University of Wisconsin-Madison
Mirus Bio Corp., Madison
Transgene, Strasbourg, France
Transfer of naked DNA into muscle cells via a high-pressure delivery system through blood vessels
Estimated start date for clinical trial
In the fall of 1988, Jon Wolff moved to the University of Wisconsin at Madison, having completed training in pediatrics and then in genetics and metabolism at the University of California-San Diego, where he’d been an assistant professor for three years.
The field of gene therapy was mostly a gleam in the eyes of a few scientists, Wolff, then 32, among them. Back then, Wolff says, “People wanted to think about taking cells that were outside the body, putting genes into them, and then putting the genetically modified cells back into the body.”
But he and a few others had a different idea — transferring genes directly into the body.
Wolff started working with viruses as vectors, but he soon began experimenting with ways to deliver genes without them. He tried cationic lipids, fat particles that carry a positive electrical charge, as gene therapy vectors. These molecules had proved among the best vehicles for putting genes into cells in a lab dish.
Injecting the genes into mouse muscle cells, the researchers compared genes (DNA) packaged in the lipids to naked DNA (delivered without any vector), fully expecting to see a difference in favor of the lipids.
But they got a surprise. The naked DNA actually worked better than the lipid-encased DNA. “Nobody could have anticipated something like this would be this easy,” Wolff says. “It really seemed like something crazy.”
Crazy or not, the group published a landmark paper in the journal Science in 1990, and that’s when Wolff’s association with MDA and muscular dystrophy began.
In that 1990 paper, they reported using naked DNA, which is actually a gene spliced into a ring of DNA called a plasmid, to deliver genes to the muscles of mice. (The cationic lipids, it turns out, were a bad idea, proving toxic to the liver and lungs.)
“MDA approached me after that Science paper, and that really got me working on muscular dystrophy,” Wolff says. “Before then, I had seen patients with muscular dystrophy, but I wasn’t really thinking about it in terms of gene therapy.”
Since 1990, he’s been thinking about it steadily, first injecting naked DNA for dystrophin directly into animal muscles. More recently he’s delivered dystrophin DNA into the bloodstreams of animals, using a high-pressure (hydrodynamic) method that involves blocking off parts of the circulation at a time with a tourniquet.
Going naked gets results
Fifteen years ago, Wolff says, you were lucky to get 1 percent of the desired genes into cells in a dish. Now, you can expect to get nearly 100 percent of genes into ex vivo cells.
“So there’s been a lot of progress in cells outside the body,” Wolff says. “But the problem is designing things to work inside the body.”
Even there, things are looking up. “We got over 10 percent of muscle fibers expressing a foreign gene with one injection,” he says of a recent experiment in which genes were injected into animal blood vessels. “That’s about as good as it gets with some viral strategies.”
Wolff says nonviral delivery methods are best for avoiding an unwanted immune response that can either block the gene transfer or, worse yet, make the recipient ill.
Even though the newest viral vector for muscle tissue, AAV, seems to avoid this kind of response the first time the body sees it, the immune system may react to it with a second or later administration. Since many gene injections over time may be needed to effectively treat muscular dystrophies, that could be a problem, Wolff says.
So far, Wolff’s plasmid DNA strategy is the only one to have been tested on boys with Duchenne and Becker MDs.
In a study conducted in France and reported in 2003, nine boys with either of these diseases had naked dystrophin DNA injected directly into an arm muscle. There were no ill effects, and some dystrophin apparently was produced.
“It’s a good first step for us,” Wolff says.
He hopes to begin a clinical trial in early 2006 and says the evidence of safety from the first trial will “really help us get through the regulatory agencies, given that this part has already been done in people.”