Source: Rett Syndrome:
Crossing the Threshold to Clinical Translation
Today’s post is on the one hand very
specific to Rett syndrome, but much is applicable to broader autism and other
single gene autisms.
Today’s post did start out with the
research showing Bumetanide effective in the mouse model of Rett syndrome. This
ended up with figuring out why this should have been obvious based on what we
already know about growth factors that are disturbed in autism and very much so
in Rett.
We even know from a published human
case studies that Bumetanide can benefit those with Fragile X and indeed Down
syndrome, but the world takes little notice.
If Bumetanide benefits human Rett syndrome
would anyone take any notice? They
really should.
To readers of this blog who have a
child with Rett, the results really are important. You can even potentially link the problem
symptoms found in Rett to the biology and see how you can potentially treat
multiple symptoms with the same drug.
One feature of Rett is breathing
disturbances, which typically consist of alternating periods of
hyperventilation and hypoventilation.
Our reader Daniel sent me a link to
paper that suggest an old OTC cough medicine could be used to treat the
breathing issues.
The antitussive cloperastine improves breathing abnormalities in a
Rett Syndrome mouse model by blocking presynaptic GIRK channels and enhancing
GABA release
Rett Syndrome (RTT) is an X-linked neurodevelopmental disorder
caused mainly by mutations in the MECP2 gene. One of the major RTT features is
breathing dysfunction characterized by periodic hypo- and hyperventilation. The
breathing disorders are associated with increased brainstem neuronal
excitability, which can be alleviated with antagonistic agents.
Since neuronal hypoexcitability occurs in the forebrain of RTT
models, it is necessary to find pharmacological agents with a relative
preference to brainstem neurons. Here we show evidence for the improvement of
breathing disorders of Mecp2-null mice with the brainstem-acting drug
cloperastine (CPS) and its likely neuronal targets. CPS is an over-the-counter
cough medicine that has an inhibitory effect on brainstem neuronal networks. In
Mecp2-null mice, CPS (30 mg/kg, i.p.) decreased the occurrence of apneas/h and
breath frequency variation. GIRK currents expressed in HEK cells were inhibited
by CPS with IC50 1 μM. Whole-cell patch clamp recordings
in locus coeruleus (LC) and dorsal tegmental nucleus (DTN) neurons revealed an
overall inhibitory effect of CPS (10 μM) on neuronal
firing activity. Such an effect was reversed by the GABAA receptor antagonist
bicuculline (20 μM). Voltage clamp studies showed that
CPS increased GABAergic sIPSCs in LC cells, which was blocked by the GABAB receptor
antagonist phaclofen. Functional GABAergic connections of DTN neurons with LC
cells were shown.
These results suggest that CPS
improves breathing dysfunction in Mecp2-null mice by blocking GIRK channels in
synaptic terminals and enhancing GABA release.
Cloperastine
(CPS) is a central-acting antitussive working on brainstem neuronal networks The
drug has several characteristics. 1) It affects the brainstem integration of
multiple sensory inputs via multiple sites including K+ channels, histamine and
sigma receptors. 2) Its overall effect is inhibitory, suppressing cough and
reactive airway signals. 3) With a large safety margin, it has been approved as
an over-the-counter medicine in several Asian and European countries.
With
the evidence that DTN cells receive GABAergic recurrent inhibition, we tested
whether the inhibitory effect of CPS was caused by enhanced GABAergic
transmission. Thus, we recorded the evoked firing activity of DTN cells before
and during bath application of CPS in the presence of 20 μM bicuculline. Under
this condition, CPS failed to decrease the excitability of DTN neurons (F(1,9)
= 0.41, P > 0.05; two‐way repeated measures ANOVA) (n=9) (Fig. 8), indicating that the inhibitory
effect relies on GABAA synaptic input
It appeared to me that the breathing
issues might be considered as another consequence of the excitatory/inhibitory
(E/I) imbalance that is a core feature of much severe autism.
In the case of Rett the lack of BDNF
will make any E/I imbalance worse and that by treating the E/I imbalance we
will produce the inhibitory effect from GABAa receptors that is needed to
ensure correct breathing. Note that in
bumetanide responsive autism there is no inhibitory effect from GABAa
receptors, the effect is excitatory.
I did wonder if arrhythmia (irregular
heartbeat) is present in Rett, since the breathing problems in Rett are also seen
as being caused by a dysfunction in the autonomic nervous system. Arrhythmia is
actually a big problem for girls with Rett syndrome. Regular readers of this blog might then ask
about Propranolol, does that help? It
turns out to have been tried and it is not so helpful. What is effective is another drug we have
come across for autism, the sodium channel blocker Phenytoin. Phenytoin is antiepileptic drug (AED) and it
works by blocking voltage gated sodium channels.
Low dose phenytoin was proposed as an
autism therapy and a case study was published from Australia. In a separate
case study, phenytoin was used to treat self-injury that was triggered by
frontal lobe seizures.
When you treat arrhythmia in Rett
girls with Phenytoin does it have an impact on their breathing problems?
If you treat the girls with Phenytoin
do they still go on to develop epilepsy?
What about if you add treatment with
Bumetanide to reduce symptoms of autism?
Lots of questions looking for answers.
What is Rett
Syndrome?
Rett syndrome was first identified in
the 1950s by Dr Andreas Rett as a disorder that develops in young girls. Only as recently as 1999 was it determined
that the syndrome is caused by a mutation in the MECP2 gene on the X
chromosome. The X chromosome is very
important because girls have two copies, but boys have just one. Rett was an Austrian like many other early
researchers in autism like Kanner and Asperger. Even Freud was educated in
Vienna. Eugen Bleuler lived pretty close by in Switzerland and he coined the
terms schizophrenia, schizoid and autism.
Rett syndrome is a rare genetic
disorder that affects brain development, resulting in severe mental and
physical disability.
It is estimated to affect about 1 in
12,000 girls born each year.
Rett is a rare condition, but among
these rare conditions it is quite common and so there is a lot of research
going on to find treatments. The obvious
one is gene therapy to get the brain to make the missing MeCP2 protein.
Rett syndrome is thankfully rare in
absolute terms, but it is one of the best known development conditions that is
associated with autism symptoms.
While Rett syndrome may not officially
be an ASD in the DSM-5, the link to autism remains. Many children are diagnosed
as autistic before the MECP2 mutation is identified and then the diagnosis is
revised to RTT/Rett.
Fragile X syndrome (FXS), on the other hand, is the most
common inherited cause of intellectual disability (ID), as well as the most
frequent single gene type of autism.
In the meantime, the logical strategy
is to treat the downstream consequences of the mutated gene. Much is known
about these downstream effects and there overlaps with some broader autism and indeed
dementia.
One area known to be disturbed in
Rett, some other autisms and dementia is growth factors inside the brain. The
best known growth factors are IGF-1 (Insulin-like Growth Factor 1), BDNF (brain-derived
neurotrophic factor) and my favorite NGF (Nerve growth factor).
Without wanting to get too complicated
we need to note that BDNF acts via a receptor called TrkB. You can either increase BDNF or just find
something else to activate TrkB, as pointed out to me by Daniel.
For readers whose children respond to
Bumetanide they are benefiting from correcting elevated levels of chloride in
neurons. Too much had been entering by the transporter NKCC1 and too little
exiting via KCC2.
One of the effects of having too
little BDNF and hence not enough activation of TrkB is that chloride becomes
elevated in neurons. If you do not
activate TrkB you do not get enough KCC2, which is what allows chloride to exit
neurons.
To what extent would TrkB activation
be an alternative/complement to bumetanide in broader autism?
To what extent would TrkB activation
be success in treating some types of chronic pain (where KCC2 is known to be
down regulated)?
Low levels of BDNF are a feature of
Rett and much dementia.
So you would want to:
·
Increase BDNF
·
Activate TRKB
with something else
·
Block NKCC2 to compensate
for the lack of KCC2
Note that BDNF is not reduced in all
types of autism, just in a sub-group.
I note that there already is solid
evidence in the research:-
Reduced
expression of brain-derived neurotrophic factor (BDNF) and impaired activation
of the BDNF receptor, tropomyosin receptor kinase B (TrkB; also known as
Ntrk2), are thought to contribute significantly to the pathophysiology of Rett
syndrome (RTT), a severe neurodevelopmental disorder caused by
loss-of-function mutations in the X-linked gene encoding methyl-CpG-binding
protein 2 (MeCP2). Previous studies from this and other laboratories have shown
that enhancing BDNF expression and/or TrkB activation in Mecp2-deficient
mouse models of RTT can ameliorate or reverse abnormal neurological phenotypes
that mimic human RTT symptoms. The present study reports on the preclinical
efficacy of a novel, small-molecule, non-peptide TrkB partial agonist,
PTX-BD4-3, in heterozygous female Mecp2 mutant mice, a well-established RTT model
that recapitulates the genetic mosaicism of the human disease. PTX-BD4-3
exhibited specificity for TrkB in cell-based assays of neurotrophin receptor
activation and neuronal cell survival and in in vitro receptor
binding assays. PTX-BD4-3 also activated TrkB following systemic administration
to wild-type and Mecp2 mutant mice and was rapidly cleared from
the brain and plasma with a half-life of ∼2 h. Chronic intermittent treatment of Mecp2 mutants
with a low dose of PTX-BD4-3 (5 mg/kg, intraperitoneally, once every 3 days for
8 weeks) reversed deficits in two core RTT symptom domains – respiration and
motor control – and symptom rescue was maintained for at least 24 h after the
last dose. Together, these data indicate that significant clinically relevant
benefit can be achieved in a mouse model of RTT with a chronic intermittent,
low-dose treatment paradigm targeting the neurotrophin receptor TrkB.
Early
alterations in a mouse model of Rett syndrome: the GABA developmental shift is
abolished at birth
Genetic mutations of the Methyl-CpG-binding
protein-2 (MECP2) gene underlie Rett syndrome (RTT). Developmental processes
are often considered to be irrelevant in RTT pathogenesis but neuronal activity
at birth has not been recorded. We report that the GABA developmental shift at
birth is abolished in CA3 pyramidal neurons of Mecp2−/y mice
and the glutamatergic/GABAergic postsynaptic currents (PSCs) ratio is
increased. Two weeks later, GABA exerts strong excitatory actions, the
glutamatergic/GABAergic PSCs ratio is enhanced, hyper-synchronized activity is
present and metabotropic long-term depression (LTD) is impacted. One day before
delivery, maternal administration of the NKCC1 chloride importer antagonist
bumetanide restored these parameters but not respiratory or weight deficits,
nor the onset of mortality. Results
suggest that birth is a critical period in RTT with important alterations that
can be attenuated by bumetanide raising the possibility of early treatment of
the disorder.
GABA depolarizes and often excites immature neurons
in all animal species and brain
structures investigated due to a developmentally regulated reduction in
intracellular chloride concentration ([Cl−]i) levels. The
control of [Cl−]i levels is mediated by the chloride
cotransporters NKCC1 and KCC2, the former usually importing chloride and the
latter exporting it. The GABA polarity shift has been extensively validated in
several experimental conditions using often the NKCC1 chloride importer
antagonist bumetanide. In spite of an intrinsic heterogeneity, this shift is abolished in many
experimental conditions associated with developmental disorders including
autism, Rett syndrome, fragile X syndrome, or maternal immune activation.
Using bumetanide, an EMA- and FDA-approved agent, many clinical trials have
shown promising results with the expected side effects. Kaila et al. have
repeatedly challenged these experimental and clinical observations. Here, we
reply to the recent reviews by Kaila et al. stressing that the GABA polarity
shift is solidly accepted by the scientific community as a major discovery to
understand brain development and that bumetanide has shown promising effects in
clinical trials.
Back in 2013 a case study was published
showing Bumetanide worked for a boy with Fragile X syndrome. A decade later and
still nobody has looked to see if it works in all Fragile X.
Treating Fragile X
syndrome with the diuretic bumetanide: a case report
https://pubmed.ncbi.nlm.nih.gov/23647528/
We
report that daily administration of the diuretic NKCC1 chloride co-transporter,
bumetanide, reduces the severity of autism in a 10-year-old Fragile X boy using
CARS, ADOS, ABC, RDEG and RRB before and after treatment. In keeping
with extensive clinical use of this diuretic, the only side effect was a small
hypokalaemia. A
double-blind clinical trial is warranted to test the efficacy of bumetanide in
FRX.
What do Rett
syndrome and Fragile X have in common?
In a healthy mature neuron the level
of chloride needs to be low for it to function correctly (the neurotransmitter
GABA to be inhibitory).
Rett and Fragile X are part of a large
group of conditions that feature elevated levels of chloride in neurons.
Elevated chloride in neurons is
treatable.
Is Bumetanide a
cure for Rett syndrome, or Fragile X?
No it is not, but it is a step in that
direction because it reverses a key defect present in at least some Rett and
some Fragile X.
In the mouse model of Rett, bumetanide
corrected some, but not all the problems caused by the loss of function of the MECP2
gene.
Moving on to
IGF-1
IGF-1 is a growth hormone with
multiple functions throughout aging. Production of IGF-1 is stimulated by GH
(growth hormone).
The lowest levels occur in infancy and
old age and highest levels occur around the growth spurt before puberty.
Girls with Turner syndrome, lack their
second X chromosome and this causes a lack of growth hormones and female
hormones. They end up with short stature and with features of autism. Treatment
is possible with GH or indeed IGF-1.
In dementia one strategy is to
increase IGF-1. This same strategy is
also being applied to single gene autisms like Rett and Pitt Hopkins.
Trofinetide and NNZ-2591 are improved
synthetic analogues of peptides that occur naturally in the brain and are
related to IGF-1. Trofinetide is being developed to treat Rett and Fragile X
syndromes, NNZ-2591 is being developed to treat Angelman, Phelan-McDermid, Pitt
Hopkins and Prader-Willi syndromes.
NGF (nerve growth
factor)
Nerve growth factor does what it says
(boosting nerve growth), plus much more. NGF plays a key role in the immune
system, it is produced in mast cells, and it plays a role in how pain in perceived.
NGF acts via NGF receptors, not surprisingly,
but also via TrkA receptors. We saw earlier in this post that BDNF acts via
TrkB receptors.
Once NGF binds to the TrkA receptor it
triggers a cascade of signalling via the
Ras/MAPK pathway and the PI3K/Akt pathway.
Both pathways relate to autism and Ras itself can play a role in
intellectual disability.
These are also cancer pathways and
indeed NGF seems to play a role. Beta
cells in the pancreas produce insulin and these beta cells have TrkA receptors.
In type 1 diabetes these beta cells die.
Beta cells need NGF to activate their TrkA receptors to survive.
Clearly for multiple reasons you need
plenty of NGF.
Lack of NGF would be one cause of
dementia and that is why Rita Levi-Montalcini choose to self-treat with NGF eye
drops for 30 years. Rita won a Nobel prize for discovering NGF.
In Rett syndrome we know that the
level of NGF is very low in the brain.
Logical therapies for Rett would seem
to include:
·
NGF itself,
perhaps taken as eye drops, but tricky to administer
·
A TrkA agonist,
that would mimic the effect of NGF
·
The traditional medicinal mushroom Lion’s Mane (Hericium
erinaceus)
We should note that effect of NGF
acting via TrkA is mainly in the peripheral nervous system, not the brain.
It has long been known that Lions’ Mane
(Hericium erinaceus) increases NGF
but it was not clear why. This has very
recently been answered.
The
active chemical has been identified to be N-de phenylethyl isohericerin
(NDPIH).
The
opens the door to synthesizing NDPIH as drug to treat a wide range of
conditions from Alzheimer’s to Rett.
Mushrooms Magnify Memory by Boosting Nerve Growth
Active
compounds in the edible Lion’s Mane mushroom can help promote neurogenesis and
enhance memory, a new study reports. Preclinical trials report the compound had
a significant impact on neural growth and improved memory formation.
Researchers say the compound could have clinical applications in treating and
preventing neurodegenerative disorders such as Alzheimer’s disease.
Professor
Frederic Meunier from the Queensland Brain Institute said the team had
identified new active compounds from the mushroom, Hericium erinaceus.
“Extracts
from these so-called ‘lion’s mane’ mushrooms have been used in traditional
medicine in Asian countries for centuries, but we wanted to scientifically
determine their potential effect on brain cells,” Professor Meunier said.
“Pre-clinical
testing found the lion’s mane mushroom had a significant impact on the growth
of brain cells and improving memory.
“Laboratory
tests measured the neurotrophic effects of compounds isolated from Hericium erinaceus
on cultured brain cells, and surprisingly we found that the active compounds promote neuron
projections, extending and connecting to other neurons.
“Using super-resolution microscopy, we
found the mushroom extract and its active components largely increase the size
of growth cones, which are particularly important for brain cells to sense
their environment and establish new connections with other neurons in the
brain.”
Hericerin derivatives activates a pan‐neurotrophic pathway in central
hippocampal neurons converging to ERK1/2 signaling enhancing spatial memory
The traditional medicinal mushroom Hericium
erinaceus is known for enhancing peripheral nerve regeneration through
targeting nerve growth factor (NGF) neurotrophic activity. Here, we purified
and identified biologically new active compounds from H. erinaceus,
based on their ability to promote neurite outgrowth in hippocampal
neurons. N-de phenylethyl isohericerin
(NDPIH), an isoindoline compound from this mushroom, together with its
hydrophobic derivative hericene
A, were highly potent in promoting extensive axon outgrowth and neurite
branching in cultured hippocampal neurons even in the absence of serum,
demonstrating potent neurotrophic activity. Pharmacological inhibition of
tropomyosin receptor kinase B (TrkB) by ANA-12 only partly prevented the
NDPIH-induced neurotrophic activity, suggesting a potential link with BDNF
signaling. However, we found that NDPIH activated ERK1/2 signaling in the
absence of TrkB in HEK-293T cells, an effect that was not sensitive to ANA-12
in the presence of TrkB. Our
results demonstrate that NDPIH acts via a complementary neurotrophic pathway
independent of TrkB with converging downstream ERK1/2 activation. Mice
fed with H. erinaceus crude extract and hericene A also
exhibited increased neurotrophin expression and downstream signaling, resulting
in significantly enhanced hippocampal memory. Hericene A therefore acts through a novel
pan-neurotrophic signaling pathway, leading to improved cognitive
performance.
Since
the discovery of the first neurotrophin, NGF, more than 70 years ago, countless
studies have demonstrated their ability to promote neurite regeneration,
prevent or reverse neuronal degeneration and enhance synaptic plasticity. Neurotrophins
have attracted the attention of the scientific community in the view to
implement therapeutic strategies for the treatment of a number of neurological
disorders. Unfortunately, their actual therapeutic applications have been
limited and the potential use of their beneficial effects remain to be
exploited. Neurotrophins,
for example, have poor oral bioavailability, and very low stability in serum,
with half-lives in the order of minutes as well as minimal BBB permeability and
restricted diffusion within brain parenchyma. In addition, their receptor
signaling networks can confer undesired off-target effects such as pain,
spasticity and even neurodegeneration. As a consequence, alternative strategies
to increase neurotrophin levels, improve their pharmacokinetic limitations or
target specific receptors have been developed. Identification of bioactive
compounds derived from natural products with neurotrophic activities also
provide new hope in the development of sustainable therapeutical interventions.
Hericerin derivative are
therefore attractive compounds for their ability to promote a pan-neurotrophic
effect with converging ERK1/2 downstream signaling pathway and for their
ability to promote the expression of neurotrophins. Further work will be
needed to find the direct target of Hericerin capable of mediating such a
potent pan-neurotrophic activity and establish whether this novel pathway can
be harnessed to improve memory performance and for slowing down the cognitive
decline associated with ageing and neurodegenerative diseases.
What this means is that there are 2
good reasons why Lion’s Mane should be helpful in Rett syndrome, both
increasing BDNF and NGF.
Conclusion
Interestingly, one of the above papers
is co-authored by a researcher from the European Brain Research Institute, founded by Rita
Levi-Montalcini, the Nobel laureate who discovered NGF (Nerve growth factor).
My top pick to test next in Rett syndrome would be NGF. Administration would have
to follow Rita’s own example and be in the form of eye drops or follow the Lion’s
Mane option, that has recently been further
validated.
Rett syndrome is very well documented
and many researchers are engaged in studying it.
As with broader autism, the problem is
translating all the research into practical therapy today.
Clearly polytherapy will be required.
More than one type of neuronal
hyperexcitability seems to be in play.
It looks like one E/I imbalance is the
bumetanide responsive kind, that can be treated and will reduce autism symptoms
and improve learning skills. Then we
have the hypoventilation/apnea for which Cloperastine looks a fair bet. For the arrhythmia we have Phenytoin. If there are still seizures after all that
therapy it looks like sodium valproate is the standard treatment for Rett.
Sodium valproate is also an HDAC
inhibitor and so has possibly beneficial epigenetic effects as a bonus.
I have always liked the idea of the
Lion’s Mane mushrooms as a means to increase NGF (Nerve growth factor). In today’s post we saw that it is the NDPIH
from the mushrooms that acts to increase both BDNF and NGF. You would struggle to buy NDPIH but you can
buy these mushrooms. I did once buy the supplement version of these mushrooms
and it was contaminated, so I think the best bet is the actual chemical or the
actual mushroom. One reader did write in
once who is a big consumer of these mushrooms.
Lion's Mane Mushroom
Source: Igelstachelbart Nov 06
A Trk-B agonist that can penetrate the
blood brain barrier would look a good idea.
There are some sold by the nootropic people.
7,8-dihydroxyflavone is such an
agonist that showed a benefit in the mouse model.
7,8-dihydroxyflavone
exhibits therapeutic efficacy in a mouse model of Rett syndrome
Following
weaning, 7,8-DHF was administered in drinking water throughout life. Treated
mutant mice lived significantly longer compared with untreated mutant
littermates (80 ± 4 and 66 ± 2 days, respectively). 7,8-DHF delayed body weight
loss, increased neuronal nuclei size and enhanced voluntary locomotor (running
wheel) distance in Mecp2 mutant mice. In addition, administration of 7,8-DHF
partially improved breathing pattern irregularities and returned tidal volumes
to near wild-type levels. Thus although the specific mechanisms are not
completely known, 7,8-DHF appears to reduce disease symptoms in Mecp2 mutant
mice and may have potential as a therapeutic treatment for RTT patients.
Rett
syndrome also features mitochondrial dysfunction and a variant of metabolic
syndrome. We have quite a resource
available from broader autism, not much of it seems to have been applied in
Rett.
You can see that in Rett less oxygen
is available due to breathing issues and yet more oxygen is required due to
“faulty” mitochondria.
“Intensified
mitochondrial O2 consumption, increased mitochondrial ROS generation and
disturbed redox balance in mitochondria and cytosol may represent a causal
chain, which provokes dysregulated proteins, oxidative tissue damage, and
contributes to neuronal network dysfunction in RTT.”
https://www.frontiersin.org/articles/10.3389/fphys.2019.00479/full#:~:text=Rett%20syndrome%20(RTT)%2C%20an,inner%20membrane%20is%20leaking%20protons.
We have seen in this blog that 2 old
drugs exist to increase oxygen levels in blood.
The Western world has Diamox (Acetazolamide) and the former soviet world
has Mildronate/Meldonium. Mildronate also was suggested to have some wider
potential benefit to mitochondria.
Rett is proposed as a neurological
disorder with metabolic components, so based on what we have seen in this blog,
you would think along the lines of Metformin, Pioglitazone and a lipophilic
statin (Atorvastatin, Simvastatin or Lovastatin).
The
Anti-Diabetic Drug Metformin Rescues Aberrant Mitochondrial Activity and
Restrains Oxidative Stress in a Female Mouse Model of Rett Syndrome
Statins
improve symptoms of Rett syndrome in mice
The ultimate Rett cure will be one of
the new gene therapies given to a baby before any significant progression of the
disorder has occurred.
For everyone else, it looks like there
is scope to develop a pretty potent individualized polytherapy, just by
applying the very substantial knowledge that already exists in the research.
Good luck to Daniel and all the others
seeking answers.
