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9 posts from September 2010

September 30, 2010

Tread lightly in the valley of death...

By David Kent

Last week, a major kerfuffle erupted in the UK stem cell community as leading scientists and pro stem cell organizations argued that Britain’s range of economic austerity measures would jeopardize their ability to be a major player in translational research. Articles in Reuters and the Daily Telegraph (here and here) detail the cries of the scientific community in response to the latest measures by the UK Government to curb spending. This also comes amidst a major “science rally” being organized by Science is Vital to stop reductions to the science community in general.  

The stem cell specific cries, however, are a little different in this particular case as they are mostly focused on investment into translational research, where the concern, according to Chris Mason, is that Britain appears to have a mantra of “invented here, commercialized elsewhere”. Basically, science in the UK is given good support, but the translation of that science into the clinic is not supported and instead feels like a trip through the “valley of death” according to Sir Richard Sykes of the UK Stem Cell Foundation. Instead of being developed in Britain, companies from America and Asia are offering packages that are much more lucrative to those looking to commercialize.

I am not convinced, however, that large-scale UK government support behind such commercialization efforts is the best solution for the thirsty scientist that steps into the valley. There will always be more money available somewhere else in a more “high risk, high reward” society – the problem for me is that we are not talking about typical inventions or products: we have people’s lives at stake. The comments last week are exactly the sort of rhetoric that offers false hope to those who have been waiting for stem cell therapies to become a reality by suggesting that the only thing stopping patients from receiving bone fide stem cell treatments is a series of tax breaks or government funding supporting the biotechnology and pharmaceutical sector. While I am certainly not against the commercialization of promising therapies and/or products, I do worry that hand waving promises and financial opportunities will guide the field as opposed to good old-fashioned sensibilities.  Investment should be made, no doubt about it, but not simply because American companies might steal the intellectual property away from the UK, but rather because they are promising therapies that behoove a government to act on behalf of its citizenry. 

The reality is that while stem cells hold an enormous amount of promise with respect to regenerative medicine, the safety and validity of proposed treatments has sometimes been overlooked in the bid to create a successful biotech startup. This is why initiatives such as the International Society for Stem Cell Research’s Closer Look at Stem Cell Treatments are so critical. In this rare progressive move from a scientific society, stem cell scientists from around the globe have banded together to help police and offer advice on the numerous stem cell therapy options available, including a patient handbook in five different languages and cautionary videos from world leading researchers.

We need to proceed with cautious optimism when it comes to stem cell therapy. As we learned the very hard way in the gene therapy field not so long ago, the promise (and indeed the successful treatment of disease) can be so attractive that we allow ourselves to take major risks that have severe consequences. More funding for stem cell scientists to do it the right way is fine, but not if the caveat is to produce therapies “now” just for some extra funding over the next few years – the science community will have to answer for it sooner or later.  

September 28, 2010

Even in pharmaceuticals, it's an RNA world

by Paul Krzyzanowski

In the RNA world hypothesis, RNA based biological life can exist without the need for DNA and proteins to store information, make decisions, and in general, control cells. In 1998, the discovery that small RNAs can play important roles in controlling cells revolutionized biology and perhaps brought us closer to an RNA world.

Since then, the promise that small RNA biology offered for the development of novel biotechnologies is slowly being delivered.  Most existing pharmaceuticals function through molecular interactions between drugs or antibodies and their target proteins, but to anyone familiar with biological research labs, RNA interference (RNAi) is a common tool used to reduce expression of proteins in most cell types, including human tissues.  Small interfering RNAs (siRNAs) are used to reduce levels of messenger RNA (mRNA) before it has a chance of being translated into protein.  RNAi based drugs can therefore, in principle, work like any traditional pharmaceutical against a target molecule.

The design of novel drugs is a difficult process, and here RNA-based pharmaceuticals have one key advantage: siRNAs target genes with similar sequences. This feature simplifies the pharmaceutical design phase and results in active molecules being identified more rapidly for a particular application.

Not surprisingly, siRNA molecules have already appeared in numerous development pipelines. Targets can range from transcripts encoding proteins used by viruses for replication (Alnylam's ALN-RSV01 interferes with Respiratory Syncytial Virus) to human enzymes that contribute to chronic disease states like high cholesterol levels (Alnylam is also developing molecules to reduce Low Density Cholesterol levels).

However, several debilitating human diseases require more subtle approaches. Instead of simply eliminating offending mRNA, in some cases siRNA-based pharmaceuticals can also alter proteins being produced.  This goal requires some more knowledge of the biology surrounding the intended target genes. 

In higher organisms like humans, mRNAs encoding protein are usually spliced before being translated to include or exclude portions. This naturally occurring process allows the production of alternative proteins from the same initial mRNA, and siRNAs can be used to control this process. During cell development, it's been recently reported that large-scale RNA switching occurs in developing muscle cells and that splicing also plays important roles in embryonic stem cell differentiation. RNA splicing was already recognized as a potential target for antisense therapeutics back in 2003 by researchers Peter Sazani and Ryszard Kole at the University of North Carolina.

One of the leading examples of RNA splicing drugs is Isis pharma's ISIS-SMNRx which is currently going through pre-clinical development as an experimental cure for Spinal Muscular Atrophy (SMA).  In SMA, the deletion of a gene eliminates expression of the Survival Motor Neuron (SMN) protein.  ISIS-SMNRx can effectively alter the splicing of a similar gene to mimic production of SMN, a technique that has been shown to increase levels of this protein in mice, and may ultimately be able to compensate for the underlying genetic defect in humans.

Development of similar products to control Dystrophin RNA splicing is being pursued by AVI Biopharma as one cure for Duchenne Muscular Dystrophy. AVI acquired Ercole Biotech Inc. in 2008 for approximately $7.5 million, a company developing siRNA switching technology which was co-founded by Ryszard Kole.

The siRNA field is exciting and it’s clear that the biological mechanisms capable of being targeted pharmaceutically are becoming much more complex as science develops. As the technology matures, fundamental processes of stem cell growth being revealed might also be controlled by siRNAs to enhance healing and regeneration, and these discoveries may foreshadow future opportunities in therapeutic settings for ambitious researchers.

September 23, 2010

Fluid shear stress promotes cell differentiation

by Allison Van Winkle

The transplantation of stem cells for use in regenerative medicine, where diseased or degenerated tissue is replaced with a new cell source, is an exciting field of research. However, prior to the implementation of a cell therapy, large amounts of cells will be required, and a consistent protocol for the development of functional cells, in large homogeneous populations must be developed for each cell type of interest.  

Endothelial cells have potential for use in cardiovascular applications, such as promoting vascularisation within the host tissue near an implanted tissue engineered product. Endothelial cells have been previously differentiated from embryonic stem cells; one common differentiation method is to culture the cells in a medium containing growth factors relevant to endothelial lineage differentiation. However, this method results in a low efficiency of endothelial differentiation.

In vivo, endothelial cells line the surface of blood vessels. They regularly experience a fluid shear stress due to the constant fluid movement of blood through the circulatory system.  By applying a shear stress to murine embryonic stem cells in vitro, at levels similar to those experienced by endothelial cells in vivo, researchers were able to obtain a much higher yield of differentiation; 1% of cells differentiated without shear stress, while 40% of cells differentiated with the application of constant shear stress. Other studies have also looked at endothelial cell differentiation with respect to shear stress, from initial cell populations of bone marrow mesenchymal stem cells, and adipose-derived stem cells, where shear stress was also found to increase endothelial differentiation.

These significant increases in differentiation yields are important with respect to both the potential use of endothelial cells in regenerative medicine, as well as in other stem cell fields. While recreating the in vivo environment is not a new approach, it is shown to be a very effective strategy with regards to enhancing differentiation. The in vivo environment may be something that other researchers should consider in when developing differentiation protocols for stem cells.

Future research considerations may include applying this technique to human cells and the purification of the resulting cell population, so that transplanted cells perform only the desired function.

September 21, 2010

In the blood – part two

Tidy_plasmaimgby Michelle Ly

In my last post, I introduced the use of clinical stem cell therapy in treating multiple myelomas and lymphomas. The treatment focuses blood stem cells, known as hematopoietic progenitor cells (HPCs). By transplanting healthy HPCs into patients, nearly normal white blood cell counts can be restored after cell-destroying cancer treatment.

The process begins not in the lab, but in the hospital, with a procedure called apheresis. In this procedure, the patient's or donor's blood passes through a cell-separator which uses a centrifuge to separate the whole blood into layers. The HPC and plasma layers are skimmed into a sterile collection bag and this “blood product” is speedily brought to the lab for processing. The remaining layers are returned to the donor's circulation.

Back in the lab, several technicians are required to complete the processing. A sealed-off “clean” area is dedicated to processing the patient sample. Unlike a research lab, a lab working with patient samples must carefully track all materials used. Each item, from pipettes to PVC tubing to plastic disposal bags, are labelled with tracking numbers and (for sterile items) expiry dates.

Preparing HPCs for storage is a three stage procedure. The initial stage is cell recovery. The technician checks the quantity and quality of the sample by taking a small sample of blood and determining the number of nucleated cells. If within reason, the blood product is transferred from the collection bag to a centrifuge bag and spun down. This separates the plasma layer from the HPC-rich layer and also reduces the volume of product to freeze.

The plasma is carefully siphoned off into a separate bag and discarded, while the remaining blood product is weighed and a cell count taken. If the cell count is approximately 90% or better of the original count, then it is considered a good recovery and the procedure can continue.

Now the blood product is rich in HPCs but not yet in a form that is appropriate for long term storage. Left as is, the cells would die. Here, we enter the second stage of the process, preparation for storage.

From the centrifuge bag, the blood product is again transferred, this time to a sterile freezing bag. Separately, a preparation of 20% dimethyl sulfoxide (DMSO) is made in isotonic solution, equal to the volume of cells. Both bags are pre-cooled and the DMSO solution is added to the freezing bag. The final solution is 10% DMSO by volume, which protects the cells from the effects of freezing - the final stage of the process.

Slowly, the HPC-DMSO mixture is frozen in a specialized freezer before being transferred to liquid nitrogen tanks. It is here that the pouches of HPC-rich product reside, at a numbing -195°C, until the day that the patient requires a transplant.

(Photo by Dale Tidy)

September 17, 2010

What's in a label?

By Katie Moisse

Replacing dead or dying cells with new, healthy ones is the holy grail of regenerative medicine. Even sustaining damaged cells with toxin-mopping, growth-factor-spewing stem-cell-derived support cells would be a tremendous feat.  

But while clinical trials have hinted at stem cell therapy's tremendous potential to replace parts or at least aid in necessary maintenance, the details of how these cells work their therapeutic magic often fall through the methodological cracks.  

It's all very well to conclude that injecting stem cells into damaged organs does some good. But in order to understand whether these cells are homing to injury sites and setting up shop, or even surviving at all, they have to be traceable. 

Let’s look at the heart. A study assessing the acute and long-term effects of intracoronary stem cell transplantation in patients with chronic heart failure (the STAR-heart study), published in the July issue of the European Journal of Heart Failure but re-reported August 29 at the European Society of Cardiology annual meeting in Stockholm, Sweden, suggests the therapy improves ventricular performance, quality of life and survival. 

But the mechanism by which 6.6 x 107 bone marrow cells confer protection when injected into damaged hearts is not explained in the report. The researchers posit that the transplant “may overcome the possibly detrimental effects of ventricular remodeling,” but later add that “further studies are required which focus on cell based therapy.” 

Not knowing where these cells are going or what type of role they’re taking on not only makes it difficult to optimize the therapy, it also represents a missed opportunity to learn something about the diseases or conditions themselves.

Incorporating reporter genes, magnetic or radioactive labels could allow researchers to follow transplanted cells to their final destination using non-invasive imaging techniques like PET and MRI, offering clues to their newfound function. There are obvious obstacles to consider before applying such approaches widely in humans though, such as immunogenicity of integrated gene products, the possibility that patients might depend on medical devices (such as pacemakers) that preclude them from having MRIs, and radiation exposure.

However, until these marvelous but mysterious cells can be tracked -- even just identified in post-mortem tissue -- we simply won’t know for sure what they’re doing. This creates problems in the era of stem cell “clinics” selling phony medical miracles based on cell therapies. Clinical trials that show stem cells work but offer no explanation as to how continue to leave open the door for such abuse. 

 

September 14, 2010

Enhancing cancer stem cell drugs

by Chris Kamel

BroccoliWhen cells are subject certain types of mild stress, it activates a protein called NF-κB and downstream pathways that can lead to future stress resistance in a process called pre-conditioning. This is useful for preparing cells for transplant into harsh environments. But there are other situations where you don't want these same survival proteins active and keeping cells alive.

One such instance is when targetting cells with chemotherapy. Cells that survive treatment can regrow into tumours that can be resistant to further treatment. Sorafenib is a drug that inhibits pathways involved in tumour blood-vessel growth, and tumour progression in general, and has been implicated in targeting cancer stem cells. It has been used to treat kidney and liver cancers, and more recently tested for use with pancreatic cancer.

Pancreatic cancer is increasingly thought to be driven by cancer stem cells. Sorafenib has shown anti-tumour activity in pancreatic cancer, but its effectiveness is limited: after an initial responsive period, the cells bounce back with enhanced tumour progression and increased metastasis. This is possibly due to a cancer stem cell population that survives treatment, leading to this renewed growth. One reason these cells survive is that sorafenib activates NF-κB survival pathways.

Recent research published in Cancer Research attempts to enhance the effectiveness of sorafenib by interfering with NF-κB. In this paper, the authors show that sorafenib reduces -- but doesn't completely eliminate -- the stem cell properties of pancreatic cancer cell lines. Some cells survive, and with time regrow. The same was shown in vivo when cancer stem cell rich pancreatic cells were transplanted into mice. However, when the cells were pre-treated with a different chemical, sulforaphane, before applying the drug, there was enhanced cell death and increased inhibition of stem cell characteristics such as morphology and colony and spheroid formation compared to either compound on its own. Furthermore, addition of sulforaphane interfered with cell survival pathways, completely abolishing sorafenib-dependent NF-κB activation. This enhancement of sorafenib activity was seen both in vitro and in mouse models for pancreatic cancer. This suggests that combination with an NF-κB inhibitor such as sulforaphane could be a therapeutic option for improving certain cancer therapies that target cancer stem cells.

So where does sulforaphane come from? You can get it from your diet. Foods like broccoli and cauliflower are rich in the anti-cancer compound, hinting at nutritional ways to break cancer stem cell resistance to therapy.

All the more reason to eat your veggies.

September 10, 2010

Suspension issued by US Court of Appeals on temporary injunction

by Ubaka Ogbogu

The US Court of Appeals in Washington DC yesterday suspended a recent temporary injunction issued by a federal district judge against the use of federal funds for hESC research. Read my earlier post on the injunction. The suspension is temporary, and is intended to allow the appellate court sufficient opportunity to review an emergency appeal filed by the federal government against the district judge’s ruling. In the meantime, activities affected by the injunction can resume

Whether or not the government’s appeal succeeds, this state of affairs does not bode well for US stem cell policy. The courts are hardly the right venue for setting science policy, especially in an area fraught with social controversy. Opponents of hESC research are also more likely to view any pro-hESC research policy as legitimate if it issued by Congress instead of by way of technical judicial interpretations. 


September 08, 2010

Not your ordinary art contest

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This year marks the third annual Cells I See art contest, and it's ramping up to be another hotly contested battle to see who wins the highly coveted grand prize for the best stem cell-inspired art. This year, it's $500 cash, courtesy of contest sponsor Fate Therapeutics. But that grand prize is only one of many great reasons to enter the contest--and here's a list of the top ten reasons you should enter:

10. Stem cells are beautiful. Don’t believe us? Check out the gallery of past Cells I See entries, and try to tell us otherwise.

9. Cells I See has a great new sponsor: Fate Therapeutics. Named in 2010 as one of the world’s 50 most innovative companies, they ought to know a good thing when they see it.

8. Fame. Well, a little of it – entries from past years have been showcased on postcards, calendars, international magazines, web sites and science exhibits.

7. Fortune. The winning entrant will receive a grand prize of $500 in cold, hard cash. Not too shabby!

6. The cool factor. Who doesn’t want to include prize-winning artist on their résumé?

5. Own a bit of the podium. The winner gets three minutes on the soapbox to talk about their image, their work and their thoughts about stem cells during the closing plenary at the SCN annual scientific meeting.

4. Get more people interested in stem cells. It’s a visual world, and for many, seeing is believing.

3. Build brain cells. Stretch your creative mind to see just what you can do with cells, stain and a microscope.

2. Notoriety in the stem cell circle. All entries will be viewed and judged by more than 450 Canadian and international stem cell experts during the SCN annual scientific meeting. That's 900 eyeballs on your work.

1. It’s easy and free to enter—what have you got to lose?

Check out stemcellnetwork.ca/CellsISee for full contest and entry details.

September 02, 2010

Ghosts of stem cells past

by Chris Kamel

One of the coolest breakthroughs of the last five years is the ability to reprogram adult, differentiated cells into pluripotent cells, effectively allowing us to change one cell type into virtually any other. Reprogramming is achieved by expression of a set of genes that yield induced pluripotent stem cells (iPS cells), which have many of the properties of embryonic stem cells (ES cells). These iPS cells are exciting both as a potential source of stem cells for regenerative medicine, and as a tool for better understanding disease and developmental processes. This excitement led Nature Methods to name iPS cells "Method of the Year" in 2009. (Two other methods for reprogramming adult cells are somatic cell nuclear transfer and cell fusion; all three methods are nicely summarized in a recent Nature review.)

Despite their strong similarities to ES cells, such as self-renewal and pluripotency, there is still some question as to exactly how ES-like iPS cells are. Two recent papers ask this question, with interesting results: each iPS cell retained "memories" of its former life.

Research published in Nature Biotechnology demonstrated that despite being genetically identical, iPS cells derived from different parental cell types had differential gene expression that seemed to reflect the cell of origin. For example, iPS cells derived from mouse granulocytes, a type of white blood cell, showed higher expression of myeloid-cell associated genes such as lysozyme and Gr-1 compared to iPS cell derived from skeletal muscle. Likewise, muscle-derived iPS cells had higher levels of muscle-related genes. This was due to different epigenetic patterns - hereditable modifications that can dictate which genes are expressed and which aren't without altering the underlying sequence. Furthermore, iPS cells from different origins had different differentiation potential. Compared to muscle- or fibroblast-derived cells, those derived from granulocytes or B-cells more readily differentiated into blood cells, forming more erythrocyte progenitors, macrophages and mixed hematopoietic colonies.

In work published in Nature, similar results were seen. Pluripotent cells derived from murine blood cells consistently produced more hematopoietic colonies compared to fibroblast-derived cells, which themselves more readily differentiated into bone-forming cells. Again, induced pluripotency led to different epigenetic and gene expression profiles and stem cells derived by somatic cell nuclear transfer were more similar to classic embryonic stem cells than the iPS lines.

Both these studies indicate that not all iPS cells are equal and that induced pluripotent cells retain an epigenetic memory of their former lives that can limit differentiation potential. This is important to consider when comparing basic research or drug discovery studies that may use iPS cells of different origins. Both papers suggest ways to get around the epigenetic memory to create more ES-like iPS cells. Serial reprogramming (differentiating an iPS cell, then reinducing pluripotency) or treatment with chromatin-modifying drugs were able to alter epigenetic programming and differentiation potential. Serial passage of cells was also able to reduce epigenetic and functional differences between iPS cell lines, though this approach does come with its own complications.

However, restricted differentiation potential of iPS cells isn't necessarily bad news. Given the challenges of producing stem cells for clinical use and the nuances of directed differentiation, choosing the right starting cell can be a strategic decision. The right choice could lead to improved scaling or transplant efficiency, or exploited to obtain cell types that have, so far, been difficult to produce from ES cells.