Download repurposing drugs for duchenne muscular dystrophy

Professor Stanley Froehner tells International Innovation how he came to realise the human
impact of his work in repurposing drugs to treat Duchenne muscular dystrophy
muscle loss in DMD, and the nitric oxide
pathway seemed like a good place to start.
Have you encountered any major
challenges in the course of your research?
How have these been overcome?
When did you become interested
in researching Duchenne muscular
dystrophy (DMD)?
In the late 1980s, we were attempting to
identify novel proteins at the neuromuscular
synapse, using an unbiased monoclonal
antibody approach. When the DMD gene
was identified in 1987 by Dr Lou Kunkel’s
laboratory, we realised that we had antibodies
to dystrophin and some of its associated
proteins. This was very exciting. Research in
our laboratory had already been supported
by the Muscular Dystrophy Association
(MDA) for several years, and suddenly
we found ourselves working on the most
devastating of all the muscular dystrophies.
What caused you to turn your efforts toward
identifying drugs for DMD treatment?
In 2001, I was invited by Pat Furlong to attend
the annual meeting of Parent Project Muscular
Dystrophy (PPMD), to present a lay level talk
about our research. Pat had lost two sons to
DMD and started PPMD to help other Duchenne
families deal with the devastating diagnosis
and promote research into this disease. At
the conference, attended by several hundred
parents of DMD boys, I realised that I had never
before met anyone with any connection to
DMD. The parents were intensely interested in
our research and had many questions. I left the
meeting thinking about how we might use our
basic science findings to find treatments. We
continued our basic research but also began to
consider drugs that might slow the relentless
Maintaining adequate funding for the
laboratory is a problem that all biomedical
scientists are facing. We depend more
now on private sources and foundations.
Also, while collaborations with biotech
and pharma are more common, the goals
of academic medical research and the
commercial world are not always compatible.
With the shift in emphasis to translational
research at the National Institutes of
Health (NIH), applications that propose
basic research are severely disadvantaged.
This is unfortunate, because with so much
not understood about the pathogenic
mechanisms that cause DMD, important
therapeutic targets may remain unidentified.
As in many areas of biomedical research,
collaboration among individuals with a broad
range of expertise is absolutely necessary in
the study of DMD. Our lab brings together
individuals with expertise in biochemistry,
molecular biology, cardiac and skeletal muscle
physiology and animal husbandry, all with the
single purpose of understanding the molecular
pathogenesis of DMD. Of course, we collaborate
widely with our colleagues here at the University
of Washington and also at other research
institutions, both nationally and internationally.
(FDA), even for a different disease, has already
been through toxicology and pharmacokinetic
studies, an expensive and time-consuming
process. Of course, if the drug has not been
approved for use in children, then additional
studies might be required. Nevertheless, the
information available about a drug already
in use can be an enormous help in meeting
the requirements for a clinical trial and allow
human studies to proceed much more rapidly.
PROFESSOR STANLEY FROEHNER
Teaching old drugs new tricks
You are currently a member of the Scientific
Advisory Committee for PPMD. What
activities do you undertake within this role?
As a member of the PPMD Committee, I
participate in reviews of grant applications
submitted to the foundation and serve on an
oversight and advisory committee for a large
research project funded by PPMD. Members
of the Scientific Advisory Committee also
advise PPMD on priorities as the foundation
tries to balance the need for funding basic
and translational research, clinical trials
and high-risk, high-payoff proposals.
How does the process of trialling an
existing drug for new uses differ from
testing a completely novel drug?
An existing drug that is already approved for
use by the US Food and Drug Administration
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PROFESSOR STANLEY FROEHNER
Halting muscular degeneration
Progress in the basic research of a rare muscular dystrophy has not yet translated into effective treatments,
but novel insights from scientists at the University of Washington are providing new hope for patients
DUCHENNE
MUSCULAR
DYSTROPHY
(DMD) is a severe genetic disorder characterised
by progressive muscle degeneration and
consequent muscle weakness. As a recessive,
X-linked condition, it almost exclusively impacts
males – with approximately 200,000 thought to
be affected worldwide.
DMD starts to show its effects during early
childhood. The typical patient has difficulty
standing upright from the prone position and
struggles to climb stairs, together with a range
of more subtle motor deficits. The individual’s
motor functions rapidly deteriorate with disease
progression and, by the age of only 10 years old,
children with DMD are usually wheelchair-bound.
Eventually the heart and breathing muscles are
compromised, leading to a premature death
from heart or respiratory failure. The best
available treatments are palliative, or else aim
to delay onset and slow progression, as there is
currently no cure. The average life expectancy of
DMD patients is only 25-30.
Despite this dearth of effective treatment
strategies, a fair amount is known about the
genetic and molecular pathology of DMD. The
gene responsible for DMD, dystrophin, was
identified over 20 years ago. The dystrophin gene is
2.4 million bases long – the largest found in nature
to date – and contains 79 exons that take over 16
hours to be transcribed and spliced. Dystrophin
is part of a large complex of proteins linking
the subsarcolemmal actin cytoskeleton with
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extracellular membrane proteins. This structure is
thought to be important for muscle health, since
many dystrophies are the result of mutations in
genes that encode members of this complex.
One of the leading theories of DMD pathology
is known as the ‘membrane fragility hypothesis’.
This posits that dystrophin plays a primarily
structural role, providing stability to the
membrane and protecting it from damage that
can be induced during the natural process of
muscle contraction. It is thought that in the
absence of dystrophin the membrane might
be more easily damaged. More recent studies,
however, have revealed that this theory is too
simplistic and certainly not the full story.
EXISTING STRATEGIES
Several genetic treatments are being developed
for use in DMD. These therapies target the
faulty gene specifically, either by correcting
the genetic mutation through gene therapy or
using a technique known as exon skipping. Gene
therapy involves the introduction – usually by
viral vector – of a healthy copy of a diseaserelated gene into the cells of a patient, where it
can restore healthy function. Despite its promise
as an effective treatment for many genetic
disorders, gene therapy for DMD is problematic
because the dystrophin gene is too big for the
development of conventional viral vectors.
Methods to develop a smaller equivalent gene,
called ‘micro-dystrophin therapy’, have also
resulted in compromised dystrophin function
and are therefore not fully effective.
Exon skipping is another, similar strategy, which
is achieved using an antisense oligonucleotide
that binds to the defective region of the target
gene, forcing the cell to skip this section of
the precursor mRNA during translation. This
treatment is also a compromise, since it does
not provide a fully functional copy of the gene,
but nonetheless it could be a large step towards
effective treatment of DMD. “Gene therapy and
exon skipping may improve the DMD pathology
to be more like Becker muscular dystrophy
– usually a less severe condition,” explains
Professor Stanley Froehner, a DMD expert from
the University of Washington. “This would be an
enormous accomplishment”. Despite the future
promise of genetic therapies, there is a need for
alternatives that can be more readily delivered
to patients and at a much lower cost.
SHIFTING THE PARADIGM
In an effort to find an alternative to the
widely accepted, yet questionable, membrane
fragility theory, Froehner and his team have
pursued the idea that the dystrophin complex
is intimately linked with a variety of muscle
cell signalling mechanisms. He suggests that
the dystrophin complex can act as a molecular
scaffold, which facilitates the organisation of
signalling proteins in the muscle membrane.
Froehner’s team has been building substantial
INTELLIGENCE
REPURPOSING DRUGS FOR
DUCHENNE MUSCULAR DYSTROPHY
OBJECTIVES
• To explore the dystrophin complex as a
scaffold that organises signalling proteins
on the muscle membrane
• To repurpose existing US Food and Drug
Administration (FDA)-approved drugs
that target these signalling pathways for
the treatment of Duchenne muscular
dystrophy (DMD)
Despite the future promise of
genetic therapies, there is a need
for alternatives that can be more
readily delivered to patients
evidence to back this up, having discovered a
family of signal transducing adapter proteins
called syntrophins, which associate with
dystrophin and target several classes of
signalling proteins to the sarcolemma (muscle
fibre cell membrane).
These signalling proteins include ion channels,
enzymes, transporters and G protein-coupled
receptors – key mediators of many pathways.
The alteration or loss of these signalling
pathways is likely to have many downstream
pathological effects and may therefore play a
significant role in the pathology of DMD. If this
is true, then it represents an extremely exciting
finding for the field. Signalling pathways provide
an accessible route for the development of
novel treatments.
Froehner is now shifting his focus towards the
development of treatments for DMD. Drugs
that restore these pathways could be rapidly
implemented clinically, especially if such
treatments arise through the repurposing of
existing drugs – something that should be
straightforward and inexpensive in the case of
generic signalling pathways. “Although not likely
to be cures, these drugs could improve quality
of life and lengthen survival, thus extending the
time for genetic treatments,” Froehner explains.
OTHER AVENUES
This line of research and drug development has
led Froehner to the study of nitric oxide (NO) and
its production by the enzyme neuronal nitric oxide
synthase (nNOSµ). In healthy muscle, nNOSµ
is localised to the muscle membrane through
dystrophin and alpha-syntrophin dependent
processes, but this is not the case in DMD muscle
– a shortcoming that corresponds with reduced
production of NO at the muscle cell membrane.
NO is involved in many signalling pathways, one
of which leads to an increase in the production
of cyclic guanosine monophosphate (cGMP) and
consequent protein phosphorylation. What is
interesting about this pathway is that there are
already clinically effective drugs that target it,
specifically sildenafil and tadalafil (also known
as Viagra and Cialis), both of which inhibit the
phosphodiesterase (PDE)-mediated degradation
of cGMP. These drugs are used for a variety of
ailments, from erectile dysfunction to artery
hypertension, and they represent a promising
therapeutic strategy that can be readily applied
to DMD.
PRECLINICAL TRIALS
The Washington team has been conducting
some preclinical trials of these drugs and found
that they improved both respiratory and cardiac
function. The two drugs are currently being
tested for efficacy in human patients: “So
far, two limited trials with sildenafil have not
demonstrated skeletal muscle improvement,”
reveals Froehner. “However, a clinical trial
of Cialis, a more potent PDE inhibitor with
higher specificity, is currently recruiting DMD
participants aged seven to 14 years.”
Reactive oxygen species (ROS)-mediated
oxidative stress has also caught Froehner’s
attention, since ROS generation is known to
occur in DMD skeletal muscle. The team found
that a pathological upregulation of NOX2 in
DMD leads to excessive ROS production in
skeletal muscle of the mdx mouse. Froehner
hopes that drugs which reduce oxidative stress,
scavenge ROS or inhibit their production
through NOX2 may provide the basis for
effective treatments in the future.
These lines of inquiry are beginning to show
promise in terms of DMD treatment, and
represent an important paradigm shift in basic
DMD research. However, these novel and
repurposed treatments may not be enough
to tackle all DMD symptoms: “Combinatorial
approaches that target distinct pathways
altered in DMD offer a very promising approach
that deserves more attention from both
researchers and clinicians,” explains Froehner.
“The limitations of genetic therapies may be
improved by co-administration with drugs
that target specific signalling pathways.” This
may one day provide realistic hope of effective
treatment for those afflicted by this otherwise
devastating condition.
KEY COLLABORATORS
Marvin Adams, PhD; Nicholas
Whitehead, PhD; Min Jeong Kim,
PhD; Kenneth Bible, DVM, University
of Washington, Seattle, USA • Kathryn
R Wagner, MD, PhD, Kennedy Krieger
Institute/Johns Hopkins, USA
FUNDING
National Institutes of Health (NIH)
Raymond and Beverly Sackler Foundation
Parent Project Muscular Dystrophy
CONTACT
Professor Stanley Froehner
Raymond and Beverly Sackler Professor
and Chair
Department of Physiology and Biophysics
University of Washington
1705 NE Pacific Street
HSB Campus Box 357290
Seattle
Washington 98195
USA
T +1 206 543 0950
E [email protected]
http://depts.washington.edu/dmdlab/
http://linkd.in/1yRK1Vp
STANLEY C FROEHNER is the Raymond
and Beverly Sackler Professor and
Chair of the Department of Physiology
and Biophysics at the University of
Washington, Seattle. Previous to this
he was Professor and Chair of Cell and
Molecular Physiology at the University
of North Carolina at Chapel Hill and
Professor of Biochemistry at Dartmouth
Medical School (from 1978-92). He has a
PhD in Biochemistry and Neurophysiology
from the California Institute of
Technology. Froehner has served as a
visiting professor in Australia and France
and was recognised with an NIH Javits
Neuroscience Investigator Award.
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