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Protein Promise

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Checking Oil's ID

An amazing molecule shows promise in the fight against
the nation's No. 1 killer.

In the mid-1990s, a small group of Germans whose hearts were straining to keep them alive signed up to try an experimental drug. As the first people to try the risky treatment, the 20 patients—all suffering from severe heart disease—were facing long odds: doctors had given them two years before the disease would claim their lives.

In the U.S., despite the best modern medicine can do, heart disease kills someone every 37 seconds. The malady remains the deadliest in the country, making the search for the next great heart drug a very glum business. Patients die taking experimental drugs to give future heart disease sufferers a fighting chance.

The Germans chose that risk. So surgeons opened the patients' chests and injected directly into their hearts a compound known as fibroblast growth factor-1—a protein that induces cells to divide.

The results were both thrilling and elegant. As hoped, new vessels sprouted from existing ones, re-routed blood around the patients' coronary arteries that were blocked by fatty plaques. Oxygen-rich blood was able to flow once again to the starving parts of the patients' hearts.

After a second successful test run in Germany, fibroblast growth factor-1 was looking like a magic molecule. On that promise, early testing began in the U.S. and is still under way.

But as with any experimental therapy no matter how tantalizing, this fascinating protein known as FGF-1 has its downsides.

"As far as properties go for a drug, this one is awful," said Michael Blaber, professor of biomedical science at the College of Medicine. Perhaps most importantly, it's a finicky, unstable protein that unfurls when it gets warm.

Blaber ought to know. He has been studying the protein's structure since 1994, around the same time the German trials were running. He has been tweaking the protein atom by atom to see if he could make the protein more stable, and has engineered hundreds of versions of it in the process.

"We've been just blazing away, making all kinds of mutants, asking basic biophysical questions," he said.

Developing a new drug was far from Blaber's mind when he first started tinkering with the problematic protein, but along the way, he just may have created one with palpable promise.

When Blaber was introduced to the growth-factor protein years ago, he did it out of pure, scientific curiosity. As a post-doctoral researcher, he became the first to solve the protein's crystal structure (there are 22 types of fibroblast growth factor). He figured out that the protein's chain of 140 amino acids fold up into a globular shape with a hollow middle and three-fold symmetry—that is, if the protein is standing on one end, it looks triangular from above.

"It was amazing to think that in living systems, part of their underlying atomic structure and basis could be something that was symmetric and absolutely wonderful, and I wanted to study that as an idea," he said. "What are the limitations of symmetric proteins?"

Blaber became fascinated by the growth factor's properties. The symmetry of the protein reminded him of the intricate lattices of snowflakes. He became absorbed in changing the structure to see if the symmetry somehow played a part in its stability. He wondered whether the hollow structure of the protein was responsible for its instability.

Blaber's first hunch was that these structural factors were directly responsible for the protein's instability.

"Along with that," he said, "this architecture has not been found in bacteria. And you have bacteria that live at 100 degrees centigrade (212F). Maybe this architecture isn't that useful to an organism that lives at 100 degrees."

To test his theory, Blaber started with what scientists call the "wild type" protein, the native version that is found in human cells. To make lots of it, he turned bacteria into little manufacturing plants. He inserted DNA for human fibroblast growth factor-1 into bacteria to churn out the precise amino acid sequence that comprises the protein. To make different versions of it, he tweaked the DNA that programs the protein and put that modified DNA in the bacteria so they would produce the custom-designed proteins.

To test the mutants, Blaber broke open the bacteria, separated the growth factor, and heated it up. If it takes more heat to unfold the modified protein than the original, then the mutant is more stable. Thinking he might be able to improve the stability by only a little bit, Blaber started heating up his altered proteins. A few degrees later, many of them remained intact. He added more and more heat.

Blaber was stunned. He had created modified proteins that didn't melt even as the temperature broke water's boiling point.

"(The protein's shape is) intrinsically capable of unbelievable stability," he said. "We have one (modified protein) that has 18 mutations in it. It still folds up in the same architecture, and it won't unfold even at 100 degrees centigrade."

Around 2001, Blaber heard about the success of the clinical trials in Germany and quickly recognized the potentially profound implications his own work could have for the direction of research the German researchers had set.

In FGF-1, the cardiovascular surgeon who led the German trials, Dr. Thomas Stegmann, had discovered a promising drug candidate, but it was unstable, which can be a disastrous trait for a drug. The growth factor could be stabilized with the addition of heparin, a complex sugar, but the molecule is not always innocuous. Heparin has to be isolated from pigs, an expensive process, which also has the risk of being tainted by disease, Blaber said. And it causes allergic reactions in some recipients.

When it became clear that his molecules could be used to treat a deadly disease, and do it more safely than the native protein, Blaber's next move was to take his basic science research to the next level and apply it to something more tangible.

First, he had to start looking at function. A stable protein is useless if it no longer works the way it's supposed to. Blaber set out to test his mutants, feeding dishes of cells with his modified proteins to see if the cells would respond the way they would to FGF-1 in its original form.

His luck held out. Despite the changes Blaber's lab made to the protein, many of the forms still did what the wild type protein is designed to do. Not only that, but some versions caused more cell growth than the original protein.

Blabers' mutants were looking like very promising drug candidates. But any one of them has more tests to pass. As mutants, when administered as a drug in the human body, the immune system might recognize them as a threat and mount an attack that could be harmful, even deadly. But there are ways to try to shield the altered proteins from roaming immune cells on the lookout for foreigners.

Blaber is also not the only one pursuing the perfect FGF-1. He knows of two other labs that are trying to do what he's doing, but Blaber started long before its direct application became obvious.

"Our advantage is that we've been doing this for a while," Blaber said. "We have a large body of data that will be useful. The other folks have been looking at it only in the last year or two. I think we have a great head start."

Ten years after the small group of Germans became the first to receive FGF-1, most had survived, elated, for a celebratory reunion hosted by the original researchers. News reports said two had passed away from causes unrelated to their heart disease.

Though trials involving the protein have only enrolled small numbers of people, no serious side effects have emerged so far. Researchers and patients are hopeful this could be the next life-saving drug for patients suffering from heart disease.

It's too early to tell how Blaber's work will contribute to the development of the protein as a drug, but his research shows enough promise that a private company is investing in it to begin testing his protein forms in animals. He has filed two provisional patents for his modified proteins, with several more in the pipeline.

If the testing goes as smoothly as past trials with the protein have run, in years to come many more cardiac patients will be rejoicing literally with all their heart.


  Protein Promise

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