Funded By:
Good morning. It's a pleasure to be here. It's very enjoyable
and friendly to be able to come here and address this particular
conference each and every year. Today, the topic we have:
Is research the answer to neuromuscular disease? First of
all, I'd like to go ahead and define what it is we mean by
"neuromuscular disease." What is it that we have
in common?
This is a picture of the lower motor unit of the neuron from the spinal cord down into the nerve all the way to its connection to the nerve in the muscle, and then into the muscle itself. And what we mean by "neuromuscular disease" is any problem or disease affecting this lower motor neuron, anywhere from the spinal cord which we would see with motor neuron disease, such as ALS, spinal muscular atrophy, where we are losing these motor cells in the spinal cord. It could be nerve problems such as in Charcot-Marie-Tooth disease, a problem with the connection between the nerve and the muscle, as in Myasthenia Gravis, or a problem with the muscle itself as is seen in muscular dystrophy.
There are a number
of potential causes of neuromuscular diseases. Most of these
disorders are genetic in origin, or hereditary, but there
are also acquired causes as well -- immune related, auto-immune
related causes, infectious causes, or environmental factors.
The genetic causes and acquired issues sometimes interact.
Occasionally individuals have a genetic predisposition to
an acquired problem, such as genetic predisposition to some
sort of environmental factor.
Well, first I'd like
to go into some recent advances with Amyotrophic Lateral Sclerosis,
or ALS, which is a motor neuron disease. There have been tremendous
advances recently in our understanding of the causes and the
mechanisms behind ALS. We now know that there are genetic
factors involved. There is a familial form of ALS, which includes
about 10% percent of all ALS cases. There recently has been
greater understanding in regards to the role of a substance
called glutamate, which is the primary excitatory nerve transmitter
in the central nervous system, which is the chemical messenger
by which nerves communicate with one another. Also, oxidative
stress seems to play a role in ALS. It could produce free
radical formation. Aging and the whole concept of our motor
neurons in the spinal cord having a genetically pre-programmed
life span, and there is a genetically pre-programmed "death"
than can occur, and then this may play a role in it as well.
And also, there is possibly a diminished factor we refer to
as neurotrophic factors, which are chemical substances that
circulate in the body and help maintain the health of nerves
and muscles.
Question: What was the term "oxidative stress," I believe you used?
Answer: The concept
is that with diseases in the central nervous system, oftentimes
through oxidative metabolism, there can be free radicals formed
which can actually themselves be damaging to tissues. And
so, that involves the use of anti-oxidants for some of these
conditions. Obviously, there are more potent anti-oxidants
than vitamin E, but many clinicians who are treating ALS patients
now are at least recommending their patients be on reasonable
doses of vitamin E, pretty benign treatment obviously, as
a potential adjunct from the other therapies that they're
on.
Comment from another physician in the audience: I'd like to talk about the oxidative free radicals that are well-demonstrated in traumatic brain injury & spinal cord injury and are, at the animal model level, ways of preventing breakdown in the cell wall in brain tissue, in spinal cord tissue, and probably in muscle cell tissue itself. The groups of anti-oxidants which are called lazaroids, one of the steroids, which are currently being used, at least in a lot of cellular injury, both in brain tissue, spinal cord, and hopefully will be used in muscle tissue too, that in animal models seem to protect against the damage done by these peroxidase oxidants that occur in a variety of muscle diseases.
Glutamate, Riluzole, Gabapentin.
Now, with our greater
understanding in this, has actually led to treatment for ALS.
I'd like to describe now the role of this chemical called
glutamate in ALS. Glutamate, as I mentioned, is the primary
nerve transmitter, the spinal cord transmitter, in the brain.
Glutamate levels have been found to be elevated, abnormally
high, in blood, in spinal fluid, and in brain tissue in ALS
patients. And we know that there is reduced clearance of glutamate;
the glutamate hangs around for too long in ALS patients, in
these critical motor areas of the central nervous system.
We also know that there is a binding protein that binds onto
glutamate called glutamate transfer protein, which is reduced,
so that probably influences the clearance of the glutamate
itself. This has led to the concept of actual glutamate inhibition
for the treatment of ALS, and this has led to two recent medications,
Riluzole, which is now FDA approved for the treatment of ALS,
and gabapantin, which is also on the horizon for the treatment
of ALS. Now with respect to Riluzole, it acts by inhibiting
glutamate release in the nervous system. It has a protective
effect, and in some animal studies, it protects motor neurons,
in some animal models from the toxic effects of glutamate
or glutamic acid. And there have now been two very well done
studies, two well controlled studies, large studies in ALS,
which show the treatment with Riluzole has led to increased
survival. It's a very small and modest increase, but there
is an increase in survival of several months, and also a slower
rate of deterioration in the brain.
Now, as with any medication,
of course there are side effects of Riluzole, and fatigue,
and drowsiness can be a problem, dizziness, abdominal pain,
diarrhea, pneumonia, problems with lung function itself. But
in general, Riluzole was very well tolerated, and in the study,
only about 14% of subjects or patients discontinued the medications
due to adverse effects. It was actually very well tolerated.
The next medication
that is on the horizon is gabapentin, which is actually FDA
approved, but for a different use. It's used for seizure control,
it's used for nerve pain in some individuals, and it also
acts with a similar but slightly different mechanism as Riluzole.
There have been some recent studies, a Phase II trial, which
have provided encouraging results with regard to gabapentin,
a slower rate of decline in arm strength. This medication
has been very well tolerated with few side effects.
You have probably heard
in the press about the use of anti-oxidants in ALS, and the
idea here is that there could be some oxidative stress factors,
free-radical formations, which might be important in the beginning
of this disease, or disease initiation. And actually, in some
animal models for ALS, the institution of vitamin E, which
is an anti-oxidant, actually delays the onset of ALS in this
particular mouse model. And there are many physicians around
the country in ALS clinics who treat ALS patients who actually
recommend that their patients be taking vitamin E.
Now, neurotrophic factors
in ALS treatment. There are a number of that are on the horizon:
insulin-like growth factor I, or myotrophin; another called
brain-derived neurotrophic factor, and then another one ciliary
neurotrophic factors, CNTF. Now, these neurotrophic factors
essentially circulate throughout the body and help maintain
the health of the nerve and the muscle itself.
Now, first of all,
IGF-1, or myotrophin, insulin-like growth factor 1: it's naturally
occurring, our body actually produces insulin-like growth
factor 1, so it's a natural substance. It's synthesized in
the muscle, and in the brain and the central nervous system.
There are receptors that we think pick up IGF-1 in motor neurons,
in the spinal cord, in the muscle, and also in the brain.
And some very elaborate animal studies have shown that the
IGF-1 actually enhances nerves and nerve regeneration in sprouting
after the nerve has been disconnected from the muscle, after
denervation. So, it enhances neuronal regeneration. Also,
IGF-1 seems to reduce muscular atrophy after denervation.
So this is actually very encouraging. It's taken similar to
insulin by means of a subcutaneous injection. Now there has
been a double blind placebo-controlled study in ALS for IGF-1,
and the preliminary studies indicate that a high dosage of
IGF-1 produces slowing in functional decline, slowing in loss
of bulbar functions, slowing in loss of breathing capability,
slowing in deterioration of muscle strength and arm and leg
function. The improvement seems to be dose related: the higher
the dosage, the better the outcome. There was another study
in Europe which had similar results, but not quite as impressive
in terms of these clinical backups. In terms of side effects,
again this is a naturally occurring substance, so the primary
side effects were that of injection site pain recorded by
about 70% of individuals, but there were really few other
side effects.
Now, brain-derived
neurotrophic factors, BDNF. This shows also great promise
in experimental models. Phase I trials, just looking at toxicity
related issues, have been completed. Some initial Phase II
trials have been completed, and it's showing slowing in the
spine and pulmonary decline in this randomized, double-blinded
trial. The optimal dose for this yet has not yet been determined,
but again, this is looking quite promising.
If you have a neurotrophic
factor, obviously you want the medication to be delivered
at its primary site of action. This would both minimize the
side effects of the medication and also increase the effectiveness
of the medication as well. And this brings up this new concept
now of "direct spinal fluid delivery" of some of
these neurotrophic factors, which I think is on the horizon
of ALS treatment. The idea is that, and in particular the
last one is that, the CNTF, ciliary neurotrophic factor would
not be tolerated when patients took it orally. There were
far too many side effects, and in fact, there was even some
increased mortality, so really oral trials were completely
discontinued.
But the idea that we
can deliver the medication at its site of action, we can minimize
the side effects because there would be less diffuse circulation
of the medication throughout the body, and also we can increase
the effectiveness, and this is why this concept of "spinal
fluid delivery." Now this sounds it might be like a "pie
in the sky concept": How will we get this into a spinal
fluid, but in actuality, there is currently an infusion system
which is commercially available that we put in patients around
the country on a daily basis for other reasons, for spasticity
management, for pain management. And there is an infusion
pump, which actually can be implanted under the skin, the
catheter can be tunneled into the spinal canal, and the medication
can actually be dripped in or infused into the spinal canal.
There are already some early trials using this infusion system,
which are underway in Chicago, looking at the infusion of
some of these neurotrophic factors in and around the spinal
cord to try to prevent the deterioration of the motor cells
within the spinal cord.
In our other experience
with the use of this pump, for another indication, spasticity
management, what we have found is that actually by delivering
the medication in this means, at least with this other medication,
spinal levels up to fifty times higher can be achieved with
one one-hundredth (1/100) of the oral dose. So we can give
one one-hundredth of the dose, yet get fifty times the medication
in the spinal fluid. So people are really quite excited about
this: the dosages itself, and the infusion rate, can actually
be adjusted by use of a regular telemetry unit, communicating
with the pump itself. So I think this is on the horizon, initial
trials are already underway, and we anxiously await the results
of that. The pump itself can be refilled by a needle. So I
think with regard to the future of treatment for ALS, at last
we have optimism and great promise. I think that ultimately
the approach to the treatment of ALS is going to focus on
combination therapy using a variety of drugs with a variety
of different mechanisms, and I think that when somebody sees
their physician, they are really going to be faced with a
variety of different agents which have very different mechanisms,
but the goal is the same end result. So the physician ultimately
may be approaching this very much like we would approach a
patient with cancer, using a variety of different chemotherapeutic
agents, some of which might act in unison, synergistically,
to actually enhance the effectiveness of each other.
On to some of our genetically
caused neuromuscular diseases: these are abnormalities in
genes which ultimately create abnormalities in proteins. Proteins
then can be abnormal in structure or completely absent altogether,
and since I joined the faculty in 1992, there has been literally
an explosion in advances in molecular genetic information
on these disorders. And it really has become quite staggering
and very difficult to keep up with, as a matter of fact. As
of last count in terms of the recent advances in the molecular
genetic disease locations in neuromuscular disorders, the
gene is actually been located or identified in 86 of these
disorders now. Eighty-six neuromuscular disorders. We have
40 other mitochondrial disorders causing problems with brain
and muscle, with which gene abnormalities have also been identified.
So years ago, when I gave this talk, I think our count was
about 30 conditions. Now we're up to 86 conditions where the
gene has actually been identified.
I just want to go through
some examples of spinal muscular atrophy, first of all. There
are a variety of types of spinal muscular atrophy. The most
common type primarily involves the shoulder and hip musculature,
and it's called predominantly proximal spinal muscular atrophy
with types I, II, and III. There are a variety of other types
of spinal muscular atrophy with autosomal recessive inheritance,
where you would need two copies of the gene from both parents.
There are some autosomal dominant varieties of spinal muscular
atrophy, x-linked spinal muscular atrophy, and sporadic spinal
muscular atrophy as well with known inherited factors. Now,
the more common type of muscular atrophy is caused by gene
abnormality in chromosome 5. So what has been quite interesting
with this particular condition is that it's not just one gene
which may be involved; there seems to be at least three genes
which may be, at least in part, responsible for this condition.
NAIP, Survival Motor Neuron, Basal transcription Factor.
Now, to summarize,
in 1990, all three forms of spinal muscular atrophy (type
I, type II, type III) were all mapped to the same chromosome
at the same gene locus. It took really quite a long time to
work out the specific gene because the complexities in the
genetic material at that locus, but in 1995, a group from
France first reported the presence of the survival motor neuron
gene. Subsequently, later that year, shortly thereafter a
Canadian group reported a second gene seeming to be associated
with spinal muscular atrophy, called NAIP, or neuronal apoptosis
inhibitory protein, and then more recently (this was in the
last few months), a United States group just identified a
third gene, basal transcription factor II, BTF2 gene, which
may also play a role in spinal muscular atrophy. With regard
to the spinal motor neuron gene, this gene is abnormal in
93% to 98% of individuals with SMA I, II, and III, really
providing very strong evidence that this is a disease determining
gene, the survival motor neuron gene. Because of this, we
now have a very active diagnostic test available; we can test
parents, siblings, other family members for carrier status.
The protein that the gene codes for is yet to be identified.
The NAIP gene seems to be present in fewer percentages of
the spinal muscular atrophy population. Forty-five percent
of SMA I, eighteen percent of SMA II and III, and this protein
that this gene codes for seems to be involved in this concept
of genetically programmed "cell-death" that occurs
in the spinal cord. And normally there is in inhibition of
this motor neuron "cell-death," but this seems to
be altered in spinal muscular atrophy.
So we now have a much
better understanding in terms of what leads to the quite varied
differences in severity from child to child with spinal muscular
atrophy. What explains this incredible diversity of severity
(diversity with regards to the progression of which these
children and adults develop their problems)? And this had
brought up the concept of a multiple gene model, or a multiple
allele model for SMA, and the idea that there are multiple
genes, (and this again might be applicable to other neuromuscular
disease conditions as well), the idea is that the varied SMA
genes may actually act in concert to produce the varying differences
in severity. The possibility of a single gene, the survival
motor neuron gene, which acts primarily to determine the disease,
and maybe the other genes modulate the severity. As an alternative,
there may be different genes which code for different proteins,
and then those proteins then interact to form a common structure,
or a large protein complex. The other possibility is that
perhaps there are more severe genes, or severe disease alleles,
which are paired then as mild-disease alleles. You could bring
up the concept that maybe SMA I of a severe gene allele paired
with another severe gene allele; SMA II has a severe allele
paired with a mild allele; and then SMA III might be a mild
allele I paired with a severe allele II, or mild with a mild,
or perhaps a severe allele I paired with a completely normal
allele II, and this brings about this varied pattern of progression
of this disorder.
Friedreich's Ataxia, Frataxin.
There are many other
disorders where the gene has now been localized, and there
is active research being done to determine the protein that
that gene codes for and also the function of that protein,
which is really the first step in determining treatment. And
now with Friedreich's ataxia, this has been localized to chromosome
9; the protein has been identified as Frataxin. There is also
a different form, a very rare form of Friedreich's ataxia,
with selective vitamin E deficiency, that is from chromosome
10. The vitamin E deficiency really needs to be ruled out
in any Friedreich's ataxia agent.
Charcot-Marie-Tooth
peripheral nerve degeneration: there is a peripheral myelin
protein which coded for by a gene from chromosome 17. Now
what's been very informative about this particular condition,
is here is a situation where normally, we have two copies
of this gene at that region. In Charcot-Marie-Tooth, type
IA, which is the most common form of Charcot-Marie-Tooth,
we actually have three copies of the gene at that region,
so there is overexpression of the gene. But there is another
disorder where individuals are predisposed to getting pinched
nerves at various locations, where they have an actual deletion
at the region. So in one instance, an overexpression of the
gene leads to disease; in the other instance, an underexpression
of the gene leads to disease. And so, this really has added
to the complexity because it's not just a matter of getting
as much of the gene back in as possible. We have to get the
correct amount of the gene back into the tissue in question.
Now, really rapidly and very complex because we used to think
that there was a demyelinated form of Charcot-Marie-Tooth
called type I. Well, now we know that there are different
genes which cause different protein abnormalities and actual
different forms of Charcot-Marie-Tooth type I: type IA, IB,
and IC. Last year when I presented this information, we had
Charcot-Marie-Tooth type 2A and we had type 2B. Well, this
year, we now have type 2A, type 2B, type 2C, and we have type
2D that just came out. And so we have multiple differences
in gene abnormalities which can cause conditions which can
look very similar, but obviously the proteins that are involved
are probably quite different. There's also some recessive
Charcot-Marie-Tooth type IV, but it's not quite so simple.
We have type IVA now, chromosome 8; type IVB, chromosome 11;
probably we have a protein even for type IVC that has not
been identified as of yet, but again different gene abnormalities
causing different protein abnormalities causing a disease
which looks fairly similar to this condition. And there is
an X-linked CMT as well, and again, of course, needless to
say, we have X-linked type I, type II, and type III. So this
is really getting incredibly complex, for Charcot-Marie-Tooth
is not just one or two different conditions; we're talking
about 12 or 14 different conditions caused by different gene
abnormalities with different protein abnormalities.
Question: How hard is it to determine the type of CMT that one has?
Answer: That's
an excellent question. We now have well over fourteen different
types of CMT, and what type does my child have or what type
of CMT do I have? Unfortunately, there are only a couple of
the varieties of CMT which can actually be elucidated through
commercially available tests. And the CMT Type 1A is one of
them, the HMTP; there is one of the other varieties, the 1B
can also be determined as well. I think this has brought up
an important question, and with all this genetic research
that has come about, theoretically, it might help us considerably
in determining what specific sub-type of a neuromuscular disease
condition an individual has. Theoretically, it has implications
for family counseling, and so forth. Currently, the actually
sub-typing of the disorder such as Charcot-Marie-Tooth is
not changing our management, in terms of how we treat patients,
in terms of medications that might be tried, or supportive
rehabilitation strategies that are used. Now, in the era of
managed care and health care reform, third-party providers
are oftentimes unwilling to pay for laboratory investigations
and studies which don't ultimately change the specific management
of the individual. I think certainly if gene transfer therapy,
gene therapy becomes available in the future, this will be
of critical importance. The other point I want to make is
that oftentimes, commercial laboratories are set up to receive
specimens, process them rapidly and efficiently, and get the
results back to the physician and the patient. Oftentimes,
we end up sending on specimens to research laboratories to
help further the research in a particular disease process.
One thing we try to do is to provide realistic information
to the patient and to their families, but this particular
test, this blood test may not lead to any useful information
whatsoever. They may not find any specific gene abnormality.
But oftentimes this can be a source of frustration to the
patient who has then submitted a sample for research investigation.
Oftentimes, I'll have to tell you this, the researchers at
these various universities try to collect batches of specimens
so that they can run them all at once in a more cost effective
manner. Well, they may be handling these specimens, keeping
them in a refrigerator for 12 months or 18 months until they
get enough of these specimens to run all at once. Some of
it is being used primarily for research at this point in time.
Obviously, if drug treatments become available which are very
specific to different sub-types of these disorders, it's going
to be very, very critical to identify the specific sub-type.
Other questions? That was an excellent question.
Question: I have one more. When it's hereditary, does that mean that the father and daughter will have the same type, or will they be different types?
Answer: In terms
of the sub-type, in terms of whether or not we're talking
about a 1A sub-type or 1B sub-type, generally it will be the
same type. But within those sub-types, there can be tremendous
variability in degree of severity. There's not necessarily
one-to-one correspondence in that. Another question over there?
Fascioscapulohumoral Muscular Dystrophy.
FSH, fascioscapulohumoral
muscular dystrophy, has now been recently localized at chromosome
4; the protein has not been fully characterized, but there
is some very exciting work being done in this region and they
are very close to sequencing the particular gene.
Myotonic muscular dystrophy
has been localized in a different location, chromosome 19,
producing this particular protein. This is a situation where
we have unstable DNA with an abnormal amount of repeated segments
of DNA, and in this situation with successive generations,
maternal transmissions to child and so forth with each successive
generation, we oftentimes see this disorder getting progressively
worse. And the reason is because of these expansions, the
expansion in this particular gene region, the expansion of
the connective material gets bigger and bigger with each successive
generation.
Question: Yeah, what's the difference between myotonia congenita, and myotonic muscular dystrophy?
Answer: All
those disorders actually have problems with relaxation of
muscle tissue, but myotonic muscular dystrophy has a dystrophic
component to that disorder, meaning that there's actual loss
of muscle tissue occurring over time, whereas that's not the
case with myotonia congenita, or some of the others. There
are a really a variety of different myotonias, which probably
have different genetic causes, different protein abnormalities.
But really the main two differences between those disorders
have to do with severity of progression over time. Oftentimes
myotonia congenita can have real problems with relaxation
at very cold temperatures, even more so than dystrophic patients.
But the other individuals are actually slowly, very slowly
losing some muscle tissue over time. And that's really what
we mean by the term "dystrophy." Other questions?
Question: In myotonic muscular dystrophy, can it have a very rapid progression?
Answer: Generally,
it's a very slow progression. If it's rapid, it is generally
in the congenital myotonic muscular dystrophy form, different
from myotonia congenita. And that oftentimes will present
in the delivery room with infants having difficulty breathing
and requiring ventilatory support from the get-go, and they
oftentimes have a more rapid degree of progression. But in
general, myotonic muscular dystrophy has a very slow progression.
Question: Well, it just seems like, for my son, I see him just deteriorate before my very eyes. And I have one more question: In the breathing for myotonic muscular dystrophy, is it normal that he will have hiccups for weeks at a time, is that normal? Is that expected or common?
Answer: Hiccups
per se, I don't think are necessarily that specific to myotonic
muscular dystrophy, I don't think so. But certainly there
can be some respiratory breathing compromise in selective
individuals with myotonic muscular dystrophy, specifically
with childhood-onset, more so than in those with adult onset.
Yes?
Question: In the myotonic muscular dystrophy, is there any kind of medicine to keep from advancing the disorder? Is there any kind of prescription available?
Answer: Most
of the medications that have been used, such as dilantin,
have really been geared at systematic treatment of the myotonia
itself and haven't been geared towards changing the actual
progression of myotonic muscular dystrophy or the disease
progression itself. There currently isn't any available treatment
which has been shown to actually impact the natural history
in progression for myotonic muscular dystrophy. Now, the fellow
back there mentioned an agent which, I understand, is under
investigation at Rochester, but at least in my reading of
literature, there's not many agents yet that have actually
impacted the rate of progression.
Question: The question of myotonic muscular dystrophy, if you have an accident, or something, could that make it rapid? Like if you fall, or you had some kind of surgery?
Answer: Well,
I think oftentimes when we speed muscular dystrophy towards
dystrophic conditions, if there is an accident causing immobility
for a prolonged period of time, there can perhaps be a little
bit of acceleration with regard to weakness. If there is immobilization,
perhaps the joints can become more stiff from contractures.
And certainly any immobilization such as that can certainly
have deleterious effects on the condition. That might be perceived
in the individual being sort of a rapid, more rapid progression
of the actual disease. I think it's just a complication of
these immobile or developing contractures in the joints.
Duchenne and Becker Muscular Dystrophy.
Now, onto muscular
dystrophy. Our understanding of muscular dystrophy has really
rapidly expanded in the last few years, and this is having
implications now for Limb Girdle muscular dystrophy, as well
as Duchenne muscular dystrophy, Becker muscular dystrophy,
and other forms of muscular dystrophy. This is the current
working model of what is causing dystrophin glycoprotein complex,
and this dystrophin lies on the inside of the muscle cell,
the muscle membrane or the muscle tissue. There are a variety
of proteins which this dystrophin binds onto, and these dystrophins,
these proteins, have actually been characterized fairly well.
These proteins then bind onto another protein which is in
the extracellular face of the extracellular matrix. And really,
abnormalities in any of this whole mechanism can produce muscular
dystrophy. This has given rise to the concept of diseases
of the dystrophin glycoprotein complex. The most common of
these being due to an abnormality in dystrophin, causing Duchenne
muscular dystrophy and Becker muscular dystrophy, so we now
have all these other previous proteins. Any one of them, if
they're abnormal, can produce a different form of muscular
dystrophy. Namely, many of Limb Girdle muscular dystrophies.
Question: I know you want to stay away from racial background, but are certain kind of muscular dystrophies more common among certain races?
Answer: That's an excellent question. I think many of the neuromuscular disorders do have racial predilections to some extent. There are certain populations around the world that have fairly high incidences of certain genetic conditions. That really varies with the particular condition. There are some conditions which have a particularly high incidence in Eastern European populations, there are some others that have low incidence among African American populations, higher incidence among Caucasian populations, and so forth. But much of that has been worked out and described by a researcher named Emery in a journal, Neuromuscular Disorders, which gets into some of the ethnicity of some of these conditions. A pretty good question, other questions?
Limb Girdle Dystrophy, Congenital Muscular Dystrophy, Merosin.
And we now know that
with these various proteins that attach onto dystrophin that
cross that membrane, any number of these proteins, if abnormal,
can produce a varied group of Limb Girdle muscular dystrophies.
This protein out here, laminin, also called merosin, if abnormal,
can produce a congenital form of muscular dystrophy, which
presents in the newborn or in early infancy. This merosin
deficiency can lead to congenital muscular dystrophy. There
is even an enzyme, protein enzyme, which if abnormal, can
produce another form of Limb-Girdle muscular dystrophy. This
enzyme is called calpain. So, we used to have a concept of
a Limb-Girdle muscular dystrophy; now we know there are autosomal
dominant varieties of Limb-Girdle muscular dystrophy -- different
genes, probably different proteins. And now we're up to seven
different sub-types of Limb-Girdle muscular dystrophies: types
IIA through IIF, all with different gene abnormalities producing
different protein abnormalities, producing a muscular dystrophy
of varying degrees of severity.
This brings up the concept of "Why is it so important to determine the location of all these genes?" Well, first of all, once we determine the gene location and sequence the gene, we can then determine what protein that gene codes for. And that's really the first step in determining the important role of that particular protein and its function, and determining the reason for which an abnormality in that protein then leads to the establishment or the progression of the disorder. Once we understand more about the mechanism of the various diseases, that's really the first step in determining rational treatment: drug therapy or pharmacological therapy. There is also the concept of gene therapy, which you hear a lot about in the news media, and the idea here is that the goal is to try to replace the defective gene with a normal copy of the gene, or a fairly normal copy of the gene, in the particular tissues that are affected. It is the muscle usually in muscular dystrophies, but it might be muscle, it might be spinal cord, it might be brain tissue, depending on the problem. There are a number of requirements for gene therapy: sufficient recombinant gene technology, which we have around the world now; the gene expression, the reading of the gene must be sufficiently understood. We have to identify what the particular gene is and its entire reading sequence. We have to have some kind of understanding about what is the mechanism of the disease, be it in the spinal cord or in the muscle. We have to have some understanding of pathogenesis, or the mechanism by which these disorders are produced. We have to understand what the target tissue is, where we need to get the gene if we're going to try to replace the gene: Do we need to get the gene back into muscle? Which muscle? Do we need to get it back into heart muscle or skeletal muscle? Do we need to get it back into spinal cord tissue or brain tissue? And then, we have to look at ways in which we can actually transfer the gene into these target tissues. And then, finally once the gene is there, we have to ensure there is sufficient or appropriate reading of the gene, or gene expression, for appropriate required amount of time, which could be for the rest of the individual's life.
Now there are obviously
some obstacles to gene transfer therapy, which I want to touch
on. This isn't just a matter of injecting the gene into the
blood stream, where it goes to the right place, and gets incorporated
and starts working. The body itself has immune responses to
foreign proteins and to foreign particles, foreign genes.
The body has responses to the viruses which are being used
to carry these genes. So there is a need for immunosuppressant
medications oftentimes. The size of the gene can be a limitation.
The gene that causes Duchenne muscular dystrophy is the largest
gene that has been identified in the human genome. It's a
very, very large gene, and there is some limitation in the
ability to attach that gene onto a particle that carries the
gene around the body; there's limitation in the size of the
gene that can be carried. Also, in delivery of the gene, we
have to get the gene into the specific location (i.e. muscle
tissue, in the case of muscular dystrophy or another muscle
disorder), and then once the gene is there, we have to worry
about getting it from outside the cell to the inside of the
cell, where the genetic reading takes place. And then with
regard to gene expression, as in the case of that one form
of Charcot-Marie-Tooth where overexpression of one gene leads
to one disease, and underexpression of the gene leads to a
different disease, we have to make sure that this gene is
read and expressed in the proper amount because we want to
ensure that there is not underexpression or overexpression.
Because theoretically, we could be creating new disorders,
new diseases.
With regard to gene
transfer therapy, I wanted to review with you some of the
current research approaches underway to deal with the various
problems. Problem #1: In gene transfer therapy, we need some
sort of "bug," or some sort of transportation vehicle
that can carry the gene around the body to the particular
location. We need a transmission vector for the gene, to deliver
the gene. And what has been used, what's currently being looked
at is this concept of viruses, using viruses to help us carry
these genes around the body. Secondly, the gene must be known,
it must be sequenced, it must be of sufficiently small size
to be carried by these transportation vehicles, and with recombinant
DNA technology, we actually create "mini-genes,î which
are not the full copy of the gene, but are a smaller copy
of the gene which contains the critical genetic material needed
to produce a functional protein. And then, these viruses,
if carrying these genes, may be recognized and destroyed by
the body's immune system, so the idea now is that people doing
this research must do modifications to the virus itself, so
the virus won't be recognized as a foreign particle and then
eliminated. And sometimes actual manipulations in the virus
itself is the genetic apparatus we made, or even little particles
can be attached to the virus to keep it from being recognized,
and these are called stealth particles, which I think is a
great term, stealth particles attached to the virus to keep
it from being recognized. And then obviously, we use immunosuppressant
medications to try to keep the major inflammatory reaction
occurring in the tissue itself, which can be very disastrous
and deleterious to the individual. Then, the transport, transportation
vehicle, the vector must travel throughout the body, and it
needs to get to the tissue where the disease occurs. These
get to the specific target tissue. If we're talking about
getting dystrophin into the target tissue, we need to get
the dystrophin protein into muscle. We don't want dystrophin
to be expressed in the pancreas or in the spleen; we have
to make sure that the dystrophin is being expressed in the
diseased tissue. We have to use an appropriate transportation
vehicle with an attractant to the diseased tissue.
And this is why research
is looking at particular viruses, adenoviruses, which have
an attraction to muscle tissue in particular. And then, finally,
once the gene is transported to the cell itself, the gene
has to get inside the cell, and sometimes it can get trapped
in the transport systems. There are other manipulations that
are being looked into that I don't want to get into detail
here. But one important concept is expression of the gene
in the appropriate tissue, muscle, and not in other tissue
by way of an on/off switch. What we do is attach to the gene
itself an appropriate on/off switch so that the gene is turned
"on" in muscle tissue, and the gene is turned "off"
in other tissue, such as the pancreas, or the spleen, or other
tissue, where we don't want the gene expressed. And so we
attach to the gene then, what are called tissue promoters
to turn "off" and "on" the gene. Now again,
once the gene is there, if the gene is too big, it might take
up to twenty hours to actually read all the DNA in a gene.
And so that's where the use of a "mini-gene," which
is a smaller version of the gene, becomes really quite important,
for efficient reading of the gene itself. And then once we
get the normal protein back into the tissue, we have to make
sure that we have the appropriate amount of protein in the
tissue at the appropriate location. And this brings up the
issue of the appropriate amount of protein expression, which
is necessary to either stabilize the symptoms or control the
systems, and this is really quite critical. We don't want
too much protein created because that could have harmful effects;
we don't want too little protein created because that won't
have the appropriate effect to stabilize the progression on
the symptoms, and that's really where the importance of animal
models comes in, if we can actually create animal models,
with various amounts of the protein actually produced.
There's one more point
I want to make, if I can. The issue of an, the concept of
a "immune-mediated response." This is where tissue
containing the restored gene product, and the restored protein
is being destroyed by the body. Keep in mind that an individual
with Duchenne muscular dystrophy, their body's immune system,
has never seen the dystrophin protein. So if all of a sudden,
you get this dystrophin protein produced, would that child's
or young adult's immune system start reacting against those
tissues because all of a sudden, this is a new formed protein?
And again, I think the need for animal studies, animal investigations,
is really critical here so that we're not doing more harm
than good by subjecting patients and clients to experimental
procedures which put out deleterious effects.
I just want to review
two recent studies published within the last two months. Last
year we talked about Dr. Chamberlain's work at Michigan, where
they took the full copy of the Duchenne muscular dystrophy
gene and actually injected that gene into an embryo at a four
stage level, and eventually by injecting that gene, the animal
then, which would definitely have muscular dystrophy, grew
up not to have muscular dystrophy. Well, injecting with a
needle a gene into an animal embryo is a far cry from human
treatment, so the next step now has been what sort of actual
transportation vehicle can be used to try to get the gene
back into the tissue? And Dr. Chamberlain's group came up
with the concept of EAM, encapsulated adenovirus mini-chromosome,
and these mini-chromosomes actually have a much bigger gene
carrying capacity than the viruses, the adenovirus itself.
So, the adenovirus is the transportation vehicle with an increased
carrying capacity. The other thing that they have done is
to completely eliminate the genetic content of the virus itself.
And now, the virus is not recognized as a foreign invader,
which would require elimination of the foreign gene, the adenovirus
gene. Again, keep in mind -- this research is still at the
laboratory level. They are basically taking muscle tissue
in a dish and injecting in the virus carrying the gene, and
so we're still at laboratory level here. They're taking muscle
tissue lacking dystrophin and injecting it, and essentially
mixing it up with this hand-made vehicle, the adenovirus.
They are also using some other viruses that have been necessary
because now that we've eliminated the virus's genetic machinery,
it can't reproduce itself, so the problem becomes how can
we get enough of this produced in the laboratory to give to
a patient? And so now we're using what are called "helper
viruses" to help grow the gutted stealth virus in the
laboratory. Now, the "helper virus" is going to
have to be removed from the stealth virus and the gutted virus.
And there are still are those small individual amount of "helper
viruses." But the results are very, very encouraging.
We got the abnormal protein, dystrophin, back into the tissue
-- that has been confirmed.
Another study at a
different center in Baylor essentially used very similar technology
and got very similar results. We have two independent groups
who have come upon the same technique: one at Michigan, one
at Baylor, and they have produced very similar results, which
is really quite encouraging with regard to gene therapy. This
gene therapy technique could conceivably be applicable to
any of the neuromuscular disorders, not just Duchenne muscular
dystrophy.
So I think to wrap
things up, in conclusion, there has really been staggering
advances over the last two years with regard to research in
neuromuscular disorders. I think for the first time, we have
promising therapy which might impact the quality of life for
individuals with ALS. There are a variety of neuromuscular
disorders where we have found gene abnormalities and the protein
abnormalities produced by those genes' defects. Gene therapy
is on the horizon. I think they are actually moving out into
actual animal models, where they are injecting the viruses
carrying the gene into the animal themselves. And I think
we are probably a few years away yet from actual human investigations.
A lot of that will stand on the results of the animal therapy
and animal experiments.
Question: As far as they're coming out, will gene therapy be limited to specific age groups?
Answer: I think that I don't see gene therapy being limited to specific age groups. I do think that there are going to be some limitations with regard to how far a given individual has progressed with regard to their disease process, and my guess would be that if an individual has lost a tremendous amount of their muscle fibers, the treatment might have more limited abilities to effectively change the natural history in those individuals. None of that has really been worked out yet, but I don't see gene therapy being limited with regard to age, in that manner. Now, myoblast transfer, which has recently been shown, even in two-year-olds, not to work for Duchenne muscular dystrophy, that certainly I think, at least shows more promise in younger patients. But with regard to gene therapy, I don't see any age-limiting factor necessarily.
