal fortune as their endogenous predecessors. But, combine stem cell regeneration of the neurons with in situ production of GDNF via gene therapy technology and you just may be able to sustain dopamine-producing cells for the long term. Or so the hope goes.

A similar idea is being tested in Svendsen’s lab for treating amyotrophic lateral sclerosis; also know as ALS or Lou Gehrig’s disease. Like Parkinson’s, ALS is caused by the progressive and irreversible loss of certain critical neural cells in the brain. Svendsen’s lab has developed rat and primate models of ALS and using human fetal-derived neural stem cells, in conjunction with gene transfer technology, have successfully implanted fully functional, GDNF producing neurons in brains of these ALS animals. The results are very encouraging at this point--they see long-term survival of the transplanted cells and sustained production of GDNF, and these correlate with resolution of symptoms.

This is not the first example using ESCs to successfully treat human diseases in animal models, but ESCs have not yet made it into the clinic. Human trials will likely begin in the next year or two and the FDA is now considering how to best monitor them for safety and efficacy--not a trivial undertaking, but, stay tuned.

Moving stem cell science along

At the symposium I also had the chance to connect with Eric Forsberg, the recently appointed Director of WiCell. This is the non-profit spin-off from the Wisconsin Alumni Research Foundation (WARF) that provides support for stem cell researchers at UW-Madison.

According to Forsberg, WiCell, not only maintains the National Stem Cell Bank, it also engages in outreach activities and provides core services for stem cell researchers that are not found elsewhere on the UW-Madison campus. Forsberg pointed out that WiCell is also happy to provide such support and training for private stem cell companies in order to foster the development of cell-based medicine in Wisconsin..

Forsberg hopes to soon partner with the Waissma Center on the UW-Madison campus to begin a trial run to grow clinical-scale batches of ESCs under the cGMP conditions that are required in order to use the cells to treat patients. The Waissman Center has the cGMP facilities to produce biological materials for clinical use.

This will be a proof-of-principle endeavor designed to show that ESCs can be produced in clinically relevant quantities while maintaining their state of differentiation. Unforeseen problems, the bane of any biotechnology research, will be identified and resolved during this trial run so they will be ready when the time comes to quickly move ESCs into clinical trials.

We rapidly approach the day when ESCs will be used in experimental therapies of human diseases. Probably the first trials will use blood or bone marrow products derived from ESCs as a donor source for marrow transplantation or red cell transfusion.
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© 2008 Steven S. Clark, PhD. Disclaimer: The authors used their best efforts in collecting and preparing the information published herein. However, Steven S. Clark, nor other authors, do not assume, and hereby disclaim, any and all liability for any loss or damage caused by errors or omissions, whether such errors or omissions resulted from negligence, accident, or other causes.

Articles contained herein, are meant to be distributed freely to interested parties. However, any excerpts from any article must credit the BioScience Biz Blog.

The stem cell frontier, 2008, on display, By Steve Clark

Cut a flat worm in two, the tail will grow a new head and the head a new tail. Cut it right down the middle, it will grow a mirror image. How does it know what to grow?

Flies can re-grow damaged tissues. Small fish can regenerate heart muscle. Why can’t humans?

In a developing infant, how do human embryonic stem cells know to grow into heart, muscle, liver, neurological and other cell types at the proper time and in the proper place?

These questions are at the center of science’s brave-new-world of stem cell biology and were the topic of the recent 3^rd Annual Stem Cell Symposium. The science was deep and detailed, and enormously enthralling. It was an intellectual playground of exciting ideas and fabulous potential.

The overriding lesson from the conference is that the mechanism by which stem cells regulate how and when they replenish themselves vs develop into different tissues is conserved in species as divergent as worms, flies, fish and mammals. This is fascinating for developmental biologists, but it also has a profound practical impact for eventually using stem cells to treat human disease. Let me explain how.

A primer on stem cell science

Consider for a moment what needs to be accomplished for an immature stem cell to differentiate into, say a beating heart cell. First, there needs to be a stimulus that initiates this program, telling the stem cell to specifically move along the cardiac muscle developmental pathway. The stem cell must then begin expressing heart cell genes while repressing the expression of all other genes that could cause it to become liver, blood, kidney and all other cell types. Quite a tall order!

Once the stem cell develops into a mature beating heart cell, it remains that type of cell. Mature cells, like zebras, cannot change their stripes. We never see a heart cell become a skin cell and vice versa. Cellular development is unidirectional and this has been one of the central tenants of developmental biology.

Then along comes Scottish scientist, Ian Wilmut, who did an experiment in the mid-1990s that no self-respecting developmental biologist would attempt since we all “knew” that cell development only moved in one direction.

What Wilmut did was to remove the nucleus from an egg cell and replace it with the nucleus from a fully mature cell taken from a different animal. Keep in mind that this donor nucleus had already been directed to express only those genes of the tissue it was taken from and to repress the expression of genes from all other tissues.

Wilmut then transferred this engineered egg into the womb of a pseudo-pregnant sheep, where the engineered egg should have died. Instead, a sheep was born that was a genetic twin of the nucleus donor sheep and the world was introduced to the first cloned animal, Dolly.

This is the type of research result that causes a scientific paradigm shift. For the first time, we realized that the genetic program of a fully developed adult cell, when placed in the proper environment, can be reprogrammed to relinquish its adult cell properties and return to its undifferentiated stem cell state, capable of developing into a fully grown sheep. 

Around the same time that Dolly was born, University of Wisconsin-Madison scientist, Jamie Thomson, published his seminal studies demonstrating the ability to grow monkey and human embryonic stem cells (or ESCs). These, of course, are the immature cells derived from five day old embryos that are able to develop into all tissues of the adult body. The way that ESCs are harvested kills these embryos making ESC research highly controversial. It would be great to be able to obtain such embryonic stem cells without having to destroy a functional human embryo.

Fast forward ten years to the conference where Professor Thomson gave an update on his recent report that he can reprogram adult cells to become stem cells without having to transplant cell nuclei. Looking at recent research from different labs, he noted that only a few regulatory genes are needed to maintain cells in their nascent developmental stage. As the research presented at the conference illustrated, these regulatory genes work across different species, so this mechanism is highly conserved in biology.

Thomson used routine gene transfer technology to induce expression of three different regulatory genes in the cells of mature fibroblasts and, amazingly, the mature cells were re-programmed to become stem cells! What Wilmut was able to do by transferring a cell nucleus to an enucleated egg can now be done in a petri dish and without the egg cell.

At the conference, Thompson explained that these “induced pluripotent cells” or iPCs seem to behave exactly like ESCs. Think about the implication of this observation: it means that mature cells from an adult can be re-programmed back to the stem cell state where they are able to generate anew, all tissues of the human body.

What next for stem cells?

UW-Madison stem cell researcher, Clive Svendson, moderator of the conference, believes that the next major advance will be the ability to develop iPCs by simply changing the environment in which adult cells are grown in the lab, which could be accomplished in about a year. This means that we would not have to insert several genes into a cell’s DNA, which has significant risks and is not a trivial procedure. Thus, it soon may be very easy to take cells from your skin, put them into a defined tissue culture environment and develop stem cells that contain your precise genetic makeup. No embryos would be destroyed and no clones would be created in the process, mitigating most of the ethical concerns.

Svendson opined that this could lead to a big boost in the tissue banking business as people store tissues when they are young for making stem cells if they should need them later. This would be necessary because, as Thomson explained, chronologically young cells are more efficient at being reprogrammed than cells from older animals. One can envision that it could become routine at birth to store placental tissue, the youngest tissue readily available that is genetically identical to the newborn baby.

As exciting as the science was at the conference, there remain some problems to deal with before these stem cells are used in the clinic. First, as with ESCs, undifferentiated iPCs form tumors called teratomas. Therefore, we need to develop a fail-safe way to completely separate or incapacitate contaminating stem cells from the functional tissues grown from them before we put them into patients. According to an article I posted here earlier, the FDA recently convened a meeting to grapple with this problem in anticipation that clinical trials will begin in the near future.

Next, even if we can use stem cells to regenerate damaged tissues, we still need to continue research into the causes of degenerative diseases because simply replacing the dying cells without dealing with what causes them to die may only be a short term fix.

Potential ethical issues may still arise

Finally, and potentially an explosive issue, there remains an ethical question regarding iPCs that no one seems to have addressed. When a mature cell is reprogrammed, how far back does it go? Do iPCs only have the potential to develop into different body tissues, or can an iPC, if given the chance, form an embryo? Of course, if the iPC cells are more like fertilized eggs than stem cells, then all bets are off--the ethical issues will arise again.

I asked both Svendson and Thompson about this and both admitted that this idea had not been tested. Svendson even owned up that no one in the field wants to test it. They do not want to know the answer because it could be very inconvenient.

My prediction

Carl Gulbrandsen, Director of the Wisconsin Alumni Foundation, shared with me that reading Thomson’s paper on reprogramming adult cells to derive embryonic iPCs made him “tingle”. Mr. Gulbrandsen does not seem to be the tingly type, but his response to Thomson’s results was not inappropriate—they are that amazing and significant.

As a scientist, I have learned to be cautious about making predictions. However, I venture one prediction here that I believe has a very good chance of being realized: Professor Jamie Thomson will, in the not-to-distant future, be awarded the Nobel prize for his outstanding work that has created a whole new field of stem cell biology and invigorated the practice of regenerative medicine. While I am at it, you can bet that he will share the prize with Ian Wilmut.

Any takers?
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This post was first published in-part by the Wisconsin Technology Network News

 

© 2008 Steven S. Clark, PhD. Disclaimer: The authors used their best efforts in collecting and preparing the information published herein. However, Steven S. Clark, nor other authors, do not assume, and hereby disclaim, any and all liability for any loss or damage caused by errors or omissions, whether such errors or omissions resulted from negligence, accident, or other causes.

Articles contained herein, are meant to be distributed freely to interested parties. However, any excerpts from any article must credit BioScience Biz.

FDA mulls embryonic stem cell therapy

It is not a matter of if, but of when clinical trials using stem cells to treat human diseases will begin. In anticipation of this, the FDA is considering ahead of time, what kind of oversight they will need to provide when the first clinical trial applications reach their door. Not bad, a government agency thinking proactively! Read on.

Posted on The Scientist NewsBlog by Andrea Gawrylewski

With biotech companies inching up on clinical trials for human embryonic stem cell-based therapies, the US Food and Drug Administration held a meeting yesterday to discuss scientific issues in properly deriving and characterizing the cells, as well as appropriate clinical trial monitoring.

Three biotechs, Geron Corporation, Advanced Cell Technology, and Novocell presented some of their scientific work on spinal cord injury, vision impairment, and diabetes, respectively, at the meeting. Geron and Advanced Cell Technology are hoping to begin testing therapies of cells derived from human embryonic stem cells sometime this year, according to Bloomberg News. Jane Lebkowski, senior vice president of regenerative medicine at Geron Corporation, told The Scientist she could not comment on whether this was true.

"The science was ready to have this kind of discussion, to make sure clinical trials are safe," Celia Witten, spokesperson for the FDA told a group of reporters after the day-long meeting, though she declined to say whether the agency has yet received any Investigational New Drug applications.

The advisory committee, which was made up of 25 independent scientists and FDA researchers, addressed issues of proper animal studies for preclinical testing and how researchers can control the embryonic stem cells for appropriate differentiation -- that is, so they don't form cancerous teratomas.

"There was a lot discussed in the range of things we're already thinking about," Lebkowski told The Scientist. "And several ideas we've been implementing already." Some of the committee's ideas were rather extreme, she added; some of the large animal model studies in non-human primates or pigs that the committee discussed were not very practical and extremely complicated -- the committee mused aloud whether allograft experiments (for example, pig embryonic stem cells transplanted to pig body) should be conducted before transplanting human embryonic stem cells to different species.

Several committee members noted that guidelines and requirements of human embryonic stem cell-based therapies will vary from disease to disease, and cell product to cell product. However, all the members seemed to agree that a common, standardized assay should be developed to determine the tumorigenicity of a specific cell product.

Kenneth Chein, from Harvard Medical School, noted that transplanting cardiomyocites and other types of differentiated embryonic stem cells has yielded mixed efficacy results so far, requiring that more definitive assays need to be developed for efficacy and safety.

The committee also addressed how researchers and clinicians will be able to control and monitor where the cells go once they are administered. While new technologies such as reporter genes may improve researchers' ability to track transplanted cells, some committee members questioned whether new techniques should be used at the same time as therapeutic embryonic stem cells, which itself is a novel type of therapy.

Some members of the committee said they were uneasy about embryonic stem cell therapies, and the committee discussed the potential of developing failsafe mechanisms in the cell products, like suicide genes. But some noted that such approaches may have their own therapeutic complications and risks.
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© 2008 Steven S. Clark, PhD. All Rights Reserved.

Disclaimer: The authors used their best efforts in collecting and preparing the information published herein. However, Steven S. Clark, nor other authors, do not assume, and hereby disclaim, any and all liability for any loss or damage caused by errors or omissions, whether such errors or omissions resulted from negligence, accident, or other causes.

Articles contained herein, are meant to be distributed freely to interested parties. However, any excerpts from any article must credit BioScience Biz.