DESIGNER COMPOUND COULD FIX SMA GENE

Scientists are working on a new strategy to compensate for the genetic defect behind spinal muscular atrophy (SMA) — a disease that kills muscle-controlling nerve cells (motor neurons) in the spinal cord.

Nearly all cases of SMA are caused by mutations in the SMN1 gene, which encodes a protein called survival motor neuron (SMN). Everyone has a "backup" gene called SMN2, but it mostly produces a shortened, inactive form of the SMN protein.

Still, in people and in mice with SMA, having more copies of the SMN2 gene lessens the severity of the disease, leading researchers to believe that using a drug to boost SMN2 might be an effective therapy.

To this end, some research groups have begun "high-throughput drug screens," testing scores of chemicals on cells from SMA patients, hoping to find some that will stimulate SMN2 to produce more of the full-length SMN protein (see "Research Updates," Quest, 2001, no. 6).

Luca Cartegni and Adrian Krainer of Cold Spring Harbor Laboratory in New York have taken a more direct approach to getting SMN2 to act like SMN1.

The key difference between the two genes lies in their RNA (the intermediate between genes and proteins). Before its converted into protein, RNA must go through splicing — a cutting and pasting process — with the help of a class of proteins called SR proteins. An essential piece of SMN2 RNA, called exon 7, lacks a signal recognized by the SR proteins and is thus removed, creating a shortened RNA and ultimately a shortened SMN protein.

To get around this problem, Cartegni and Krainer made a synthetic compound — part SR protein and part RNA — that recognizes SMN2 exon 7. When added to cell extracts, the compound caused an increase of up to 40 percent in the levels of full-length SMN2 RNA. Those experiments are part of a new study published online by Nature Structural Biology on Jan. 13.

Since the synthetic compound is designed to recognize SMN2 RNA, the researchers say its likely to have a specific effect on the gene, while the chemicals found so far via high-throughput screening might have unpredictable effects on other genes.

Unfortunately, they dont yet have a way to deliver their compounds into cells.

"It may be harder to have them taken up by cells or to target them to difficult sites, such as the spinal cord. We have not yet been successful in delivering them into cultured cells, but this work is in progress and I am fairly certain that it will eventually work," Krainer said.

SMA, caused by mutations in the SMN1 gene, might be treatable by enlisting the help of a "backup" gene, SMN2. SMN2 makes a short version of the SMN protein because its RNA lacks a signal that directs appropriate splicing by SR proteins (left). This shortened SMN protein is unable to compensate for loss of the full-length protein in SMA (center). Researchers have designed a synthetic SR compound that stimulates SMN2 to produce the full-length protein (right).

Gene Transfer Study Results Expected Soon

Researchers say theyll announce results in June of a small gene transfer study conducted in boys with Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) in France. The technology for the trial is based on the work of MDA research grantee Jon Wolff at the University of Wisconsin in Madison.

The study, which included nine boys age 15 or older, is designed to test the safety of a method for delivering the gene for dystrophin through the bloodstream to muscle cells without the use of viruses ("naked" DNA). Dystrophin is the muscle protein thats deficient in these dystrophies. The research was conducted at the Pitié-Salpêtrière Hospital in Paris.

Previous gene transfer (gene therapy) studies have almost all used viruses to transport the gene into cells. These organisms are efficient transporters but can be dangerous.

The study is being carried out by Transgene, a company based in Strasbourg, France, with offices in Boston; and Mirus of Madison, Wis.

"The present trial is only [for] safety," said Serge Braun, vice president of Research at Transgene. "A second trial would be to demonstrate efficacy" of the method, he said.

Gene Delivery Tested in Spinal Muscular Atrophy

Christine DiDonato, an MDA grantee and senior research associate at the Ottawa Health Research Institute (OHRI) in Ottawa in Ontario, is part of a team working on gene therapy for SMA.

With help from virologist Robin Parks and molecular biologist Rashmi Kothary, both scientists at OHRI & professors at the University of Ottawa, DiDonato used a virus to deliver the SMN1 gene to cells derived from people with SMA.

Although motor neurons are the ultimate target for the therapy, DiDonato tested the approach in skin cells because they're easier to obtain and grow in the lab, and they exhibit "one of the hallmarks of SMA," she said.

Normal human cells, she explained, contain "gems" — areas rich in SMN and other RNA-processing proteins — but cells from people with SMA contain a reduced number of gems or none at all. DiDonato and her colleagues showed they could restore gems to the cells by infecting them with an adeno-virus carrying SMN1.

Their study, published in the Jan. 20 issue of Human Gene Therapy, is a first step toward clinical trials of gene therapy for SMA, DiDonato said. "The next step is to go into animal models of SMA and see if we can correct the pathological defects that result from low intracellular levels of SMN," she said.

Utrophin-Based Therapy May Need to Start Early, Mouse Study Suggests

MDA grantee Kay Davies at the University of Oxford in England was part of a group that recently established that utrophin delivery in mice genetically destined to develop Duchenne muscular dystrophy (DMD) has to begin early to be effective. Utrophin is a muscle protein that can to some extent substitute for dystrophin, missing in DMD.

In a paper published in the Dec. 15 issue of Human Molecular Genetics, Davies and colleagues describe a system they designed in which the gene for utrophin is added to the dystrophin-deficient mice. In this high-tech system, utrophin production from the added gene can be turned off by giving the animals the antibiotic tetracycline in their drinking water.

Mice that didnt produce utrophin until 30 days of age experienced much less benefit from it than did dystrophin-deficient mice in which utrophin production started earlier — for instance, at birth and at 10 days.

Davies said that "early is best" for utrophin production. She added, "I am pretty confident that this will extrapolate to humans. The later stages are harder to predict. However, if we could protect DMD respiratory and cardiac muscle and maintain the use of the arms, this would be an enormous advance."

DMD Researchers Enhance Marrow-to-Muscle Switch

The stem cells in bone marrow were once thought to give rise only to blood cells, but its now recognized that they can cross tissue boundaries and become diverse cell types, from muscle fibers to nerve cells.

Hoping that this "plasticity" might allow bone marrow stem cells to repair damaged muscles, several research groups have tried transplanting bone marrow into mice with Duchenne muscular dystrophy (DMD). But when the muscles of the transplant recipients are examined, the result is always the same: Less than 1 percent of the cells are derived from the donor.

In a new study on normal mice, published in the Nov. 15 issue of Cell, Mark LaBarge and Helen Blau of Stanford University in California show that its possible to increase that number to nearly 4 percent by deliberately injuring the transplant recipients muscles.

LaBarge and Blau reasoned that normal muscles, and perhaps even muscles affected by DMD, fail to attract bone marrow stem cells because they already have stem cells of their own, called satellite cells. So, prior to giving the mice a bone marrow transplant, the researchers exposed some of them to high-dose irradiation, which destroyed their satellite cells and made room for the bone marrow cells.

After the transplant, they gave some of the mice a running wheel, which made them exercise and wear down their muscles, stimulating the bone marrow stem cells to make new muscle fibers.

Although procedures like these would never be used on children with DMD, they show its possible to increase the conversion of bone marrow to muscle. Along with previous findings, they also suggest that damaged muscles release signals to attract stem cells.

Identification of these signals might improve the prospects of bone marrow transplantation for DMD.

Myotonic MD Study Needs 60 Participants

A study of excessive breakdown of immune-system proteins known as IgG in myotonic dystrophy (MMD) continues under the direction of Harvard Medical School in Boston and needs 60 more participants who are at least 18 years old.

The study requires only a blood sample, which can be drawn by the participants local physician and mailed to Boston at the investigators expense. Each participant will receive $25. No one will be identified by name.

The investigators will check IgG protein levels and analyze the DNA from the blood sample. Potential participants will be contacted by phone for a full discussion of the study and informed consent.

For information, contact Tamara Dolan at (617) 432-7004 or tamara_ dolan@hms.harvard.edu.

New CMT Gene Identified A research team led by MDA grantees Phillip Chance and Valerie Street at the University of Washington in Seattle has linked yet another gene to Charcot-Marie-Tooth disease (CMT). CMT actually refers to a group of genetic diseases, each one caused by mutations in a distinct gene, which cause nerve degeneration and weakness. The Washington researchers discovery marks the 12th gene linked to CMT, and is expected to lead to improved genetic diagnosis. The team found the gene by studying three large families with CMT type 1. This common form of the disease involves damage to the protective covering around nerve fibers — called myelin — and is inherited in an autosomal dominant manner, meaning it takes just one defective copy of a gene to cause the disease. Previously, its been shown that 70 percent to 80 percent of CMT1 cases (designated CMT1A) are caused by mutations in the PMP22 gene. But in some cases (CMT1C), the defective gene underlying the disease was unknown. A single peripheral nerve is composed of many long nerve cell branches — or axons — that extend from the spinal cord and connect to muscle fibers. Each axon is surrounded by myelin made from the wrappings of Schwann cells. Last year, Chance and Street narrowed their search for the gene to chromosome 16 (see "Research Updates," February 2002), and now theyve found CMT1-causing mutations in the gene encoding LITAF, a protein that might regulate the breakdown of other proteins in cells that form myelin. "We predict that such mutations may account for a significant proportion of CMT1 patients lacking a mutation in previously known genes [such as PMP22]," they write in the Jan. 14 issue of Neurology. Street told MDA that she hopes genetic testing for the mutations will be available soon. Meanwhile, researchers based at the University of Antwerpen in Belgium say theyve found evidence that mutations in the GDAP1 gene are the most frequent cause of CMT inherited in an autosomal recessive manner (two defective copies of a gene are required to cause the disease). Their study appeared in the Dec. 24 issue of Neurology.

Modafinil May Relieve Extreme Sleepiness in Myotonic MD

A drug called modafinil (brand name Provigil) thats been used to treat abnormal and disruptive sleep "attacks" (narcolepsy) has also been found helpful in treating the excessive daytime somnolence (sleepiness) sometimes associated with myotonic dystrophy (MMD).

In a study conducted at McMaster University Medical Center in Hamilton, Ontario, Canada, Mark Tarnopolsky of the Neuromuscular Disease Unit and colleagues studied 40 adults with MMD who described themselves as excessively sleepy during the day.

Most patients had genetically confirmed type 1 (chromosome 19) MMD; two had genetically confirmed type 2 (chromosome 3) MMD. On average, participants had had excessive sleepiness for 10 years.

Participants filled out questionnaires about their moods, sleep experiences and physical symptoms.

Those on modafinil were less sleepy on self-rating questionnaires than those given a placebo. Those taking modafinil also judged themselves to have decreased "fatigue-inertia" and increased "vigor-activity," although they also noted moderate increases in the "tension-anxiety" part of the mood spectrum.

Modafinil appeared to decrease daytime sleepiness and lead to an enhanced mood in general, despite the slight increase in tension and anxiety. In all but two patients, the self-judged benefit of treatment with modafinil outweighed any negative effects.

The authors conclude, in a paper published in the Dec. 24 issue of Neurology, that "the results of this and other studies suggest that modafinil may have the potential for widespread use in numerous neurologic disorders in which EDS [excessive daytime somnolence] is a bothersome symptom."

Two Genes Shape Muscles of the Face

In diseases like facioscapulohumeral muscular dystrophy (FSHD) and oculopharyngeal muscular dystrophy (OPMD), certain muscles in the face become severely weakened while many other muscles in the body remain strong.

FSH dystrophy causes extreme facial weakness, making it hard to talk or smile.

The reasons for these differences are unclear, but an MDA-funded research team has found a pair of genes — called MyoR and capsulin — whose activity appears to set facial muscles apart from other voluntary muscles.

Led by Eric Olson at the University of Texas Southwestern Medical Center in Dallas, the team showed that mice lacking both genes fail to develop certain facial muscles, but have normal limb and trunk musculature.

Their discovery, published in the Dec. 20 issue of Science, may lead to a better understanding of why facial muscles are especially affected in some forms of MD.

Registry Seeking People With FSH, Myotonic MDs

Researchers at the University of Rochester (N.Y.), with funding from the National Institutes of Health, want to remind people that the National Registry of Myotonic Dystrophy (MMD) and Facioscapulohumeral Muscular Dystrophy (FSHD) Patients and Family Members still needs participants.

The registry is designed to help people with these two disorders link up with scientists studying these conditions and participate in research if they wish to do so.

To add your name, click here; then scroll to either MMD or FSHD National Registry.

Or contact MMD study coordinator Cheryl Barbieri at cheryl_barbieri@ urmc.rochester.edu; or Lynn Cos, study coordinator for FSHD, at lynn_cos@ urmc.rochester.edu

You can also call, toll-free, (888) 925-4302.

Some MDA clinics may have registry applications as well.

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