Research Updates

• Babies With Pompe's Still Doing Well
• Sifting Through SBMA Stickiness
• New Gene Therapy Strategy Found in DMD
• Oxandrolone Has No Benefit in DMD
• MG Mystery Begins to Clear
• Gentamicin Studies Yield Confusing Results in MDs
• Narcolepsy Drug Promotes Alertness in MMD
• Mouse Model Created for Nemaline Myopathy
• Disability in CMT Type 1 Underestimated, Report Says

Babies With Pompe's Still Doing Well

Three babies with Pompe's disease — a disorder that usually causes cardiac failure within the first year of life — continue to remain heart-strong, thanks to treatment they're receiving as part of an ongoing clinical trial started last fall (see Quest, "Research Updates," vol. 7, no. 6).

Pompe's disease, also known as infantile glycogen storage disease type 2 (GSD-II), or acid maltase deficiency, results from a genetic deficiency of the acid maltase enzyme, a protein that normally breaks down glycogen (sugar stored in cells). In the absence of acid maltase, glycogen accumulates to toxic levels in cardiac and skeletal muscles, causing the muscles to waste away.

The three babies are being given twice-weekly injections of a genetically engineered acid maltase protein called Pompase, developed by MDA grantee Yuan-Tsong Chen of Duke University Medical Center in Durham, N.C., and the biotechnology company Genzyme of Cambridge, Mass.

In the March/April issue of Genetics in Medicine, Chen and Genzyme scientists announced that the treatment had produced a lasting improvement of cardiac function in all three babies. Without treatment, the babies weren't expected to survive beyond 1 year of age, but they've now reached 23, 25 and 29 months.

Yuan-Tsong Chen

Several weeks into the clinical trial, all three babies showed some improvement in skeletal muscle function, but two of them soon declined to pre-treatment levels. In those two babies, an immune response against the injected enzyme apparently inhibited its effects on skeletal muscle.

The youngest child showed no immune response to the therapy, probably because he had trace amounts of his own acid maltase. At the start of the trial, his motor skills were severely underdeveloped, but he's now walking independently, according to the Genetics in Medicine report.

The child's overall improvement suggests that treatment with Pompase might similarly restore normal cardiac and skeletal muscle function to anyone with trace amounts of acid maltase, says the report. About half of babies with Pompe's, and all children and adults with later-onset forms of GSD-II, have these trace amounts, the report adds.

The biotechnology company Novazyme has developed its own version of acid maltase, called NZ-1001, and plans to begin testing it in clinical trials sometime this year.

New Gene Therapy Strategy Found in DMD

Researchers in Melbourne, Australia, are working on a new strategy to repair the genetic defects that cause Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD).

The strategy relies on a phenomenon known as homologous recombination, a natural process that's constantly breaking and rejoining the strands of DNA that carry our genes.

Years ago, scientists figured out that if they could get DNA into a cell, homologous recombination would sometimes make the cell replace its own DNA with the foreign DNA. Now, scientists Robert Kapsa and Andrew Kornberg of the Melbourne Neuro-muscular Research Institute have recognized that technique's potential for repairing the defective DNA that causes disease.

Using homologous recombination, Kapsa and Kornberg found they could replace the defective dystrophin gene with a normal dystrophin gene in 15 percent to 20 percent of cells isolated from mice with DMD. Those results were reported in the April 10 issue of Human Gene Therapy.

In the only previous attempt to use this gene repair technique, scientists achieved a 1 percent success rate in cells carrying the defective gene for cystic fibrosis, making Kapsa and Kornberg's work an unprecedented success.

Kapsa and Kornberg are receiving MDA support for experiments to refine the technology.

Oxandrolone Has No Benefit in DMD

An MDA-supported, six-month study involving 51 boys with Duchenne MD found that the steroid drug oxandrolone (Oxandrin), when compared with a placebo (inactive substance), provided no benefit in that disorder.

Oxandrolone is an anabolic steroid, the group of drugs sometimes used to improve athletic performance. The drug had shown promise in a pilot study of 10 boys with DMD.

Although there was a difference in strength between the placebo group and the treated group on some measures, the authors concluded that the use of this drug isn't justified in DMD.

Neurologist Gerald Fenichel, MDA clinic director at Vanderbilt University Medical Center in Nashville, Tenn., and one of the study's authors, says, "While the drug is free of adverse reactions during the time tested, the efficacy was not of a magnitude to justify its use."

Fenichel added that he didn't think oxandrolone should be used to offset the growth-retarding effects of prednisone (Deltasone), a use that some experts have considered worth trying.

The study is published in the April 24 issue of Neurology. The drug and statistical support were supplied by Bio-Technology Corp.

MG Mystery Begins to Clear

A new study helps to solve a mystery that's long hampered diagnosis and treatment for some people with the debilitating autoimmune disorder myasthenia gravis.

MG occurs when the immune system makes wayward antibodies that attack the connection between nerve and muscle (the neuromuscular junction). At the normal NMJ, the nerve stimulates the muscle by releasing a chemical signal called acetylcholine (ACh), which the muscle intercepts using cell-surface proteins called ACh receptors. ACh receptors are concentrated underneath the nerve to ensure efficient capture of ACh.

MG is usually caused by antibodies that destroy ACh receptors, and it nearly always responds to treatment with immunosuppressants. But 10 percent to 20 percent of people with MG don't have those antibodies in their blood. In those cases, it's difficult for doctors to distinguish MG from nonimmune diseases affecting the NMJ, like congenital myasthenic syndrome (see "Congenital Myasthenic Syndromes").

A study in the March issue of Nature Medicine suggests that most of those baffling MG cases (70 percent) are caused by antibodies against the muscle-specific receptor tyrosine kinase (MuSK), a protein that normally helps cluster ACh receptors underneath the nerve during development. In the study, a group of European researchers headed by Angela Vincent at John Radcliffe Hospital in Oxford, Britain, found that 17 out of 24 MG patients who lacked the ACh receptor antibodies had MuSK antibodies. The latter antibodies, the researchers showed, inhibited the MuSK protein's ability to promote ACh receptor clustering in cultured cells.

For people with MG caused by anti-MuSK antibodies, new blood tests will improve diagnosis, as well as the monitoring of the effectiveness of immunosuppressant therapy, the researchers say. They also suggest that genetic defects in MuSK might actually cause some forms of congenital myasthenic syndrome.

Narcolepsy Drug Promotes Alertness in MMD

In a German study, nine people with myotonic muscular dystrophy (MMD) who experienced excessive daytime sleepiness benefited from the drug modafinil (Provigil).

Provigil, manufactured in the United States by Cephalon Inc., is usually prescribed for narcolepsy, a neurological condition marked by excessive daytime sleeping. It has been prescribed for similar symptoms in MMD, and anecdotal reports have been positive.

About one-third of people with MMD have daytime sleepiness severe enough to interfere with driving, working and social life. The cause seems to be a disruption in the brain's control of sleeping and waking, although respiratory impairment can also contribute.

Modafinil is believed to act on the brain's sleep-wake control center. No psychiatric problems or other side effects occurred with the drug, and nighttime sleep wasn't impaired.

The study was conducted at the Carl Gustav Carus University Hospital in Dresden, Germany, and was supported in part by Merckle GmbH, manufacturer of modafinil in Germany. Results are published in the March 27 issue of the journal Neurology. (For more on sleep problems in neuromuscular disorders, see "Better Nights for Better Days," Quest, vol. 7, no. 5.)

Mouse Model Created for Nemaline Myopathy

In an effort to identify possible treatments, researchers have created the first mouse model of the muscle disease nemaline myopathy.

Nemaline myopathy is really a spectrum of inherited diseases that cause muscle weakness at different ages, with varying levels of severity. Despite those differences, it's universally defined by the presence of nemaline (threadlike) rods — characteristic clumps of protein in the affected muscles. It's known that those rods arise from genetic defects in one of at least four different muscle proteins.

In the Feb. 15 issue of Human Molecular Genetics, a team of Australian researchers describes mice that have a defect in the muscle protein alpha-tropomyosin-slow, which causes childhood-onset nemaline myopathy in a large Australian family.

The team, including former MDA grantee Peter Gunning of the University of Sydney, found that the mice show typical features of the human disease, including a late-onset weakness and nemaline rods.

The scientists also noted that the mice have a not-so-typical feature of the disease that might be exploited as a means of treatment. In some people with nemaline myopathy, certain fast-type muscle fibers actually become larger and stronger, in a process known as hypertrophy. The researchers observed hypertrophy in those same muscle fibers in the mice, and found it was associated with lower levels of muscle weakness.

"This feature makes the nemaline mouse a particularly powerful tool to investigate [hypertrophy as an] innate therapeutic mechanism," the researchers say at the conclusion of their study.

Disability in Type 1 CMT Underestimated, Report Says

Type 1 Charcot-Marie-Tooth disease has often been considered one of the less severe neuromuscular disorders, so researchers at the University Hospital Eppendorf in Hamburg, Germany, were somewhat surprised to learn that the disorder can have a profound impact on daily living and lifestyle choices.

In a study of 50 people with type 1 CMT, which affects the myelin sheath around peripheral nerves (see "From Clear-Cut Endings to Complex Beginnings," Quest, vol. 8, no. 1), large numbers of participants reported higher than average needs for sleep and rest, slowness or clumsiness in daily activities, pain and a wish to avoid childbearing.

CMT patients reported emotional stress comparable to that reported by patients six months after a stroke who had no mental impairment. Some 18 percent of participants reported they were depressed. These findings revealed a greater degree of impact of type 1 CMT on quality of life than previous research has suggested.

The study is in the April issue of the Journal of Neurology, Neurosurgery and Psychiatry.

Sifting Through SBMA Stickiness

In SBMA, polyglutamine (PG) in the androgen receptor (AR) becomes enlarged, sticking tightly to the gene regulatory protein CBP and sequestering it away from its target genes.

Recent studies could help scientists leap forward in their efforts to understand and treat a group of neurological disorders that includes spinal-bulbar muscular atrophy (also known as Kennedy's disease) and Huntington's disease.

On the surface, it's far from obvious that the two diseases are related. They produce different symptoms, and arise from genetic mutations in distinct proteins. In SBMA, the culprit is the androgen receptor protein, while in HD, it's huntingtin.

In both proteins, the disease-causing mutations alter polyglutamine — a piece of protein made up of the building block glutamine. Normally, polyglutamine is thought to form a sticky spot where one protein can attach to another to carry out important reactions in a cell. But in SBMA and HD, a genetic mutation expands the polyglutamine, making it so clingy that it creates a tangled mass of proteins, or an aggregate, found inside dying nerve cells (neurons) in these disorders.

Last year, Kenneth Fischbeck of the National Institutes of Health in Bethesda, Md. — who, in 1991, identified the genetic defect underlying SBMA in MDA-backed research — showed that the expanded androgen receptor protein tends to trap and disable CBP, a polyglutamine-containing protein that controls gene expression. Fischbeck found that he could protect cultured neurons from the expanded androgen receptor by giving them extra CBP, suggesting a potential SBMA treatment. The study was published in Human Molecular Genetics.

In the March 23 issue of Science, a group at Johns Hopkins University in Baltimore showed that expanded huntingtin has nearly the same effects on CBP as the expanded androgen receptor. Since many scientists are already screening for drugs that inhibit huntingtin aggregation, they might discover drugs that are equally effective for treating HD and SBMA.

"I think it's reasonably likely that [drugs] that are effective in preventing huntingtin aggregation or toxicity may have applicability to SBMA," says MDA grantee Marc Diamond of the University of California at San Francisco. Eventually, Diamond hopes to begin screening for drugs aimed specifically at the expanded androgen receptor that causes SBMA.

Gentamicin Studies Yield Confusing Results in MDs

Two studies of the antibiotic gentamicin (Garamycin) in Duchenne (DMD), Becker (BMD) and limb-girdle muscular dystrophies (LGMD) found no evidence of the hoped-for result — that the drug would restore production of a needed muscle protein.

However, both studies found that serum creatine kinase (CK) levels decreased markedly in participants who took gentamicin. Elevated CK levels in the blood are associated with ongoing damage to muscle cells, from which the CK enzyme leaks into the bloodstream. Falling CK levels could mean less damage to muscles, but researchers don't know how this could happen in the apparent absence of restorative muscle protein activity.

In mice, gentamicin allowed cells to read through a stop codon mutation (genetic "stop sign") in the gene for dystrophin. The effects of gentamicin on protein production in humans with stop codons remains unclear.

In mouse studies, gentamicin has allowed muscle cells in the animals to "read through" a genetic code that otherwise would stop production of dystrophin, the muscle protein missing in DMD and abnormal or diminished in BMD. The researchers hoped the same thing would happen in people with MD who have the same kind of mutation, known as a premature stop codon.

Premature stop codons affect a small percentage of boys with DMD and BMD and an unknown number of people with LGMD. LGMD can result from the lack of a sarcoglycan, dysferlin or any of several other needed muscle proteins.

Eric Hoffman of the Research Center for Genetic Medicine of Children's National Medical Center in Washington, who was part of one gentamicin study that looked at DMD and BMD, said, "The CKs did go down quite a bit, but without dystrophin present, it is difficult to interpret what this means. Maybe the gentamicin was working perfectly but there was just not enough time to see anything significant in terms of protein production."

Testing higher doses for longer periods would "almost certainly lead to significant side effects," Hoffman said. "So we're somewhat between a rock and a hard place here." Gentamicin can interfere with hearing and kidney function when taken at high doses.

The study on which Hoffman was an author included two boys with DMD and two with BMD. It was funded by the National Institutes of Health and is in the April issue of the journal Neurology.

Another member of that study team, Kathryn Wagner of the Department of Neurology at Johns Hopkins Hospital in Baltimore and NIH in Bethesda, Md., said she didn't consider the results encouraging. "I don't recommend anyone using gentamicin just because the CK levels went down," she said.

The other study, which was MDA-funded, included 10 boys with DMD and three people with LGMD. Two LGMD participants had stop codon mutations in genes for beta-sarcoglycan, and one had a stop codon in a gene for dysferlin. Results of this study were reported at the annual meeting of the American Academy of Neurology (AAN) in May.

Among the researchers on that team were MDA clinic co-director Jerry Mendell of the Department of Neurology at Ohio State University in Columbus and Kevin Campbell of the Department of Physiology and Biophysics at the University of Iowa in Iowa City.

Mendell, who presented the findings at the AAN meeting, said he plans to do further studies with gentamicin. Other drugs might also be capable of restoring protein production from genes with stop codons, recent research has suggested.

Both studies gave each participant 7.5 milligrams per kilogram per day of intravenous gentamicin for two weeks.