First mouse model could be used to test therapies
Davide Gabellini, an MDA grantee, was on a research team at the University of Massachusetts Medical School in Worcester that has found a new piece of the puzzle of facioscapulohumeral muscular dystrophy (FSHD) by identifying specific consequences of a previously known mutation that removes a section of DNA. The scientists also developed the first reliable mouse model of FSHD.
In 2002, Gabellini and colleagues found that a missing section of DNA on chromosome 4, called D4Z4, normally acts as a suppressor of other genes near the D4Z4 region. (See “Research Updates,” August 2002.) Since then, a search for genes in the region that, if activated instead of suppressed, might cause muscle degeneration, has turned up several candidates, but nothing definitive.
Now, scientists from the University of Massachusetts, including Gabellini, and from Milan and Pavia, Italy, who published their findings online Dec. 11 in Nature, have determined that a gene known as FRG1 (for FSHD region gene 1) is a major suspect in causing FSHD.
When they analyzed genetically altered mice that produced various levels of the FRG1 protein, they found that the higher the FRG1 levels were, the worse the MD symptoms were in the mouse. Mice with elevated FRG1 levels showed spinal curvatures, muscle wasting (atrophy), increased connective tissue in the muscles, and reduced exercise tolerance, along with muscles that appeared dystrophic under the microscope.
The muscles most affected were somewhat analogous to those affected in humans with FSHD, although differences in mouse and human anatomy in the face and upper body make direct comparisons impossible.
Further experiments showed that FRG1 produces its adverse effects indirectly, through at least two other proteins: TNNT3, which normally regulates muscles’ ability to contract, and MTMR1, which can regulate muscle atrophy.
The researchers think FRG1 is one of many genes that affect RNA splicing, a process that determines the final composition of a protein. They say the effects of high levels of FRG1 could range far beyond TNNT3 and MTMR1, possibly affecting hundreds or even thousands of other genes.
“It is likely that the cumulative effect of decreased levels of many normal protein [forms] and increased levels of many aberrant protein [forms] is responsible for disease,” they write.
Gabellini says, however, that suppressing or blocking FRG1 might be all that’s needed to remedy the effects of FSHD. He also noted that FRG1 mice could be used to test potential therapies for FSHD.
In experiments with mice, MDA grantees at Stanford (Calif.) University have achieved excellent production and distribution of dystrophin, the muscle protein needed but missing in Duchenne muscular dystrophy (DMD).
The scientists used a new technique in which genes are injected into the muscles without being carried in a virus, but with a second gene that coaxes the new genes to integrate into an existing chromosome.
Thomas Rando, an MDA-supported molecular biologist and associate professor, and Carmen Bertoni, an MDA-supported postdoctoral student working with Rando, injected the leg muscles of DMD-affected mice with dystrophin genes. They also injected genes for integrase, a protein that causes genetic material to integrate into a chromosome instead of remaining separate from the chromosomes in the cell nucleus.
The research team, whose results are in the Jan. 10 issue of Proceedings of the National Academy of Sciences, also included investigators from Stanford’s Genetics Department and the Veterans Affairs Palo Alto Health Care System.
The team found that the mouse muscles receiving both dystrophin and integrase genes had more than eight times the amount of dystrophin six months after injection than did mouse muscles that received dystrophin genes alone.
Moreover, while the muscle fibers that received dystrophin genes alone showed dystrophin production centered around the injection site, fibers that got both proteins produced dystrophin along the whole length of the fiber. That result is thought to give the fiber much better protection against damage.
The muscle fibers that received the combined gene transfer strategy were less permeable to an injected dye than dystrophin-damaged fibers, also a sign of superior protection.
“The issue of the distribution of dystrophin from end to end of the fiber is an important factor that really pertains to any form of muscle gene delivery. You need to consider not just the amount of the protein produced, but where it is,” Rando said.
Acknowledging that the possibility of using an integrating gene is a “step forward for virus-free gene therapy,” Rando said it also carries an inherent risk, because gene integration in other situations has led to genetic mutations.
“The development of this technology will involve producing more specific integrases with the hope of targeting integration to one place. Knowing where the gene is, knowing it’s safe, and getting high levels of protein production are the ultimate goals.”
A new compound that blocks myostatin, a natural inhibitor of muscle growth, has increased muscle mass in mice by up to 60 percent in two weeks, a team of scientists announced in the Dec. 13 issue of Proceedings of the National Academy of Sciences. The tested mice didn’t have muscular dystrophy.
Se-Jin Lee at Johns Hopkins University in Baltimore, with colleagues from several academic institutions and biotechnology companies, says the compound blocks myostatin by providing it with a portion of a molecule that it normally sticks to.
Known as ACVR2B, the new compound provides myostatin with a partial molecule that keeps it from interacting with its normal molecular binding partner. Without this interaction, myostatin can’t send its usual growth-inhibiting signals to muscle cells.
A previously developed myostatin blocker is now being tested in clinical trials in people with certain adult forms of muscular dystrophy. That compound, MYO-029, was developed by Wyeth Pharmaceuticals, and is based on an antibody (immune system protein) that sticks to and interferes with the myostatin protein.
Lee, a professor of molecular biology and genetics at the Johns Hopkins Institute for Basic Biomedical Sciences, has MDA funding for work on myostatin mechanisms. He says the new inhibitor is very potent and leads to dramatic effects in the mice. These effects were “larger and faster than we’ve seen with any other agent and even larger than we expected.”
He cautions, however, that the effects of ACVR2B can be attributed to its ability to block more than just myostatin signaling, which may increase the potential for side effects.
Lee also notes that increasing muscle mass alone isn’t necessarily the answer in muscular dystrophy.
“In general, I am quite optimistic that targeting this pathway will turn out to be an effective way to increase muscle growth,” he says. “But much more work will be required to determine whether this will be a viable approach.”
A slight variation in an immune system protein called PTPN22 may cause susceptibility to autoimmune diseases, disorders in which the immune system mistakenly mounts an attack on the body’s own tissues, a new study says. (Autoimmune neuromuscular diseases include myasthenia gravis, Lambert-Eaton syndrome, polymyositis and dermatomyositis.)
In the protein, the substitution of the amino acid tryptophan where most people have arginine appears to disrupt some of the fine-tuning needed for a safe and effective immune response, according to findings in the December issue of Nature Genetics. The altered protein results from a variation in the gene for PTPN22.
Torkel Vang, at the Burnham Institute in La Jolla, Calif., and colleagues, say the altered PTPN22 may interfere with the necessary dampening of an immune response normally performed by regulatory immune system cells, or with the normal destruction of self-reactive cells.
They suggest that a small molecule that blocks this alteration could be developed and potentially be used as a treatment for a variety of autoimmune diseases.
At a January meeting of the European Neuromuscular Centre, professionals from the United States and five European countries learned that type 2 myotonic dystrophy (MMD2) may be much more common than previously believed, at least in some areas. In Central Finland, the estimated prevalence is about one person in 10,000.
MMD2, which results from an expanded, repeated section of DNA on chromosome 3, is similar to type 1, which results from a similar DNA expansion on chromosome 19. But the type 2 form has been considered a rare disease, while the type 1 form, which occurs in about one in 7,500 births, is relatively common.
Unlike type 1, type 2 MMD doesn’t seem to affect newborns, doesn’t cause serious cognitive problems and affects thigh muscles early.
MDA hosts a chat for families with MMD every Wednesday evening. Visit www.mda.org and select MDA Chat. Online groups focused solely on MMD2 can be found on Yahoo! Groups (enter ”myotonic dystrophy 2” in the search box).
MDA grantee Howard Worman at Columbia University in New York, a leading physician-scientist in the field of lamin proteins, recently announced that his group has uncovered the molecular connections between at least one lamin gene mutation and heart disease. Mutations of lamins underlie Emery-Dreifuss muscular dystrophy (EDMD) and type 1B limb-girdle MD (LGMD1B).
The group’s findings were presented at a December meeting of the American Society for Cell Biology in San Francisco. It seems that type A lamins, which are produced in almost all cells in the body, may, when flawed, have specific deleterious effects on the heart.
The researchers analyzed cardiac muscle tissue from mice carrying a lamin A gene mutation that causes the chromosome 1 type of human EDMD, a disorder in which heart disease is nearly universal.
Unlike normal mouse hearts, those with the mutated lamin genes showed increased activity of genes for other proteins, called MAP kinases, which have been implicated in heart enlargement and heart failure. The investigators also found changes in gene activity for three other components that can contribute to cardiac muscle disease.
Worman said, “The finding that MAP kinases are activated in the heart in this mouse model of Emery-Dreifuss muscular dystrophy suggests that inhibitors of these enzymes may be useful as treatments. This of course remains to be tested in animal models and, if successful, possibly in human subjects.”