LONDON ¿ While many attempts at gene therapy involve replacing a defective gene with a similar but healthy one, researchers in Switzerland have shown that a completely different gene may work just as well ¿ so long as it has the same effect. Their approach, demonstrated in a mouse model of muscular dystrophy, could work for other inherited diseases, they suggest.
Markus Ruegg, associate professor for neurobiology at the University of Basel in Switzerland, told BioWorld International: ¿Our study shows that one can use a protein that is not necessarily homologous to that normally made by a disease gene, to replace a disease gene. The protein we used to rescue these mice, which normally exhibit symptoms similar to muscular dystrophy, was not the protein that you would normally think of using to treat muscular dystrophy.¿
Ruegg is a member of the management team of MyoContract Pharmaceutical Research Ltd. in Basel, which is committed to developing drugs for the treatment of neuromuscular diseases including muscular dystrophies. Together with colleagues in Switzerland, the U.S. and Germany, Ruegg reports his latest findings in a paper in Nature titled ¿An agrin minigene rescues dystrophic symptoms in a mouse model for congenital muscular dystrophy.¿
Most muscular dystrophies are caused by a mutation in one of the genes encoding the proteins that form the dystrophin-glycoprotein complex. This complex binds to the actin cytoskeleton of the muscle fibers and plays a vital role in allowing the muscle fibers to contract when given the signal to do so.
In congenital muscular dystrophy (CMD), a mutation is present in the gene encoding one of the chains of laminin, a protein that forms part of the dystrophin-glycoprotein complex. Congenital muscular dystrophy is very rare, affecting one or two in every 100,000 births in Europe. Those affected begin to develop symptoms soon after birth, and may never stand or begin to walk. Death in early childhood is usually due to respiratory failure.
Ruegg and his team had been studying the formation of nerve-muscle synapses, including the influence of a protein called agrin on the formation of neurotransmitter receptors on the postsynaptic membrane. Ruegg said, ¿When we identified the binding partners of agrin, we realized that it was just like laminin, in that it bound both sarcolemma ¿ the muscle membrane ¿ and extracellular matrix. This gave us the idea of using it to treat the symptoms of congenital muscular dystrophy, in which laminin is missing.¿
They wanted to test the idea that the binding domains of agrin that they had identified in their earlier experiments could substitute for those of laminin in the dystrophin-glycoprotein complex. So they took DNA encoding these domains and used it to form an agrin ¿minigene,¿ together with a promoter that would ensure that the DNA was expressed in muscle. They then engineered a mouse in which this transgene was expressed.
A mouse model of CMD was already available. These mice are smaller, thinner and more passive than wild-type mice. They typically avoid walking and die at an average age of 8 weeks.
When Ruegg and his colleagues crossed the transgenic mice they had made with the CMD mice, some of the offspring expressed the transgene. ¿When we compared these animals in terms of their muscle structure and their behavior to those that did not express the transgene, we found that the transgene had a beneficial effect on the disease phenotype,¿ Ruegg said. The animals expressing the agrin minigene, for example, survived for about 40 weeks, much longer than the CMD mice. Nor did they avoid walking.
While these encouraging results point toward the possible use of gene therapy in patients with CMD, Ruegg emphasized that clinical applications of this work in mice are ¿many years down the road.¿ One advantage of using a truncated agrin gene, he explained, would be to avoid the severe immune response that would be expected if the laminin gene were to be expressed in people who had never previously made this protein. Its small size should also mean that it is easier to insert into viral vectors for gene therapy, he added.
Nor does the study in mice replicate the exact situation of gene therapy in people with CMD. ¿When you want to treat patients,¿ Ruegg said, ¿they are already ill. But in our mice, the transgene was expressed throughout development and before they began to show symptoms. Next, we want to generate mice that will express the transgene only after they develop the disease, and compare their disease progression with those of CMD mice. This will allow us to find out if the transgene can slow the development of the disease or restore muscle function.¿
It may also be possible, he added, to identify drugs that could upregulate levels of agrin in the body, in order to treat CMD.