Mark Thorson
2012-07-31 06:19:45 UTC
A NEW APPROACH TO ALZHEIMER'S DISEASE
Copyright 2012 Mark Thorson (ver. 7/30/12)
This essay describes a new therapeutic approach to
Alzheimer's Disease (AD). This approach is based on the
possibility that the cause of AD is a deficiency of BH4
(a.k.a. tetrahydrobiopterin), which is a cofactor for
enzymes needed to make important neurotransmitters. The
therapeutic agent is something you can make in your kitchen
from materials you can buy without any limits or regulation.
The procedure for making the agent is completely safe and
requires no particular skills nor equipment not normally
found in a kitchen.
The remainder of this essay is divided into five parts:
I. What is the evidence that BH4 deficiency causes AD?
II. What is the best treatment for BH4 deficiency?
III. How can I make sepiapterin?
IV. What results have been obtained?
V. References
PART I. What is the evidence that BH4 causes AD?
BH4 is like a vitamin in its role as a cofactor for enzymes.
A cofactor is a molecule that an enzyme needs to carry out
its function. A neurotransmitter is a molecule released
by one nerve cell to send a signal to another nerve cell.
BH4 is a cofactor for enzymes needed to make several
neurotransmitters: dopamine, epinephrine, norepinephrine,
serotonin, and melatonin. Although the exact way in
which AD affects memory and causes confusion is not well
understood, shortages of these important neurotransmitters
could easily be part of this process.
BH4 is also a cofactor for nitric oxide synthase enzymes,
which make nitric oxide. Nitric oxide relaxes blood vessels
in the brain. Lack of nitric oxide tightens these blood
vessels and reduces blood flow. Nitric oxide also acts as
a neurotransmitter believed to play an important role in
forming long-term memories.
In Alzheimer's disease, the level of BH4 (measured
indirectly as the level of biopterin) in the brain is
greatly reduced. Studies show the level in cerebrospinal
fluid (the fluid which bathes the brain and spinal cord) is
about 30 to 50% lower in people with AD in comparison with
people of the same age without AD.
Other studies show that the level declines dramatically
with age. Between the ages of 20 and 70, the level falls by
about 50%. These same studies also show a wide variation.
Some young adults have levels similar to senior citizens,
and some senior citizens have levels similar to young
adults. This may explain why some people get AD in old age
and some don't -- there may be a threshold level at which
AD occurs, and some people reach that level in old age while
others never reach that level.
BH4 is currently available as a pharmaceutical drug called
Kuvan from BioMarin Pharmaceutical. Currently, Kuvan is
only marketed for treatment of a rare genetic disease.
The first clinical trial of Kuvan in people with AD is
currently recruiting subjects, but it will be some time
before any results are reported. For more information
about the trial, go here:
http://clinicaltrials.gov/show/NCT01439555
PART II. What is the best treatment for BH4 deficiency?
There are several difficulties in treating AD with BH4.
First, BH4 is a very unstable molecule. It is attacked
by oxygen from the air. To make BH4 stable enough to
distribute as a pharmaceutical, it must be in the form of
an acid salt, it must be mixed with antioxidants, it must
be packaged in oxygen-barrier materials with an oxygen
absorber, and it must be kept frozen at very low
temperature.
Second, BH4 is poorly absorbed. It does not easily cross
the blood-brain barrier into the central nervous system.
Third, BH4 is a pharmaceutical drug not available without
a prescription. The only purpose for which BH4 has been
approved by the FDA is treatment of tetrahydrobiopterin
deficiency (formerly known as atypical phenylketonuria).
This is a genetic disease in which one of the enzymes
necessary for the biosynthesis of BH4 is defective.
It is diagnosed in infancy, and its victims usually have
severe mental retardation and short life spans. If AD is
caused by a BH4 deficiency, it differs from the genetic form
in being a milder form which strikes long after the growth
and development of the nervous system is complete.
There is a therapeutic agent for BH4 deficiency which is
better than BH4 in all of these respects. It is much more
stable, though stability is still a problem. It has greater
efficacy in treating BH4 deficiency, which implies it has
better absorption. It is available to anyone who wishes to
make it. This agent is sepiapterin.
Once sepiapterin is absorbed, it is converted into BH4.
Sepiapterin is very similar to BH4, differing only in two
chemical bonds. In the conversion to BH4, these two bonds
are opened up and hydrogen atoms are placed on the free
ends. This change is necessary for the function of BH4 as
an enzyme cofactor, in which BH4 acts as a hydrogen donor,
but the change also makes the molecule very vulnerable to
attack by oxygen.
PART III. How can I make sepiapterin?
Making either BH4 or sepiapterin synthetically would require
chemicals, equipment, and skills which you are unlikely to
have. Also, it would require purification to remove
unnatural forms of these molecules (called isomers) which
have unknown effects in the human body.
However, there is a biological source of natural sepiapterin
you can grow which is free of unnatural isomers. This is
the sepia mutant of the Drosophila fruit fly.
Growing these fruit files requires a level of effort about
the same as making a loaf of bread, but without the baking.
Making a daily supply is like making a loaf of bread every
day, which is not a trivial amount of work, but it is within
what most people can do. It also requires space for growing
the flies about the size of a small closet.
Fruit flies are commonly grown as experimental animals for
biological laboratories, and the methods for growing them
have been worked out over more than 100 years. A good book
summarizing this knowledge is _Drosophila_Guide_ which is
available for free from various sources on the Internet,
such as this one:
http://www.public.asu.edu/~thoffman/commonfiles/lsc348/lsc348drosophilaguide.pdf
If you have no experience raising flies, you probably should
follow these time-tested methods because they will give you
consistent results and high productivity.
My growing methods are different in two main ways. First,
I do not use the mold inhibitor methyl paraben (a.k.a.
Tegosept M). Although methyl paraben has low toxicity for
humans and the amount carried in harvested adult flies is
very small, I do not wish to have it present in any amount.
Second, I do not sterilize the food (called culture medium)
that I feed my flies. I could sterilize the culture medium
with an ordinary pressure cooker, but that requires a
minimum of 1 hour at 1 atmosphere of pressure, which would
more than double the amount of time required to make a batch
of flies. I rely on living yeast to crowd out mold on the
culture medium, and I add an acid to suppress growth of
bacteria. I make the culture medium from bananas, cream of
tartar, active dry yeast, and water.
My results are not always consistent. Sometimes a batch
turns out very well, and all of the containers (called
culture vessels) produce many flies. But often, there will
be a few culture vessels that are poor performers with few
or no flies. I had believed these were due to bacterial
contamination, but now I think the problem is accumulation
of excess carbon dioxide gas from the living yeast in the
culture medium.
The culture vessels I use are disposable plastic cups with
dome lids of the type used for "bubble" tea. Specifically,
I use Fabri-Kal KC16S cups with DLKC16/24 lids. Between the
cup and the lid, I place a paper coffee filter to allow air
to enter while keeping the flies inside in the cup. I use
the smallest size of Melitta fluted coffee filters, intended
for 4-cup coffee brewers. The lids snap over the rims of
the cups and hold the coffee filters in place.
The flies I use are double mutants, both sepia mutant and
vestigial wing mutant. Vestigial wing mutants are much
easier to handle because they can't fly, although they can
hop and crawl. You can order these files here (do not
correct the spelling error in the link):
http://www.connecticutvalleybiological.com/drosophila-melanogaster-vestegial-sepia-p-11466.html
Any company that sells these mutants will also offer F1
sepia/vestigial hybrids. Do not buy those. You only want
purebred sepia/vestigial flies, because they breed true
generation after generation. F1 hybrids do not breed true.
I first tried apterous mutants, which have no wings at all.
They do not reproduce very well, probably because their lack
of wings prevents them from properly performing the mating
dance.
I harvest the cultures when they are 17 days old. The
best time to harvest will vary with temperature, because
temperature affects growth rate. I harvest one culture
every day and start one every day, so I have 17 cultures
in progress at any particular time.
From when the first flies are received until when harvesting
can begin may be more than 17 days, because time will be
needed to build up the population of flies until a surplus
is available. The amount of time will depend on the number
of flies used to get started. I ordered 12 vials in two
groups of 6, delivered one week apart. From this, it took
over three months until I had a surplus above the number
needed to maintain the population, though much of this time
was lost in experiments to optimize the recipe for the
culture medium. I had many failed experimental batches.
One vial has about enough flies to start 2 or 3 cultures,
and in another 2 or 3 weeks the vial will produce enough new
flies to start 2 or 3 more cultures. If you order enough
vials to receive 6 vials every day for 17 days, you might be
able to begin harvesting a surplus in 17 days. That would
be a rather expensive way to get started, and the supplier
may have had difficulty handling such a large order on short
notice. It is better to start slowly and gain experience
growing flies before running large batches.
I currently use 12 culture vessels for each daily batch.
I do not know how many flies is the minimum to start a
highly productive culture, but my experiments show that more
than 30 flies is a waste of flies. The surplus is about 2
to 3 grams (live weight) of flies. I'm still optimizing my
culture recipe, and I believe I can improve on this.
I use a stick blender to break up the live flies into
freshly squeezed orange juice. Orange juice is both acidic
and contains antioxidants (citric acid and ascorbic acid)
which help preserve the sepiapterin. I use the slowest
speed on the blender, to avoid whipping air into the orange
juice. The blended mixture should be consumed as soon as
possible after it is prepared. It should not be stored for
any length of time. The blended flies add no noticable
flavor or texture to the orange juice, however they are
visible as dark particles.
PART IV. What results have been obtained?
Before giving it to anybody else, I consumed five typical
batches of flies to ensure that they do not have any adverse
effects. I did not notice any effects at all.
Beginning on July 5, I've been giving it to my mother, who
has early stage AD. As of today (July 30), no clear effects
have been noticed. I was hoping for an unmistakeable,
immediate reversal of her symptoms, but that was probably
an unrealistic hope.
AD progresses slowly in small increments. If this therapy
is successful, that might only become obvious after a long
period of time. Even then it might only be seen as a
slowing or halting of the progress of the disease, which
would be hard to discern, especially without knowing what
course the disease would have taken without the therapy.
If this is an effective therapy, I do not know what an
effective dose will be. I frequently think about scaling up
production to a higher level, but for the time being I am
concentrating my effort on improving the productivity of my
cultures. I also do not know whether the results would have
been any better had I started earlier. All I know is that
this therapy is very unlikely to have any bad effects, and
the chance that it will have a good effect on the progress
of the disease is not unreasonably small.
PART V. References
There are other studies of biopterin (oxidized form of BH4)
levels in AD, but the ones which look at levels in blood or
urine are not the best data because those levels do not
accurately reflect the level inside the brain. Nearly all
of the biopterin in the brain is in the form of BH4, but
most analytical methods oxidize all forms to biopterin.
Two reports of biopterin levels in cerebrospinal fluid (CSF)
are:
"Cerebrospinal Fluid Biopterin Is Decreased in Alzheimer's
Disease", Archives of Neurology, 1986, volume 43, pages
996-999.
"Tetrabiopterin metabolism in senile dementia of the
Alzheimer type", Journal of Neurology, Neurosurgery, and
Psychiatry, 1983, volume 46, page 582. (letter)
There are several studies which looked at the decline of
biopterin with aging, but only one which graphed the level
in CSF against age:
"Aging and CSF hydroxylase cofactor", Neurology, 1980,
volume 30, pages 1244-1246.
Another study graphed the level in brain tissue obtained at
autopsy against age:
"Influence of development and aging on brain biopterin:
Implications for dopa-responsive dystonia onset", Neurology,
1998, volume 51, pages 632-634.
When I look at Figure 1 in the former and Figure C in the
latter, I wonder why Kuvan (or sepiapterin) is not already
a multi-billion-dollar anti-aging drug. Everybody should
be taking this!
In this letter, a suggestion is made that AD may be an
accelerated form of the age-related decline in BH4:
"Pathogenesis of Alzheimer's disease", Journal of the Royal
Society of Medicine, 1987, volume 80, pages 127-129.
That is a similar idea to mine, except I don't believe it
must be an accelerated form. In people who start with low
levels, the normal age-related decline may be enough to
trigger AD.
This study found a 50 milligram dose of sepiapterin has
about the same effect as a 200 milligram dose of BH4 in
treating the genetic form of tetrahydrobiopterin deficiency:
"Atypical Phenylketonuria with Defective Biopterin
Metabolism. Monotherapy with Tetrahydrobiopterin or
Sepiapterin, Screening und Study of Biosynthesis in Man",
European Journal of Pediatrics, 1982, volume 138, pages
110-112.
Copyright 2012 Mark Thorson (ver. 7/30/12)
This essay describes a new therapeutic approach to
Alzheimer's Disease (AD). This approach is based on the
possibility that the cause of AD is a deficiency of BH4
(a.k.a. tetrahydrobiopterin), which is a cofactor for
enzymes needed to make important neurotransmitters. The
therapeutic agent is something you can make in your kitchen
from materials you can buy without any limits or regulation.
The procedure for making the agent is completely safe and
requires no particular skills nor equipment not normally
found in a kitchen.
The remainder of this essay is divided into five parts:
I. What is the evidence that BH4 deficiency causes AD?
II. What is the best treatment for BH4 deficiency?
III. How can I make sepiapterin?
IV. What results have been obtained?
V. References
PART I. What is the evidence that BH4 causes AD?
BH4 is like a vitamin in its role as a cofactor for enzymes.
A cofactor is a molecule that an enzyme needs to carry out
its function. A neurotransmitter is a molecule released
by one nerve cell to send a signal to another nerve cell.
BH4 is a cofactor for enzymes needed to make several
neurotransmitters: dopamine, epinephrine, norepinephrine,
serotonin, and melatonin. Although the exact way in
which AD affects memory and causes confusion is not well
understood, shortages of these important neurotransmitters
could easily be part of this process.
BH4 is also a cofactor for nitric oxide synthase enzymes,
which make nitric oxide. Nitric oxide relaxes blood vessels
in the brain. Lack of nitric oxide tightens these blood
vessels and reduces blood flow. Nitric oxide also acts as
a neurotransmitter believed to play an important role in
forming long-term memories.
In Alzheimer's disease, the level of BH4 (measured
indirectly as the level of biopterin) in the brain is
greatly reduced. Studies show the level in cerebrospinal
fluid (the fluid which bathes the brain and spinal cord) is
about 30 to 50% lower in people with AD in comparison with
people of the same age without AD.
Other studies show that the level declines dramatically
with age. Between the ages of 20 and 70, the level falls by
about 50%. These same studies also show a wide variation.
Some young adults have levels similar to senior citizens,
and some senior citizens have levels similar to young
adults. This may explain why some people get AD in old age
and some don't -- there may be a threshold level at which
AD occurs, and some people reach that level in old age while
others never reach that level.
BH4 is currently available as a pharmaceutical drug called
Kuvan from BioMarin Pharmaceutical. Currently, Kuvan is
only marketed for treatment of a rare genetic disease.
The first clinical trial of Kuvan in people with AD is
currently recruiting subjects, but it will be some time
before any results are reported. For more information
about the trial, go here:
http://clinicaltrials.gov/show/NCT01439555
PART II. What is the best treatment for BH4 deficiency?
There are several difficulties in treating AD with BH4.
First, BH4 is a very unstable molecule. It is attacked
by oxygen from the air. To make BH4 stable enough to
distribute as a pharmaceutical, it must be in the form of
an acid salt, it must be mixed with antioxidants, it must
be packaged in oxygen-barrier materials with an oxygen
absorber, and it must be kept frozen at very low
temperature.
Second, BH4 is poorly absorbed. It does not easily cross
the blood-brain barrier into the central nervous system.
Third, BH4 is a pharmaceutical drug not available without
a prescription. The only purpose for which BH4 has been
approved by the FDA is treatment of tetrahydrobiopterin
deficiency (formerly known as atypical phenylketonuria).
This is a genetic disease in which one of the enzymes
necessary for the biosynthesis of BH4 is defective.
It is diagnosed in infancy, and its victims usually have
severe mental retardation and short life spans. If AD is
caused by a BH4 deficiency, it differs from the genetic form
in being a milder form which strikes long after the growth
and development of the nervous system is complete.
There is a therapeutic agent for BH4 deficiency which is
better than BH4 in all of these respects. It is much more
stable, though stability is still a problem. It has greater
efficacy in treating BH4 deficiency, which implies it has
better absorption. It is available to anyone who wishes to
make it. This agent is sepiapterin.
Once sepiapterin is absorbed, it is converted into BH4.
Sepiapterin is very similar to BH4, differing only in two
chemical bonds. In the conversion to BH4, these two bonds
are opened up and hydrogen atoms are placed on the free
ends. This change is necessary for the function of BH4 as
an enzyme cofactor, in which BH4 acts as a hydrogen donor,
but the change also makes the molecule very vulnerable to
attack by oxygen.
PART III. How can I make sepiapterin?
Making either BH4 or sepiapterin synthetically would require
chemicals, equipment, and skills which you are unlikely to
have. Also, it would require purification to remove
unnatural forms of these molecules (called isomers) which
have unknown effects in the human body.
However, there is a biological source of natural sepiapterin
you can grow which is free of unnatural isomers. This is
the sepia mutant of the Drosophila fruit fly.
Growing these fruit files requires a level of effort about
the same as making a loaf of bread, but without the baking.
Making a daily supply is like making a loaf of bread every
day, which is not a trivial amount of work, but it is within
what most people can do. It also requires space for growing
the flies about the size of a small closet.
Fruit flies are commonly grown as experimental animals for
biological laboratories, and the methods for growing them
have been worked out over more than 100 years. A good book
summarizing this knowledge is _Drosophila_Guide_ which is
available for free from various sources on the Internet,
such as this one:
http://www.public.asu.edu/~thoffman/commonfiles/lsc348/lsc348drosophilaguide.pdf
If you have no experience raising flies, you probably should
follow these time-tested methods because they will give you
consistent results and high productivity.
My growing methods are different in two main ways. First,
I do not use the mold inhibitor methyl paraben (a.k.a.
Tegosept M). Although methyl paraben has low toxicity for
humans and the amount carried in harvested adult flies is
very small, I do not wish to have it present in any amount.
Second, I do not sterilize the food (called culture medium)
that I feed my flies. I could sterilize the culture medium
with an ordinary pressure cooker, but that requires a
minimum of 1 hour at 1 atmosphere of pressure, which would
more than double the amount of time required to make a batch
of flies. I rely on living yeast to crowd out mold on the
culture medium, and I add an acid to suppress growth of
bacteria. I make the culture medium from bananas, cream of
tartar, active dry yeast, and water.
My results are not always consistent. Sometimes a batch
turns out very well, and all of the containers (called
culture vessels) produce many flies. But often, there will
be a few culture vessels that are poor performers with few
or no flies. I had believed these were due to bacterial
contamination, but now I think the problem is accumulation
of excess carbon dioxide gas from the living yeast in the
culture medium.
The culture vessels I use are disposable plastic cups with
dome lids of the type used for "bubble" tea. Specifically,
I use Fabri-Kal KC16S cups with DLKC16/24 lids. Between the
cup and the lid, I place a paper coffee filter to allow air
to enter while keeping the flies inside in the cup. I use
the smallest size of Melitta fluted coffee filters, intended
for 4-cup coffee brewers. The lids snap over the rims of
the cups and hold the coffee filters in place.
The flies I use are double mutants, both sepia mutant and
vestigial wing mutant. Vestigial wing mutants are much
easier to handle because they can't fly, although they can
hop and crawl. You can order these files here (do not
correct the spelling error in the link):
http://www.connecticutvalleybiological.com/drosophila-melanogaster-vestegial-sepia-p-11466.html
Any company that sells these mutants will also offer F1
sepia/vestigial hybrids. Do not buy those. You only want
purebred sepia/vestigial flies, because they breed true
generation after generation. F1 hybrids do not breed true.
I first tried apterous mutants, which have no wings at all.
They do not reproduce very well, probably because their lack
of wings prevents them from properly performing the mating
dance.
I harvest the cultures when they are 17 days old. The
best time to harvest will vary with temperature, because
temperature affects growth rate. I harvest one culture
every day and start one every day, so I have 17 cultures
in progress at any particular time.
From when the first flies are received until when harvesting
can begin may be more than 17 days, because time will be
needed to build up the population of flies until a surplus
is available. The amount of time will depend on the number
of flies used to get started. I ordered 12 vials in two
groups of 6, delivered one week apart. From this, it took
over three months until I had a surplus above the number
needed to maintain the population, though much of this time
was lost in experiments to optimize the recipe for the
culture medium. I had many failed experimental batches.
One vial has about enough flies to start 2 or 3 cultures,
and in another 2 or 3 weeks the vial will produce enough new
flies to start 2 or 3 more cultures. If you order enough
vials to receive 6 vials every day for 17 days, you might be
able to begin harvesting a surplus in 17 days. That would
be a rather expensive way to get started, and the supplier
may have had difficulty handling such a large order on short
notice. It is better to start slowly and gain experience
growing flies before running large batches.
I currently use 12 culture vessels for each daily batch.
I do not know how many flies is the minimum to start a
highly productive culture, but my experiments show that more
than 30 flies is a waste of flies. The surplus is about 2
to 3 grams (live weight) of flies. I'm still optimizing my
culture recipe, and I believe I can improve on this.
I use a stick blender to break up the live flies into
freshly squeezed orange juice. Orange juice is both acidic
and contains antioxidants (citric acid and ascorbic acid)
which help preserve the sepiapterin. I use the slowest
speed on the blender, to avoid whipping air into the orange
juice. The blended mixture should be consumed as soon as
possible after it is prepared. It should not be stored for
any length of time. The blended flies add no noticable
flavor or texture to the orange juice, however they are
visible as dark particles.
PART IV. What results have been obtained?
Before giving it to anybody else, I consumed five typical
batches of flies to ensure that they do not have any adverse
effects. I did not notice any effects at all.
Beginning on July 5, I've been giving it to my mother, who
has early stage AD. As of today (July 30), no clear effects
have been noticed. I was hoping for an unmistakeable,
immediate reversal of her symptoms, but that was probably
an unrealistic hope.
AD progresses slowly in small increments. If this therapy
is successful, that might only become obvious after a long
period of time. Even then it might only be seen as a
slowing or halting of the progress of the disease, which
would be hard to discern, especially without knowing what
course the disease would have taken without the therapy.
If this is an effective therapy, I do not know what an
effective dose will be. I frequently think about scaling up
production to a higher level, but for the time being I am
concentrating my effort on improving the productivity of my
cultures. I also do not know whether the results would have
been any better had I started earlier. All I know is that
this therapy is very unlikely to have any bad effects, and
the chance that it will have a good effect on the progress
of the disease is not unreasonably small.
PART V. References
There are other studies of biopterin (oxidized form of BH4)
levels in AD, but the ones which look at levels in blood or
urine are not the best data because those levels do not
accurately reflect the level inside the brain. Nearly all
of the biopterin in the brain is in the form of BH4, but
most analytical methods oxidize all forms to biopterin.
Two reports of biopterin levels in cerebrospinal fluid (CSF)
are:
"Cerebrospinal Fluid Biopterin Is Decreased in Alzheimer's
Disease", Archives of Neurology, 1986, volume 43, pages
996-999.
"Tetrabiopterin metabolism in senile dementia of the
Alzheimer type", Journal of Neurology, Neurosurgery, and
Psychiatry, 1983, volume 46, page 582. (letter)
There are several studies which looked at the decline of
biopterin with aging, but only one which graphed the level
in CSF against age:
"Aging and CSF hydroxylase cofactor", Neurology, 1980,
volume 30, pages 1244-1246.
Another study graphed the level in brain tissue obtained at
autopsy against age:
"Influence of development and aging on brain biopterin:
Implications for dopa-responsive dystonia onset", Neurology,
1998, volume 51, pages 632-634.
When I look at Figure 1 in the former and Figure C in the
latter, I wonder why Kuvan (or sepiapterin) is not already
a multi-billion-dollar anti-aging drug. Everybody should
be taking this!
In this letter, a suggestion is made that AD may be an
accelerated form of the age-related decline in BH4:
"Pathogenesis of Alzheimer's disease", Journal of the Royal
Society of Medicine, 1987, volume 80, pages 127-129.
That is a similar idea to mine, except I don't believe it
must be an accelerated form. In people who start with low
levels, the normal age-related decline may be enough to
trigger AD.
This study found a 50 milligram dose of sepiapterin has
about the same effect as a 200 milligram dose of BH4 in
treating the genetic form of tetrahydrobiopterin deficiency:
"Atypical Phenylketonuria with Defective Biopterin
Metabolism. Monotherapy with Tetrahydrobiopterin or
Sepiapterin, Screening und Study of Biosynthesis in Man",
European Journal of Pediatrics, 1982, volume 138, pages
110-112.