University of Washington, Seattle
Delivering microdystrophin gene in a type 6 adeno-associated viral (AAV) vector, eventually via the bloodstream
Estimated start date for clinical trial
2006 for intramuscular gene delivery; later for intravascular gene delivery
In the late 1970s, a young Jeffrey Chamberlain began working toward his doctorate in biochemistry under professor Stephen Hauschka, a muscle biologist at the University of Washington in Seattle.
“Steve’s lab was doing a lot of basic research in muscle development and muscle gene expression in the early ’80s,” Chamberlain says, “and he was supported by MDA. He was on some of MDA’s advisory boards and would tell us about muscular dystrophy, even though we weren’t really working on that disease per se. But MDA was interested at that time in understanding as much about muscle biology as they could because they didn’t know what caused any of the muscle diseases.
“That’s what got me interested in muscular dystrophy,” says Chamberlain, now a professor of neurogenetics, biochemistry and medical genetics at the university.
Chamberlain became captivated by muscle genes and went to Baylor College of Medicine in Houston to do postdoctoral studies with Thomas Caskey, another prominent medical genetics researcher then supported by MDA.
In 1986, the gene for dystrophin, the protein missing in Duchenne muscular dystrophy, was identified by MDA-supported researchers, and the field changed completely.
“About that same time,” Chamberlain says, “it became clear that a naturally occurring mutation in a mouse that had been described a few years earlier turned out to be a dystrophin mutation.” Chamberlain embarked on studying this and other mouse versions (“models”) of Duchenne dystrophy. “I decided that’s where I wanted to go with this. Having a good mouse model, I thought, was a really good way to understand the disease.”
In 1990, Chamberlain moved to the University of Michigan in Ann Arbor to set up his first independent laboratory, and he and his colleagues began an intensive study of dystrophin.
“A lot of what I was doing at Michigan was trying to develop as much knowledge about the mdx [dystrophin-deficient] mouse as I could,” says Chamberlain, who in 1999 became director of the university’s Center for Gene Therapy.
The dystrophin gene posed many challenges. First, it’s the largest gene in nature, making it particularly unwieldy to work with. Second, it carried instructions for a structural protein, one that appeared to play a supporting role near the membrane of the muscle cell, and researchers weren’t sure whether they could get that effect from gene transfer.
Chamberlain selected the adenovirus (which causes a coldlike illness in its natural state) as a potential vector, modifying it to carry the dystrophin gene to muscle.
His team worked at the problem from “both ends” — miniaturizing the large gene so it would fit inside the virus, and at the same time hollowing out the virus so it could carry the full gene.
They also disguised most of the viral characteristics so as not to provoke an attack by the immune system. (The importance of that strategy became clear when, in 1999, a participant in a non-MDA gene therapy trial died of what was later determined to be an overwhelming immune response to the adenovirus.)
Chamberlain’s group determined which parts of the dystrophin gene could be eliminated and still leave a working protein. But making an adenoviral shell that the body’s immune system wouldn’t recognize as an unwanted invader proved challenging. When the adenovirus was fully gutted of its own genes, the immune system appeared to tolerate its presence, but the vectors were very difficult to prepare in quantities needed for clinical trials.
Many investigators, including Chamberlain, began shifting their attention to the more benign adeno-associated virus, or AAV (so named because it was discovered in association with an adenovirus infection).
But an old problem resurfaced: The AAV was too small to carry even a miniaturized version of the dystrophin gene.
It was back to the drawing board for Chamberlain, who further miniaturized the dystrophin gene, finding exactly which parts had to be retained in the lab’s new microdystrophin molecule.
“We’ve had a lot of exciting results recently with AAV vectors, particularly with the type 6 version of AAV. But we don’t want to put all our eggs in one basket, so we’re looking at other AAV types and other types of vectors. We’re not just relying on what can be found naturally in the world, but engineering features in the vectors to make them do a little more of what we would like them to do.”
The team recently achieved widespread delivery to mouse muscles by injecting a type 6 AAV vector carrying a microdystrophin gene into the tail veins of mice. At the same time they injected VEGF (vascular endothelial growth factor), which temporarily allowed the AAV vector to cross blood vessel walls and enter muscle cells.
“We’re putting a lot of effort into improving that strategy to make it simpler, safer, and something that can be achieved at lower doses,” Chamberlain says.
Chamberlain is ironing out administrative and regulatory wrinkles for a planned clinical trial of the strategy. He hopes to deliver the microdystrophin gene to the muscles of children with DMD in 2006 and to have a vascular gene therapy trial in place some time after that (delivered through the circulatory system).
“We have a lot of very good collaborators at the MDA clinic here in Seattle. We’ve actually got more than 12 physicians who have joined our Wellstone Muscular Dystrophy Cooperative Research Center and who will be part of the clinical trial team. [This center is co-funded by MDA and the National Institutes of Health.]
“My lab won’t conduct the actual trials, since I’m not a physician,” Chamberlain says. “However, we will develop the system in preclinical studies and generate a microdystrophin vector that can be tested in the clinic.”