Ever since Louis M. Kunkel discovered the cause of Duchenne muscular dystrophy (DMD) in 1986, he has been laboring to find a cure for this muscle-wasting disease. DMD—the result of an error in a single gene—attacks 1 out of every 4,000 newborn boys, progressively crippling and then killing them at an early age.

Kunkel saw that patients with DMD lacked a protein, dystrophin, which this gene would have produced if it were functioning normally. So he knew he had to replace the protein somehow. He and others tried many methods—gene therapy to deliver a normal gene to the defective muscle cell, drugs to help restore the missing protein, and cell therapy to inject normal cells into muscle or blood—but despite some partial successes in animals, nothing really worked.

Kunkel's lab worked mostly with mdx mice, a naturally mutant strain that lack dystrophin. When he and his colleagues attempted to cure these crippled mice with injections of muscle stem cells from normal mice, "some of the donor cells did go into the damaged muscles," he recalls, "but we never got more than 1 to 2 percent of the muscles repaired. Part of the problem was that when you inject cells into a mouse's tail vein, which is the most accessible part of its circulation, the donor cells go through all the organs—the lungs, liver, heart, and so on—and out through the arterial system. Most of the cells get filtered and lost, and don't contribute to the therapy."

Today, however, Kunkel feels he is on the verge of success. The big breakthrough came last summer when a team of Italian scientists headed by Giulio Cossu of Milan's Stem Cell Research Institute announced it had found a new route for the injection of stem cells into dystrophic mice directly into an artery. The cells seemed to lodge within the capillary system near the injection site. From there, about 30 percent of them migrated to the diseased muscles. "Not only did the cells get there," he says, "but at later time points, you could see a larger number of donor cells than at the earliest point, as if they were trying to divide."

"Can we improve on this?" asks Kunkel with a glint in his eye. "If we can get the stem cells into 50 percent of the dystrophic muscles, that's basically a cure."

They had trouble at first because "the mouse artery was 10 times smaller than our smallest injection needles—it was like trying to hit it with a hammer!" Kunkel says. "Though a tail vein is even smaller than an artery, it can be hit much more easily because it is right under the surface of the skin and can be made to swell up by warming it. In the new system, the mouse had to be anesthetized and opened up to expose its artery, which was lifted out—a complex procedure.

"It wasn't until we started collaborating with some vascular surgeons who had been doing heart transplants in mice that we were able to get the stem cells into the mouse arteries efficiently," he says. In humans, of course, reaching an artery would not be a problem given that human arteries are so much larger.

Getting the stem cells into the muscles was just the first step. Unless these cells supplied enough dystrophin, the diseased muscles would not be repaired. So Kunkel also tried to find different stem cells that could do the job more effectively. In 1999 his lab and that of his colleague Richard Mulligan announced they could restore some of the missing dystrophin in mdx mice with the aid of a new kind of stem cells called "side population" (SP) cells, which seemed to work much better. These SP cells had to be taken from muscle tissue, however. Last year Kunkel's lab succeeded in deriving similar SP cells from adult skin, which is easier to obtain. Since they originate in adult tissue, both kinds of SP cells will be much less controversial than embryonic stem cells.

"It's my belief that you can do a lot of therapeutic intervention with adult-derived cells," says Kunkel. He notes that the new stem cells seem ready to differentiate into every type of muscle tissue, which implies that they have the potential to treat many forms of muscle disease.

The combination of a new cell type and a new delivery system "may revolutionize how one does therapy for muscle diseases," Kunkel suggests. "When we get it perfected in mice, we'll go to humans." He thinks this might happen "in a couple of years."

—Maya Pines

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Reprinted from the HHMI Bulletin,
Spring 2004, pages 12-17.
©2004 Howard Hughes Medical Institute