DIABETES: THE SPACE CONNECTION

Astronaut and protein crystallographer Larry DeLucas, PhD, has launched several experimental insulins into orbit. These include Humalog, the long-awaited, recently approved, short-acting insulin lispro. Outer space has provided him with a laboratory environment unavailable on Earth. In the gravity-free environment of space, DeLucas can grow flawless crystals of complex proteins. Such crystals are easier for crystallographers to examine, and they provide more accurate "blueprints" for pharmaceutical manufacturers.

It may seem odd to think of insulin as a crystal. DeLucas points out, however, that the cloudiness in vials of NPH and ultralente insulin is nothing more than undissolved insulin crystals. Regular insulin is clear because the insulin is already dissolved--and thus faster acting. Some animals--rats, for instance, actually have insulin crystals undissolved in their pancreas.

For more than four decades, scientists have experimented with new forms of insulin. Most of these variants, or analogs, offer insulin users different durations of action. Insulin lispro, for instance, begins acting within minutes after injection to control the rise of blood glucose immediately after meals. On the other hand, researchers are developing an extra-long acting variety of insulin. Such an insulin analog might deliver a basal dose lasting 2 days or more, instead of the maximum 28-hour cover now available with ultralente.

Eli Lilly's Humalog was approved for sales in the U.S. market last June. A long-acting analog, however, is on the more distant horizon--literally.

Why Grow Crystals in Space?

In 1992, DeLucas donned a spacesuit himself as a payload specialist for the National Aeronautics and Space Agency (NASA). He wanted to learn the rigors and practical realities of research in a weightless environment. Now that he has designed a successful crystal-growth system, he prepares experiments on Earth and ships them to space aboard space shuttles, where other astronauts tend the projects.

"Crystals produced on Earth are usually flawed because of gravity-induced movements within the liquid, crystal-growing medium," explains DeLucas. "But in space, the liquid is motionless, which greatly slows the growth of some protein crystals. The more slowly you grow crystals, the fewer the flaws in their structure."

The atoms and molecules of space-grown crystals are aligned more perfectly. This allows their structures to be seen at higher resolutions.

DeLucas, who studies crystals at the Center for Macromolecular Crystallography at the University of Alabama at Birmingham, said he could not have designed his system without his actual space trip in 1992. "I could see what changes I needed to make," he said. "So many simple things happen differently in space, like stirring a drop of liquid with a syringe tip -- the drop of liquid wouldn't stay in position in the container. Instead the drop of liquid stuck to the syringe tip, moving with the tip as I used it to try to mix the liquid drop. Another nuisance was bubbles in the syringe. To get rid of them, I had to create artificial gravity -- using my arm to swing the syringe around in a bag."

DeLucas said that he has worked with insulin for years as a "practice" crystal to develop his techniques and equipment for crystal-growing in space. Insulin stopped being just for practice, however, when DeLucas began collaborating with Dr. G. David Smith. Smith, at the Hauptman-Woodward Medical Research Institute in Buffalo, New York, has worked extensively on the insulin lispro analog with Hauptman-Woodward colleague Dr. Ewa Ciszak, PhD. They grew insulin lispro crystals on two space flights during 1995. Smith published the structure of insulin lispro recently and has answered some of the difficult questions explaining its fast-acting behavior.

Smith spent 14 years studying insulin structures with the goal of developing an ultra-slow-dissolving insulin. Smith had experimentally altered ordinary insulin by adding a small "guest molecule." This, in turn, "buries" the insulin's zinc ions, which play an important role in stabilizing the insulin. Isolating the zinc makes insulin harder to dissolve. Smith tried two guest molecules-- Tylenol and p-hydroxybenzamide (BZN). Unfortunately, the Tylenol made the insulin slightly toxic. While Earth-grown BZN-altered crystals were adequate to determine the insulin structures, it was hoped that the space-grown crystals of BZN could be studied more closely.

Surprises From Space

After careful review to choose the most promising experiments, a BZN-insulin growing chamber was sent into orbit on the NASA-sponsored Space Habitat mission number STS60, in 1994. Back on Earth, the crystal surprised Smith, even after 10 years of insulin research. To his amazement, the BZN additive had actually attached itself in pairs to an insulin molecule.

"This was the first time something like this had ever been observed," Smith noted. "In hindsight, there were indications from the Earth-grown crystals, but we might have missed the paired structure without the space-grown crystals." Smith said that the BZN-insulin is unlikely to result in a new product at this time. However, he says that he and DeLucas will probably be asked to grow other insulin crystals in upcoming space flights.

DeLucas has grown several other diabetes-related proteins in space. Space-grown variants of aldose reductase crystals have also been produced. Aldose reductase, an enzyme, has been implicated as a cause of various diabetes complications. By building perfect, space-grown crystals of aldose reductase bound to different inhibitors, DeLucas says scientists hope to design a matching compound to more effectively block the enzyme's activity. Previous attempts to design such a drug have not been completely successful. The precision of space-grown crystals offers another chance of finding a medication that actually works.

(Note: This article appeared in DIABETES DATELINE, Fall 1996, published by the National Diabetes Information Clearinghouse, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health.)