Thursday, 10 January 2013

Regulating Single Protein Prompts Fibroblasts to Become Neurons

Regulating Single Protein Prompts Fibroblasts to Become Neurons

Thursday, 10 January 2013

Human fibroblast
This is a confocal micrograph of a primary
human fibroblast cell grown in culture stained
blue for actin, a highly abundant protein that
makes up the cytoskeleton of cells. Energy-
producing mitochondria are shown in green.
Credit: Image courtesy of Matthew Daniels,
University of Oxford and Wellcome Images. 
Repression of a single protein in ordinary fibroblasts is sufficient to directly convert the cells – abundantly found in connective tissues – into functional neurons. The findings, which could have far-reaching implications for the development of new treatments for neurodegenerative diseases like Huntington's, Parkinson's and Alzheimer's, will be published online in advance of the January 17 issue of the journal Cell.

In recent years, scientists have dramatically advanced the ability to induce pluripotent stem cells to become almost any type of cell, a major step in many diverse therapeutic efforts. The new study focuses upon the surprising and singular role of PTB, an RNA-binding protein long known for its role in the regulation of alternative RNA splicing.

In in vitro experiments, scientists at University of California, San Diego School of Medicine and Wuhan University in China describe the protein's notable regulatory role in a feedback loop that also involves microRNA – a class of small molecules that modulate the expression of up to 60 percent of genes in humans. Approximately 800 miRNAs have been identified and characterized to various degrees.

One of these miRNAs, known as miR-124, specifically modulates levels of PTB during brain development. The researchers found that when diverse cell types were depleted of PTB, they became neuronal-like cells or even functional neurons – an unexpected effect. The protein, they determined, functions in a complicated loop that involves a group of transcription factors dubbed REST that silences the expression of neuronal genes in non-neuronal cells.

According to principal investigator Xiang-Dong Fu, PhD, professor of cellular and molecular medicine at UC San Diego, it's not known which neuronal signal or signals turn on the loop, which in principle can happen at any point in the circle. But the ability to artificially manipulate PTB levels in cells, inducing them to become neurons, offers tantalizing possibilities for scientists seeking new treatments for an array of neurodegenerative diseases.

It is estimated that over a lifetime, one in four Americans will suffer from a neurodegenerative disease, from Alzheimer's and Parkinson's to multiple sclerosis and amyotrophic lateral sclerosis (Lou Gehrig's disease).

"All of these diseases are currently incurable. Existing therapies focus on simply trying to preserve neurons or slow the rate of degeneration," said Fu.

"People are working with the idea of replacing lost neurons using embryonic stem cells, but there are a lot of challenges, including issues like the use of foreign DNA and the fact that it's a very complex process with low efficiency."

Fu explained that REST is expressed in cells everywhere except in neurons. PTB is itself a target of miR-124, but also acts as a break for this microRNA to attack other cellular targets that include REST, which is responsible for repressing miR-124.

In non-neuronal cells, REST keeps miR-124 down and PTB enforces this negative feedback loop, but during neural induction, miR-124 is induced, which diminishes PTB, and without PTB as a break, REST is dismantled, and without REST, additional miR-124 is produced. This loop therefore becomes a positive feed forward, which turns non-neuronal cells into neurons.

"If we learn how to manipulate PTB, which appears to be a kind of master regulator, we might eventually be able to avoid some of these problems by creating new neurons in patients using their own cells adjacent deteriorating neurons," said Fu.

Contact: Scott LaFee
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Saturday, 5 January 2013

When Will Genomic Research Translate into Clinical Care - and at What Cost?

New study applies quantitative modelling to genomics

Saturday, 05 January 2013

Genomic research is widely expected to transform medicine, but progress has been slower than expected. While critics argue that the genomics "promise" has been broken – and that money might be better spent elsewhere – proponents say the deliberate pace underscores the complexity of the relationship between medicine and disease and, indeed, argues for more funding.

But thus far, these competing narratives have been based mostly on anecdotes. Ramy Arnaout, MD, DPhil, a founding member of the Genomic Medicine Initiative at Beth Israel Deaconess Medical Center (BIDMC), decided it was time to look at genomics from a new perspective. So he turned to quantitative modelling, a numerical forecasting approach used to predict everything from weather events to the outcomes of political elections, and an extremely useful way to both set expectations and assist in decision-making.

Arnaout and colleagues knew that drug-related adverse outcomes cost the health-care system upwards of $80 billion a year, and that many such cases should be avoidable by choosing and dosing drug prescriptions according to a person's genome. So they developed a quantitative model to estimate how much time and money would be required to use genomics, specifically pharmacogenomics, to cut these adverse outcomes in half. Their findings, currently published online in the journal Clinical Chemistry, provide one of the first examples of data-driven estimates being applied to genomic medicine and offer a template for the use of quantitative modelling in this field.

How do the numbers add up? After analysing their model for a range of situations, the research team found that the cost can be expected to be less than $10 billion, spread out over approximately 20 years.

"If you look across medicine, you can see specific places here and there where genomics is really starting to change things, but it's been hard to know how it all adds up in the big picture," explains Arnaout, who is also an Assistant Professor of Pathology at Harvard Medical School (HMS) and Associate Director of Clinical Microbiology at BIDMC.

"Quantitative modelling is a standard approach for forecasting and setting expectations in many fields as we all remember from the recent presidential election and from the hurricane season. Genomics is so important and is so often on the minds of our patients, students and staff, that it seemed like a good idea to use modelling to get some hard numbers on where we're headed."

The idea for the study originated nearly two years ago, while Arnaout (whose laboratory studies genomics) and Sukhatme, BIDMC's Chief Academic Officer, were attending a lecture, shortly after the 10-year anniversary of the sequencing of the genome.

"Vikas asked me, 'So when is genomics really going to change medicine?'" remembers Arnaout.

"I realized I didn't know. And that got me thinking."

Arnaout and Sukhatme, together with co-authors Thomas Buck, MD, and Paulvalery Roulette, MD, of BIDMC and HMS, decided to try and answer this question by applying forecasting methods to a big clinical problem – drug-related adverse outcomes.

"We know that preventable causes of these adverse outcomes -- patients' non-adherence, interactions between multiple drugs, and medical error, for example -- account for only a fraction of the millions of adverse outcomes that patients experience each year," explains Arnaout.

"This leaves a significant number that are currently considered non-preventable and are thought to be caused by genomic variation."

By way of example, Arnaout explains that 30 million Americans currently use the blood-thinning drug warfarin. But because, in some cases, patients' genomes contain variants that make the standard dose of warfarin too high for them, these individuals are likely to experience bleeding, an extremely dangerous side effect. In fact, researchers now estimate that three-quarters of the variability in warfarin dosing requirement is due to these genomic variants, and they have already identified a set of variants in six specific genes that explain two-thirds of the variability.

"This kind of progress suggested an interesting thought experiment," says Arnaout.

"What if we took existing examples in which there appears to be a carefully vetted, clinically useful connection between a specific adverse outcome and a specific genetic variant, found out how much it cost and how long it took to discover, and applied that model to all drugs? How much would it cost and how long would it take to cut adverse outcomes by 25 percent? How about by half?"

As data for the model, the authors selected eight associations involving six prescription drugs (clopidogrel, warfarin, escitalopram, carbamazepine, the nicotine-replacement patch and abacavir) and one drug class, the statin class of anti-cholesterol drugs.

Using an approach called Monte Carlo modelling, the team ran simulations to forecast the research investment required to learn how to cut adverse outcomes by meaningful amounts, and how long that research work would be expected to take. For statistical confidence, they ran their simulations thousands of times and explored a wide range of assumptions.

"The results were surprising," says Arnaout.

"Before we did this work, I couldn't have told you whether it would take a million dollars or a trillion dollars or whether it would take five years or a hundred years. But now, we've got a basis for thinking that we're looking at single-digit billions of dollars and a couple of decades. That may sound like a lot or a little, depending on your point of view. But with these numbers, we can now have a more informed conversation about planning for the future of genomic medicine."

The most important determinant of the numbers is the extent to which the examples used in the model will turn out to be representative of drugs as a whole.

"It's a broad set of drugs that were used, but we know how the genome can surprise us," says senior author Sukhatme.

"For example, you won't be able to use genomics to cut adverse outcomes in half if genomics turns out to explain less than half of the adverse outcomes. But even in that case, we found that pharmacogenomics will be able to make a significant dent in adverse outcomes – cutting them by a quarter – for multi-billion-dollar investments."

Also surprising, say the authors, was the timing.

"As a rule, the fruits of research come only after research dollars have already been spent," points out Arnaout. This means that, in this case, hundreds of millions of dollars will be spent for "pump-priming" long before the public can expect to see any meaningful clinical impact.

"It's one thing to say, 'Be patient,' based on just faith," he adds.

"It's another to be able to say so based on data and a model. We now have that. This enables the conversation to shift to which indicators of progress to look for, over the five or so years of pump-priming, to make sure we're on track."

Can we go faster?

"If we could enrol an ethnically diverse set of patients who are taking each of the 40 or 50 most commonly prescribed drugs, get their blood samples, and keep track of the adverse outcomes that some of them are bound to experience, we should be able to move faster, for less money," adds Arnaout, who describes this idea as a "50,000 Pharmacogenomes Project," a pursuit along the lines of the 1,000 Genomes Project, the UK10K or the Veteran's Association Million Veteran Program.

"This model provides the start of a provocative conversation and illustrates the value of quantitative modelling in this very practical and publically relevant aspect of genomics," adds BIDMC Chief of Pathology Jeffrey Saffitz, MD, PhD.

"Such models should help both decision makers and the public set expectations and priorities for translating genomic research into better patient care."

Contact: Bonnie Prescott
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For more on stem cells and cloning, go to CellNEWS at

Gene Therapy Reprograms Scar Tissue in Damaged Hearts into Healthy Heart Muscle

Gene Therapy Reprograms Scar Tissue in Damaged Hearts into Healthy Heart Muscle

Saturday, 05 January 2013

A cocktail of three specific genes can reprogram cells in the scars caused by heart attacks into functioning muscle cells, and the addition of a gene that stimulates the growth of blood vessels enhances that effect, said researchers from Weill Cornell Medical College, Baylor College of Medicine and Stony Brook University Medical Center in a report that appears online in the Journal of the American Heart Association.

"The idea of reprogramming scar tissue in the heart into functioning heart muscle was exciting," said Dr. Todd K. Rosengart, chair of the Michael E. DeBakey Department of Surgery at BCM and the report's corresponding author.

"The theory is that if you have a big heart attack, your doctor can just inject these three genes into the scar tissue during surgery and change it back into heart muscle. However, in these animal studies, we found that even the effect is enhanced when combined with the VEGF gene."

"This experiment is a proof of principle," said Dr. Ronald G. Crystal, chairman and professor of genetic medicine at Weill Cornell Medical College and a pioneer in gene therapy, who played an important role in the research.

"Now we need to go further to understand the activity of these genes and determine if they are effective in even larger hearts."

During a heart attack, blood supply is cut off to the heart, resulting in the death of heart muscle. The damage leaves behind a scar and a much weakened heart. Eventually, most people who have had serious heart attacks will develop heart failure.

Changing the scar into heart muscle would strengthen the heart. To accomplish this, during surgery, Rosengart and his colleagues transferred three forms of the vascular endothelial growth factor (VEGF) gene that enhances blood vessel growth or an inactive material (both attached to a gene vector) into the hearts of rats. Three weeks later, the rats received either Gata4, Mef 2c and Tbx5 (the cocktail of transcription factor genes called GMT) or an inactive material. (A transcription factor binds to specific DNA sequences and starts the process that translates the genetic information into a protein.)

The GMT genes alone reduced the amount of scar tissue by half compared to animals that did not receive the genes, and there were more heart muscle cells in the animals that were treated with GMT. The hearts of animals that received GMT alone also worked better as defined by ejection fraction than those who had not received genes. (Ejection fraction refers to the percentage of blood that is pumped out of a filled ventricle or pumping chamber of the heart.)

The hearts of the animals that had received both the GMT and the VEGF gene transfers had an ejection fraction four times greater than that of the animals that had received only the GMT transfer.

Rosengart emphasizes that more work needs to be completed to show that the effect of the VEGF is real, but it has real promise as part of a new treatment for heart attack that would minimize heart damage.

"We have shown both that GMT can effect change that enhances the activity of the heart and that the VEGF gene is effective in improving heart function even more," said Dr. Crystal.

The idea started with the notion of induced pluripotent stem cells – reprograming mature specialized cells into stem cells that are immature and can differentiate into different specific cells needed in the body. Dr. Shinya Yamanaka and Sir John B. Gurdon received the Nobel Prize in Medicine and Physiology for their work toward this goal this year.

However, use of induced pluripotent stem cells has the potential to cause tumors. To get around that, researchers in Dallas and San Francisco used the GMT cocktail to reprogram the scar cells into cardiomyocytes (cells that become heart muscle) in the living animals.

Now Rosengart and his colleagues have gone a step farther – encouraging the production of new blood vessels to provide circulation to the new cells.

Contact: Lauren Woods
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