Fish and Sea Life Yield Some Medical Answers

by Food and Drug Administration (FDA)

Don Hochstein raises a thin glass tube up to his eye level and flicks it with a fingernail. Inside the pencil-width vessel, a substance with the texture of gelatin shimmies and wobbles but doesn't move from the tube's bottom.

"There's endotoxin in there, you can bet on it," he says, slipping the tube back into a rack.

Hochstein, former deputy director of product quality control (he retired last Sept. 3) in the Food and Drug Administration's Center for Biologics Evaluation and Research, is demonstrating a simple analytical test. It's one that medical professionals, drug companies, pharmacies, and others use worldwide to detect the presence of endotoxins — dangerous toxic byproducts of "gram-negative" bacteria such as Salmonella and E. coli.

The test is the limulus amebocyte lysate assay and is, Hochstein says, "remarkable" for its origin: the horseshoe crab. The limulus test, along with an osteoporosis treatment derived from salmon and a bone filler made from coral, are approved medical products that come from the sea.

Until recently, virtually all medical products had terrestrial sources. For example, organisms found in soil have yielded products such as penicillin, amoxicillin, and other antibiotic compounds responsible for saving millions of Americans from suffering and death.

Sea-based products are rare, but some experts say the world's oceans and waterways may harbor the next generation of drugs, biologics, and even a few medical devices. Dozens of promising products, including a cancer therapy made from algae and a painkiller taken from snails, are in development at research laboratories right now. Other products, such as an anti-inflammatory drug extracted from an organism called the Caribbean sea whip, are under FDA review. Three approved products already have brought the healing power of the sea successfully into the world of public health.

A Lucky Horseshoe

Along the Eastern Seaboard of the United States, it's not unusual when strolling on the shore to find horseshoe crabs that have "beached" or shed their shells. These crabs, the limulus species, are important players in the ecology and marine life of shore areas from Maine to Florida. Their importance increased when, more than two decades ago, researchers discovered that, due to some unique properties, the crabs' blood could be used to detect dangerous endotoxins in drugs, medical devices, and even water.

Endotoxins are produced when E. coli and other gram-negative bacteria break down. The effect on humans exposed to the toxins ranges from fever to hemorrhagic stroke. "This underscores the importance of the test in finding these toxins before they can do any damage," says Hochstein.

Before the limulus amebocyte lysate (LAL) test was marketed, medical professionals gauged endotoxin presence by injecting the substance being analyzed into a rabbit's ear. If the animal developed a fever, endotoxins were present. Rabbit tests still are done but are "falling out of favor," says Hochstein, because "they are just too complicated." The tests take four to five hours, and labs must keep caged rabbits on hand.

By contrast, the LAL test uses a glass tube and takes only one hour. Drawing blood from horseshoe crabs causes the animals no harm, and they can be returned to their habitat within 48 hours.

By many accounts, the discovery of the LAL test was serendipitous. In 1971, National Institutes of Health researcher Jack Levin was studying various marine animals when he discovered that blood in horseshoe crabs exposed to E. coli bacteria had clotted. He then drew fresh blood from some horseshoe crabs and exposed it to E. coli in the laboratory. The blood clotted to a gel-like consistency. Further experiments in the NIH Bureau of Biologics, which later became part of FDA, confirmed that if any endotoxins are present, the blood will clot.

Hochstein was a major participant in those early tests, and he recalls setting up shop at a NASA facility on the Eastern Shore of Virginia to catch and draw blood from 1,000 horseshoe crabs at a time. He and his colleagues also kept as many as 200 crabs in tanks filled with ocean water in labs outside Washington, D.C., to ensure an available blood supply.

The team ultimately developed a method for separating amebocytes, which are similar to human white blood cells, from the rest of the crab's blood. These cells then were spun in a centrifuge to intentionally rupture them and create a "lysate," the essence of the LAL test, which is freeze-dried and looks like grains of salt.

In 1973, FDA published regulatory guidelines for producing the LAL test, and in 1977, the agency licensed the first LAL product to Massachusetts-based Associates of Cape Cod. Five other companies have developed their own LAL products since then. Hochstein says FDA's LAL work is an excellent example of transferring technology from the public to the private sectors.

The test has a large market in drug companies that use LAL to detect endotoxin contamination in injectable products, says Melissa Juntunen, marketing coordinator for Associates of Cape Cod. "Probably every major pharmaceutical company uses it," she says. Medical device firms also use the test to ensure that catheters, pacemakers, and other invasive devices are endotoxin-free.

About the Author

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FDA is A United States government body that oversees medical devices, including contact lenses, intraocular lenses, excimer lasers and eyedrops. In the US, these products must be approved by the FDA before they can be marketed.