Research Updates
Halted LGMD Trial of Gene Therapy Shows No Benefit, No Harm
Two people with limb-girdle muscular dystrophy (LGMD) due to a deficiency
of the alpha-sarcoglycan protein who were injected with genes for that protein
tolerated the gene transfer well, but neither showed any benefit from the
experimental treatment, the researchers say.
These findings, based on a 1999 MDA-supported gene therapy trial, were presented
at the annual meeting of the American Academy of Neurology in Denver in April.
The trial was halted prematurely in January 2000 because of the death of a
participant in an unrelated gene therapy trial and subsequent rulings by the
U.S. Food and Drug Administration.
In the LGMD trial, a 37-year-old man and a 14-year-old boy were each injected in
one foot muscle with genes for alpha-sarcoglycan and given a placebo injection
in the other foot muscle. Neither researchers nor participants knew which foot
got the gene transfer until recently.
 Jerry Mendell |
"Both patients tolerated the injections into the EDB [extensor digitorum brevis
muscle] without adverse effects," reported the research team led by Jerry
Mendell of Ohio State University in Columbus. Mendell is a neurologist and
co-director of the MDA clinic at Ohio State.
"Neither developed signs of local or systemic reaction at any time. [However,]
no differences could be discerned in histopathology [the appearance of
thetissue] comparing the right and left EDBs of either patient. Dystrophic
changes were observed on both sides."
The researchers concluded that the EDB muscle posed problems for the
interpretation of gene transfer results, and that in future trials the gene
should be injected into limb muscles.
Mendell's department is testing people with LGMD for genetic mutations as a
basis for possible inclusion in a future gene therapy trial. For information,
contact Mendell at (614) 293-9016, or e-mail him at mendell.1@osu.edu.
MDA announced in February that it has awarded a grant to researchers at the
University of Florida in Gainesville to explore new approaches to LGMD gene
therapy (see "Research Updates,"
Quest, April 2002).
Gentamicin Trial for DMD Stop Codons Expanded
The antibiotic gentamicin has shown confusing results in studies of boys
with Duchenne muscular dystrophy (DMD) and other forms of MD.
Now, MDA-supported researcher Jerry Mendell at Ohio State University in
Columbus wants to get to the bottom of the story with an additional study.
In 1999, when researchers gave gentamicin to mice with DMD (mdx mice), they
found evidence that the drug allowed muscle cells in the animals to "ignore" a premature
stop codon and begin to make dystrophin, the protein missing in DMD.
(See "Research
Updates," vol. 6, no. 4, August 1999.)
A premature stop codon is the type of mutation thought to affect some 5 percent
to 15 percent of boys with DMD. Premature stop codons tell the cell to stop
"reading" the gene too soon, so the protein the cell would ordinarily make from
the gene is too short.
Most boys with DMD have a deletion mutation, which removes part of the
instructions for protein manufacturing, with similar results.
In two small clinical trials of gentamicin in people with stop codons, results
were puzzling. (See "Research
Updates," June 2001.)
The investigators didn't see any increase in dystrophin in the muscle cells, but
they found decreased blood levels of the enzyme creatine kinase, a marker of
muscle destruction. This led researchers to think gentamicin might have
improved the health of the patients' muscle cells. (See "Simply
Stated," Quest, vol. 7, no. 1, February 2000.)
Mendell thinks some increased dystrophin production may have occurred, but
another interpretation is that the gentamicin had an effect, not on stop
codons, but on some other aspect of muscle cell preservation. (For instance,
the drug may have stabilized the muscle cell membrane.)
To help clarify the mechanism by which gentamicin appears to lower CK levels in
DMD, the Ohio State research team gave a short course of intravenous gentamicin
to boys with DMD who had deletion mutations. The team is now analyzing these
results.
Normal
DYSTROPHIN FULL LENGTH GENE |
Stop Codon Mutation + Gentamicin
GETAMICIN DYSTROPHIN GENE WITH STOP CODON |
DYSTROPHIN FULL-LENGTH PROTEIN |
DYSTROPHIN FULL-LENGTH PROTEIN |
Stop Codon Mutation
DYSTROPHIN GENE WITH STOP CODON |
| In mice, gentamicin allowed cells to read through a stop codon mutation (genetic "stop sign") in the gene for dystrophin. A new trial could help to explain how gentamicin may affect boys with Duchenne MD. |
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SHORTENED DYSTROPHIN PROTEIN |
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In the next few months, with MDA support, the researchers will begin a six-month
trial of gentamicin in boys with DMD and premature stop codons. The drug will
be administered on an outpatient basis, and some visits to Columbus will be
necessary.
The team is testing boys ages 5 to 15 who have DMD and meet certain other
criteria for the gentamicin study to see what kind of mutation they have.
For more information on testing and trial criteria, contact Cheryl Wall or
Jennifer Berarducci, clinical trial coordinators, at (614) 293-9016 or e-mail wall.49@osu.edu or mendell.1@osu.edu.
Studies Reveal Causes, Potential Treatments for Myotonic Dystrophy
MMD1: Genetic Trigger Defined
The most common form of myotonic muscular dystrophy (MMD1) is caused by a trinucleotide
repeat expansion. But what causes this mutation to occur in the first
place?
MDA grantee Christopher Pearson, a molecular biologist at the Hospital for Sick
Children and the University of Toronto in Canada, believes the answer might
hold clues to treatment. In work supported by MDA, he's found that the
expansions are at least partly triggered by peculiarities in neighboring, or
flanking, DNA.
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NORMAL
repeats
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MMD
expanded repeats
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DNA |
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RNA |
RNA |
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| MMD1 & MMD2 are caused by mutations that increase or "expand" repeated
segments of DNA. The expanded DNA is converted into abnormally long RNA, which
appears to cause widespread defects in RNA processing. |
"If we could fix the flanking region [of DNA] maybe we could control the
expansion," Pearson said. Such a treatment, hypothetical at this point, could
be used to prevent the passage of MMD1 from parents to their children. In
addition, although it wouldn't reverse symptoms in people who already have the
disease, it could stop them from getting any worse, he said.
A trinucleotide repeat expansion is sort of like a run-on sentence in DNA.
Normally, the DMPK gene, located on chromosome 19, contains four to 24 repeats
of the three-letter phrase CTG (representing chemicals that make up DNA). But
in families with MMD1, those repeats are unstable, poised to multiply (expand)
whenever new copies of the gene are made in new cells. Having more than about
50 CTG repeats triggers symptoms of the disease, and in severe cases, the
repeats can climb into the thousands.
If the expansions occur in a person's sperm or eggs, MMD1 can jump — and
escalate — from one generation to the next. If they occur in other cells of the
body, such as muscle or nerve cells, the disease could worsen over a person's
lifetime, scientists believe.
To figure out what causes the expansions, Pearson developed a system to examine
them in monkey cells. These experiments, described in the May issue of Nature
Genetics, show that the repeats are influenced by flanking DNA. Pearson also
designed a system using extracts from human cells, described in the April 19
issue of the Journal of Biological Chemistry.
Pearson's research could have implications for other "repeat" diseases,
including MMD2 (caused by a four-nucleotide repeat expansion in the ZNF9
gene) and Kennedy's disease.
MMD1 & MMD2: Myotonia Explained
Researchers believe the repeat expansions underlying MMD1 and MMD2 interfere
with the way cells handle RNA, the chemical intermediate between DNA and
proteins. In fact, it's thought that a widespread defect in RNA processing
probably accounts for the diverse symptoms of MMD, which include muscle
wasting, cardiac problems and cataracts.
Now, MDA grantees Ami Mankodi and Charles Thornton of the University of
Rochester in New York have tied defective RNA processing to one of the most
prevalent symptoms of MMD — myotonia (muscle stiffness).
Mankodi and Thornton took a hint from myotonia congenita, a disease that
can look similar to MMD, but is caused by genetic defects in chloride channels,
pores that allow negatively charged chloride particles to flow into muscle.
When those pores open, the muscle relaxes, but when they don't open enough, the
muscle can become myotonic.
Mankodi and Thornton examined muscle cells from mice with MMD1 and from people
with MMD1 and MMD2, and found extensive abnormalities in the RNA that produces
chloride channels.
The study, reported in April at the American Academy of Neurology annual meeting
in Denver, suggests that drugs effective against myotonia congenita could also
alleviate MMD-associated myotonia. One such drug, mexiletine (Mexitil), is
already in clinical trials for MMD (see "Research
Updates," April 2002).
MMD1: Immune System Explored
Besides their effects on RNA, there's evidence that the repeat expansions in the
DMPK gene might have more direct effects on several genes on chromosome 19. For
instance, researchers have noted that the expansions turn down the activity of
certain genes near DMPK.
Marilyn Webster, Alex Ebralidze and Richard Junghans of Beth Israel Deaconess
Medical Center in Boston suspect that this "position effect" explains a
long-standing mystery in MMD1. Over 30 years ago, Webster says, investigators
found that people with MMD1 had abnormally low levels of IgG antibodies (a part
of the immune system's defenses). Reduced levels of IgG don't cause immune
system problems in people with MMD1, but "could be used to uncover mechanisms
behind the disease," she says.
Webster and her colleagues theorize that the repeat expansions in DMPK adversely
affect a nearby gene called FCGRT, which plays a role in stabilizing IgG in the
blood. To test the idea, they're collecting blood samples from people with
MMD1, which they'll use to measure IgG levels and probe for variations in the
FCGRT gene.
Each participant in this MDA-funded study will receive $25 for donating a blood
sample, which can be mailed to Boston. For more information, contact Alex
Ebralidze at (617) 432-7013 or aebralidze@hms.harvard.edu,or
Marilyn Webster at (617) 432-7004 or marilyn_webster@hms.harvard.edu.
Enzyme Protects Mice Against Duchenne MD
In an MDA-funded study, researchers have found that boosting the levels of a
protein called CT GalNac transferase can block muscle wasting in mice
with Duchenne muscular dystrophy (DMD).
"In terms of therapy for DMD, this [protein] is a new target to look at that was
previously not evident to anybody," says the study's lead researcher Paul
Martin, a neurobiologist at the University of California in San Diego. His
finding is unique because CT GalNac transferase has a function quite different
from that of dystrophin, the protein that's missing in DMD.
Dystrophin forms part of an essential complex of proteins in the membrane
(surface) of muscle fibers, and its absence makes the complex fall apart,
except in small patches that contain dystrophin's sister protein utrophin.
 Paul Martin |
Many scientists hope to restore the complex, and thus halt DMD, by boosting the
levels of dystrophin or utrophin.
Integrin, another protein found in muscle membrane, has shown similar promise.
In contrast to dystrophin and its counterparts, CT GalNac transferase is one of
many enzymes that attach small sugar groups (in this case, GalNac) onto
proteins,a process known as glycosylation. Glycosylation is important
for the function of many muscle proteins, including those at the root of other
forms of muscular dystrophy.
Martin and his team became interested in CT GalNac transferase when they noticed
that it's concentrated in utrophin complexes in muscle cells. Guessing that the
enzyme might help stabilize those complexes, they introduced extra copies of
the gene for the enzyme into mice with DMD. In those mice, the enzyme and the
utrophin complexes spread over the entire surface of each muscle fiber, and
kept the fibers from degenerating.
The study was published in the April 16 issue of the Proceedings of the National
Academy of Sciences.
Delivering the enzyme via gene therapy or activating it with a drug might be an
effective way to treat DMD and other MDs, in particular merosin-deficient
congenital muscular dystrophy, Martin says. Similar treatments for DMD
are being pursued with dystrophin and utrophin, but the enzyme might be
superior because just a little of it could have far-reaching effects, he says.
Green Tea Protects Muscle in DMD Mice
Green tea appears to reduce or at least delay muscle cell death in mice with Duchenne
muscular dystrophy (DMD), say Urs Ruegg and colleagues at the School of
Pharmacy at the University of Lausanne in Switzerland, who published a paper on
the subject in the April issue of the American Journal of Clinical Nutrition
(see also "Staying Healthy From the
Outside In,"). The substance may have potential for staving off cell
death in boys with the disorder, although they'd have to drink seven cups of it
a day to reach levels equivalent to what the mice received.
DMD-affected mice that were fed an extract of green tea as newborns (directly
and possibly through their mothers' milk, since the mothers also ingested it)
showed less muscle destruction than untreated mice in one leg muscle, but
didn't show the same benefit in another leg muscle. The two leg muscles have
cells with slightly different chemical properties, which may account for the
difference in benefit, the researchers say.
The investigators interpreted the findings to mean that the tea probably
combated oxidative stress, a type of cellular damage seen in many
disorders and one that has been implicated in DMD (see "Simply
Stated," Quest, vol. 7, no. 2, April 2000).
Mark Tarnopolsky of the Neuromuscular Disease Unit of McMaster University
Medical Center in Hamilton, Ontario, Canada, has had MDA funding to study
energy production in muscle cells and is interested in antioxidants,
compounds that fight off oxidative stress by neutralizing the chemicals that
cause it (known as free radicals).
"There's a potential for antioxidants to be of benefit in Duchenne dystrophy,"
he said. "There's clearly an increase in oxidative damage when dystrophin [the
protein missing in DMD and diminished in Becker MD] is abnormal. The
next step is to see which compounds are safe and effective."
New Look at Inflammation in Duchenne Dystrophy May Open Treatment Doors
In a study of mice with Duchenne muscular dystrophy (DMD), scientists
have found that inflammation plays a large part in the disease, perhaps ending
an old debate and opening a new door to treatment.
Inflammation, an invasion of immune cells into damaged tissue, is often seen in
muscles affected by DMD, but scientists have long questioned whether it's a
contributing factor — or just a consequence — of muscle wasting. Some point out
that anti-inflammatory steroids like prednisone can slow the disease, but
others say these drugs may have some unknown mechanism of action.
The authors of the MDA-funded study say their results should encourage
scientists to revisit inflammation in DMD with an eye toward developing
anti-inflammatory drugs superior to prednisone, which often causes adverse side
effects. The study was published in the February issue of Human Molecular
Genetics.
"I think there is a strong potential for immunotherapy in DMD," says lead author
John Porter of Case Western University and University Hospitals of Cleveland.
Using "gene chip" technology (see "Fast-Track
Pharmacy," Quest, vol. 8, no. 4, August 2001), Porter, co-investigator
Francisco Andrade and their team found a spike in the activity of 242 genes in
muscles from mice with mid-stage DMD. Many of those genes encode proteins found
on or secreted from immune cells; altogether, nearly 30 percent of them have
some function related to inflammation.
To Porter, each one of the genes is a possible target for therapeutic
intervention. "The long-term potential is thatpharmaceutical [companies] are
developing specific drugs to block the recruitment of inflammatory cells," he
says. "If we can block the recruitment of inflammatory cells and it helps
alleviate the disease in mice, the same approach may become very plausible in
DMD patients."
Biopsy Samples Needed From Those With OPMD
If you have oculopharyngeal muscular dystrophy (OPMD) and have undergone
a muscle biopsy, MDA-supported molecular geneticist Eric Hoffman would like to
hear from you.
 Eric Hoffman |
Hoffman's group, which is studying the reasons underlying the onset and
progression of OPMD, would like to receive "flash-frozen" OPMD muscle samples
that weigh at least 0.1 grams. Hoffman also wants information about your
clinical status and history, and about your genetic mutation (PABP2 or other
mutation), if available. But all identifying data (such as your name) should be
removed from each sample.
The samples should be sent on 10 pounds of dry ice. Hoffman's lab will pay any
processing or postage charges.
You can have your physician contact Hoffman at ehoffman@cnmc.org.
Samples should be sent to Eric Hoffman, Ph.D., Director, Research Center for
Genetic Medicine, Children's National Medical Center, 111 Michigan Ave. NW,
Washington, DC 20010.
Oxandrolone, Interferon Eyed in IBM Myositis
A small study of the drug oxandrolone, an anabolic steroid related
to the male hormone testosterone, showed modest benefits in treating the
weakness associated with inclusion-body myositis (IBM) and may have
increased muscle bulk as well.
Fourteen men and two women with IBM participated in a clinical trial of
oxandrolone at Beth Israel Deaconess Medical Center in Boston and the
University of Connecticut in Farmington.
The drug had a borderline significant effect in improving whole-body strength
and significantly improved arm strength as measured by maximal voluntary
isometric contraction testing. It also increased lean body mass (the weight of
the body minus its fat content) by 1.1 percent, compared to an average decrease
of 0.6 percent seen in untreated patients.
"That lean body mass increased significantly with oxandrolone as compared with
placebo ... helps support the supposition that at least some muscles of
patients with IBM are responsive to anabolic compounds," the researchers wrote
in their paper, published in the April 9 issue of Neurology.
Anabolic steroids generally build muscle but can have masculinizing effects in
women and can cause liver damage, difficulty sleeping and other side effects in
both sexes.
The researchers say further study of oxandrolone, possibly in combination with a
drug that acts on the immune system (thought by some to function abnormally in
IBM), is warranted, although no further studies are planned at this time.
The study was funded by a grant from the National Institutes of Health and by
BioTechnology General Corp., makers of the Oxandrin brand of oxandrolone.
A multicenter study of interferon beta-1a (brand name Avonex) in
IBM, funded by the National Institutes of Health, is closed to recruitment,
with results expected in 2003.
Interferon beta-1a is thought to fight viruses and have complex effects on the
immune system. It's on the market for relapsing multiple sclerosis, a disorder
in which the immune system attacks a part of the nervous system called the myelin
sheath.
First Fruits of MD-CARE Act Lead to Meeting on Data Gathering
In December 2001, President Bush signed into law the Muscular Dystrophy
Community Assistance, Research and Education Amendments. Known as the MD-CARE
Act, the bill directs agencies of the federal government to put increased
emphasis on developing treatments and cures for all forms of muscular dystrophy.
In March, Sharon Hesterlee, MDA's research development director, attended a
meeting of the Centers for Disease Control and Prevention (CDC) in Atlanta,
where a panel of representatives from voluntary health agencies joined CDC
staff members in discussing what kinds of data would be helpful to gather about Duchenne
and Becker muscular dystrophies. The CDC has been granted $2 million to
gather the information.
In addition to Hesterlee, the panel included three MDA clinic directors and an
MDA-funded investigator.
Discussions centered around what type of statistical information would most aid
research and clinical care efforts. Topping the list of priorities were finding
out how DMD and BMD are distributed in the U.S. population, what correlations
can be made between genetic mutations and clinical course in each disease, and
what kinds of services families need most. 
MORE MDA RESEARCH NEWS
For up-to-the-minute news on MDA research developments, visit MDA's Web site at www.mda.org. Click on "Research"
for information on recent research developments and active clinical trials, and
links to major medical/research sites. Look at the Web site's "What's
New" section for news bulletins about breaking research announcements.
For research news about amyotrophic lateral sclerosis, see The MDA/ALS
Newsletter or go to www.als-mda.org. |
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