Pompes Disease Trials Expanded to 3 More U.S. Centers

With encouraging results from small pilot trials, the biotech company Genzyme is recruiting participants for a large, multicenter trial to further evaluate an experimental treatment for Pompes disease, also known as acid maltase deficiency.

The disease is caused by genetic defects in the enzyme acid maltase, also called acid alpha-glucosidase (GAA). The enzyme normally breaks down glycogen (stored sugar) in muscle cells, and without it, glycogen accumulates in cellular compartments called lysosomes. The most severe form of the disease causes cardiac and skeletal muscle weakness in infancy, and is often fatal within the first year of life.

In two pilot trials, 11 babies with Pompes have received enzyme replacement therapy, and some are now healthy, walking toddlers (see "Pompes Disease," March-April). The therapy involves intravenous infusions of GAA.

In March, Genzyme, of Cambridge, Mass., announced that it was launching a third, larger trial, to include up to 16 babies between ages 6 months and 3 years. That trial is recruiting participants through Duke University in Durham, N.C., the University of Florida College of Medicine in Gainesville, Childrens Hospital Medical Center in Cincinnati, and two sites in England and France (For more information, visit www.mda.org/research/ctrials.aspx.)

MDA has offered to provide up to $50,000 to each U.S. center in order to help trial participants cover expenses for physical, speech and respiratory therapy, social services, and nutritional consultations. Most participants will need these services during the several trial period and are likely to be outside of their health insurance coverage areas.

In February, MDA Director of Research Development Sharon Hesterlee met with Genzyme officials to lay the groundwork for a natural history study of late-onset Pompes disease, which causes skeletal muscle weakness but no cardiac problems. This study would be a step toward designing trials of enzyme replacement therapy in children and adults with Pompes.

Andy Amalfitano

Meanwhile, MDA grantee Andy Amalfitano at Duke is making progress toward gene therapy for the disease. Injections of the GAA gene might be required just once or twice a year, compared to the two to eight times per month now required for injections of the GAA protein, he says.

In experiments on mice with Pompes, he showed that a virus carrying the GAA gene can be targeted to the liver, which then releases the protein into the bloodstream, restoring GAA activity to muscles for up to six months.

His latest study, published online by the Journal of Gene Medicine in December, showed that the same approach could be used to restore GAA activity for at least 28 days in the muscles of quail with Pompes. The quail model provided validation for the enzyme replacement therapy.

New Tests Detect More Dystrophin Gene Flaws

The large size and complexity of the dystrophin gene, which carries instructions for a muscle protein and is flawed in Duchenne and Becker muscular dystrophies (DMD and BMD), has made dystrophin genetic testing particularly challenging.

Now, MDA-supported researchers have developed tests that can detect a much higher percentage of dystrophin mutations, including some very minor DNA changes.

Most commercially available tests can detect only the type of mutation in which large pieces of the gene are absent. This "deletion analysis" testing is positive for only about 60 percent of people with DMD.

Knowing the precise genetic flaw in a person with DMD can help a family predict the risk of passing the disease to children, qualify for certain research studies, and eventually perhaps to predict the severity of the disease or suggest specific treatments.

Carolyn Sue Richards, a molecular biologist and MDA grantee at Baylor College of Medicine in Houston, is working on a type of dystrophin gene analysis based on the "denaturing high performance liquid chromatography" (DHPLC) technique.

Richards group wants to test the technique on at least 100 people with a clinical (symptom-based) diagnosis of DMD or BMD, a muscle biopsy result suggesting these conditions, and negative results on a dystrophin gene deletion test.

For information on getting into this study, which includes test results for the family and doesnt require travel to Houston, contact Madhuri Hegde or Patricia Ward at the Baylor Diagnostic Sequencing Laboratory at (800) 411-GENE, (713) 798-5722 or dsl@bcm. tmc.edu.

Kevin Flanigan, an MDA grantee at the Eccles Institute of Human Genetics at the University of Utah in Salt Lake City, was among those who recently developed another approach to improved dystrophin gene testing. MDA clinic co-director Jerry Mendell, at Ohio State University in Columbus, provided patient DNA samples used to evaluate the new, more accurate procedures.

	Kevin Flanigan

The researchers published an article in the April issue of the American Journal of Human Genetics about their technique, called "single condition amplification/internal primer," or SCAIP, sequencing.

The test is expected to become available for less than $1,000 this spring through the Human Genome Depot (www.genome.utah.edu) at the University of Utah.

For information, call (801) 587-9540 or e-mail kevin.flanigan@genetics.utah.edu.

Complete dystrophin gene analysis using DOVAM, for "detection of virtually all mutations," became available about two years ago, with MDA support, through the City of Hope National Medical Center in Duarte, Calif.

You can get more information at www.cityofhope.org/cmdl, by calling (888) 826-4362, or by e-mail to cmdl@coh.org. The cost ranges from $900 to about $1,600.

Better Understanding, Testing in Type 2 MMD

MDA-supported researchers Laura Ranum and John Day at the University of Minnesota in Minneapolis have developed a DNA test for myotonic dystrophy (MMD) type 2.

Ranum, Day and colleagues published their results in the Feb. 25 issue of Neurology, along with a thorough description of the disease.

Laura Ranum and John Day

MMD2 seems to lack the severe, congenital form of the disorder often seen in type 1 MMD, and it doesnt seem to worsen with each generation, as does MMD1. Type 2 tends to cause more weakness in the proximal (close to the trunk) muscles, while type 1 causes more distal (far from the trunk) muscle weakness.

In many other respects, the two diseases are similar.

The genetic test is available through Athena Diagnostics of Worcester, Mass. See www.athenadiagnostics.com, or call (800) 394-4493 for details.

Destroying Excess Genetic Material New Lead in MMD1

MDA-supported researcher Jack Puymirat of the Human Genetics Unit of Laval University in Quebec is part of a team thats on the trail of a new treatment strategy for type 1 myotonic dystrophy (MMD1), a multisystem disease that leads to muscle weakness and can also involve heart and gastrointestinal problems and central nervous system dysfunction.

	Jack Puymirat

Using a new technique called antisense RNA, the researchers were able to destroy at least in cells in a lab dish the abnormally long sequence of genetic material that causes MMD1. In the disease, these long sequences are found in the DNA on chromosome 19. These are converted by the cells to correspondingly long strands of RNA, which, many experts say, may be the root cause of the most serious trouble in MMD1.

The researchers found that the antisense construct blocked the abnormally long RNA. The treated MMD-affected cells then morphed from immature, separated muscle cells to mature, fused muscle fibers in a normal fashion; and they reacted normally to insulin and sugar. The investigators say these activities normally wouldnt be carried out correctly in cells with the MMD defect.

"Our data support a future role for antisense as a new gene therapy for myotonic dystrophy," the authors say in the April 22 issue of Gene Therapy.

Antisense could theoretically be used to treat other disorders in which excess DNA or RNA, rather than missing DNA, plays a role. Among other MDA-supported diseases in this category are MMD2 and oculopharyngeal MD.

New Findings Expected to Improve CMT Diagnosis

Researchers have linked two new genes to Charcot-Marie-Tooth disease (CMT).

There are over a dozen forms of CMT, many of them traceable to genes that encode specific components of nerve fibers or myelin, the protective covering around nerve fibers. The new genes, which bring the total number of identified CMT genes to 14, appear to have more general functions.

An MDA-funded team led by Vincent Timmerman of the University of Antwerp in Belgium has found the culprit gene behind CMT type 2B, which causes such a severe loss of sensation that wounds to the feet often go unnoticed and rapidly become infected. The gene, RAB7, encodes a protein that controls the transport of nutrients and other cargo within cells; its not clear why RAB7 defects specifically cause damage to nerve cells.

The same mystery holds for the second new CMT gene: glycyl tRNA synthetase (GARS), which is part of the machinery that all cells use to make proteins. A team led by Eric Green of the National Human Genome Research Institute in Bethesda, Md., found GARS mutations in families with CMT type 2D and the clinically similar disease spinal muscular atrophy type 5, both of which cause pronounced weakness in the upper extremities.

Kumaraswamy Sivakumar, co-director of MDAs clinic at St. Josephs Hospital in Phoenix, was part of the CMT2D study, which appears in the May issue of the American Journal of Human Genetics. Timmermans study appeared in the journals April issue.

If current trends continue, each newly discovered CMT gene could lead to better diagnostic tests for the disease.

Athena Diagnostics (www.athenadiagnostics.com), a company based in Worcester, Mass., offers a panel of tests for the six genes most often responsible for CMT, including periaxin, linked to the disease in early 2001.

In January, MDA grantees Valerie Street and Phillip Chance of the University of Washington in Seattle reported that mutations in the LITAF gene cause CMT type 1C, and now theyre working toward a genetic test (see "Research Updates," March-April). Theyve put out a call for neurologists to help them identify patients who might have CMT1C.

New Clues Found to SBMA Mechanism, Treatment

A study in the Jan. 23 issue of Nature confirms that the accumulation of mutant proteins is a key part of neurological diseases like spinal-bulbar muscular atrophy (SBMA). It also describes a compound that can block the accumulation and thus might make a good drug candidate for treating these diseases.

In SBMA, nerve cells in the spinal cord and in the bulbar (bulblike) part of the brainstem degenerate, causing the muscles connected to them to grow weak and waste away.

The disease is caused by a genetic defect in the androgen receptor, a protein that allows cells to respond to testosterone and other masculinizing hormones. In SBMA, the receptor contains an abnormal structure called expanded polyglutamine, which causes it to stick to itself and other proteins, creating clumps of debris called aggregates.

At least eight other neurodegenerative diseases, including Huntingtons, are caused by similar genetic defects in other proteins. In all of these diseases, expanded polyglutamine leads to the formation of aggregates, but researchers have debated whether the aggregates contribute to nerve cell death or just reflect a dying cells inability to clear away debris.

Junying Yuan and colleagues at Harvard Medical School in Boston investigated that question by experimenting with Congo red, a dye that attaches itself to aggregated proteins and is often used to examine postmortem brain tissue.

In experiments on cells containing expanded polyglutamine, they showed that Congo red could reduce the formation of aggregates, dissolve pre-existing aggregates and protect against cell death. Those results provide strong evidence that aggregates cause cell death in Huntingtons, SBMA and related diseases, Yuan says.

Other results show that compounds like Congo red might be used to treat these diseases. When Yuan and his group gave Congo red to mice with Huntingtons disease, the mice performed better on movement tests and lived longer than untreated mice.

Nonviral Gene Transfer Uses Sound Waves, Gas Bubbles

A method of gene delivery that uses microscopic gas bubbles and high-frequency sound waves (ultrasound) to help propel DNA into cells may prove safe and effective, investigators say.

Viruses have been the preferred delivery method for gene transfer, but they have safety risks. Inserting DNA without viruses ("naked" DNA) has so far proved less efficient than viral delivery.

MDA grantee Terry Partridge, in the Department of Muscle Cell Biology at Hammersmith Hospital in London, is part of a group of researchers who recently published two papers, in the mid-January and early March issues of Gene Therapy, on the "microbubble" with ultrasound gene transfer technique.

The microscopic bubbles are made of octa-fluoropropane gas coated with albumin protein. As a product called Optison, the bubbles are used to light up tissues for medical imaging. Ultrasound, used for viewing organs and tissues, is generally considered safer than X-rays.

The new technique was apparently safe and effective in mice that received injections into their leg muscles. The microbubbles seem to have had a protective effect against the muscle damage often seen with gene transfer injections.

In their March paper, the authors say, "Whatever the mechanisms involved, the protective effect of the microbubbles is a most exciting prospect for further exploitation in any gene delivery system where tissue damage is a concern."

MDA grantee George Karpati at Montreal Neurological Institute has also been studying ultrasound-enhanced delivery of DNA. MDA grantee Jon Wolff at the University of Wisconsin is experimenting with various techniques to deliver naked DNA.

Early Prednisone May Preserve Strength in DMD

A trial of alternate-day administration of the anti-inflammatory drug prednisone in very young children with Duchenne muscular dystrophy (DMD) was recently conducted in Bologna, Italy, leading researchers to conclude that such early prednisone may be justified in this disease.

Five boys with DMD between ages 2 and 4 began taking 1.25 milligrams of prednisone per kilogram on alternate days, while three boys of the same age with DMD didnt take the drug.

At the end of the study, which is in the February issue of Muscle & Nerve, the treated boys had taken prednisone for an average of four years and seven months. All were still able to get up from the floor, while two of the boys who hadnt taken the drug had lost this ability.

Children in the prednisone group also had higher muscle strength scores. They didnt show significantly more bone loss than those in the nonprednisone group.

Luciano Merlini at the Istituto Ortopedici Rizzoli in Bologna and colleagues conclude that alternate-day prednisone maintained the ability to rise from the floor in patients with DMD started on treatment before age 4 and that side effects werent worse than those observed in children treated later in the disease.

They note, "As long-term steroid treatment is effective in prolonging function but not in recovering lost function, its early use seems appropriate."

Creatine Could Help in Type 2 MMD

A German study suggests that the dietary supplement creatine may help in type 2 myotonic dystrophy (MMD2), at least with perceived strength and endurance.

Creatine may help muscle cells produce energy more easily.

In the study, 10 people with MMD2 received 10 grams of creatine per day for three months, while 10 others got a similar-looking placebo.

Christiane Schneider-Gold of the University of Wurzburg in Germany, who published these results in the Feb. 11 issue of Neurology, found no significant improvement in the creatine group on objective tests of muscle strength.

However, seven people who got creatine and one person in the placebo group reported perceived improvement of strength. The drug may also have contributed to muscle pain relief. There were no severe side effects.

The investigators suggest that a larger trial is warranted.

Service Dog Benefits Under Study

Diane Collins, occupational therapist and graduate student at the University of Pittsburgh (Pa.), is conducting a phone and mail survey to determine whether service dogs enhance functional independence, promote psychosocial well-being, or improve socioeconomic status for people with disabilities. Collins will compare the status of people who have service dogs with that of people who either arent seeking a service dog or who are waiting for one.

For more information, contact Diane Collins at dmcst84@pitt.edu or (412) 365-4544, or Shirley Fitzgerald at (412) 365-4840.