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Thursday, 12 April 2018

HDAC Inhibitors for which Cancer/Autism?


Most types of autism can be viewed as the miss-expression of a few hundred genes, in some cases this has been caused by an initial defect in just one gene.  These single gene autisms are the ones that are usually studied.

Epigenetics has been covered previously in this blog and can either be made to look ultra-complex, which is the reality, or quite simple. The simple view is that in some people genes are miss-expressed because they have been tagged with heritable and removable markers; these can be wiped away. One type of epigenetic marker can be modified by an HDAC inhibitor or HDI.  
Some medical conditions featured genes turned off when they should be on. For example tumor suppressor gene (and autism gene) PTEN is turned off in the prostate of many males with prostate cancer; a neat therapy would be to switch it back on.  Deacetylation of PTEN by SIRT1 deacetylase and, by HDAC1, can stimulate its activity, so probably a good thing for people with this kind of common cancer.
In some types of autism there is a deficiency of a single protein because one of the two copies of the gene that encodes it does not work (Haploinsufficiency) and a neat therapy would be to make the remaining copy of that gene work harder. When I originally looked at epigenetics I thought it would not be possible to epigenetically tag the good copy of the specific gene, to switch it on. However it seems that we do not need to tag a specific gene, just provide the “post-it” notes and let the body do the tagging.
All this leads to the use of HDIs to treat cancer, leaving the body to figure out the hard part of which genes.  In reality an HDI will change the expression of numerous genes, not just the one(s) you wanted.

Different Colours of Tags
Just as those useful Post-It notes come in multiple colours, epigenetic markers come in different varieties.  This has been well studied in the cancer research.
HDAC1 inhibitors only affect part of the epigenome; there are other modifiers that are required to affect other genes.
In autism, as in cancer, you need to know which genes are miss-expressed and then you can see if an epigenetic therapy exists that covers them.  Put more simply if HDAC1 inhibitors affect only yellow post-its, which cancers/autisms would become treatable?
The more complex explanation regarding different colours of post-its:








“Important epigenetic modifications known to regulate gene expression. a DNA methylation of CpG islands in promoter regions by DNA methyltransferases (DNMT) represses gene activity. Posttranslational covalent histone modifications of lysine (K), arginine (R) or serine (S) residues in the “histone tail” also influence gene expression in different ways. b Histone acetylation (Ac) catalysed by histone acetyltransferases (HAT) is usually correlated to increased gene activity, whereas histone deacetylation caused by histone deacetylases (HDAC) is considered to decrease gene expression, even though histone hyperacetylation not always matches regions of increased gene activity. c Histone methylation (Me) and demethylation by histone methyltransferases (HMT) and histone demethylases (HDM) at lysine or arginine residues show different effects on gene activity depending on number and position of methyl groups. d Histone ubiquitinylation (Ub) at lysine residues alters histone structure and allows access of enzymes involved in transcription. e Histone phosphorylation (P) at distinct serine residues is known to be associated with increased gene expression, and it is also involved in DNA damage response and chromatin remodelling. Phosphorylation at linker histone (LH) H1 is considered to be a signal for the release of histone H1 from chromatin. In general, epigenetic regulation depends on the addition of epigenetic marks by writer enzymes (e.g. DNMT, HMT, HAT) and the removal of these marks by epigenetic eraser enzymes (e.g. HDAC and HDM) as well as epigenetic reader enzymes (not shown in this figure)”


Treating cancer is always going to be more difficult than treating autism because by the time it has been identified a whole cascade of changes is already underway and whereas autism is not degenerative, cancer by definition is. So even a very partially effective cancer drug might be potent enough for autism, or just a tiny dose of an effective cancer drug.





This post is about HDAC1&2 / yellow Post-its 

1.  The Grant Application 

The goal of this study is to discover novel, mechanism-based pharmacological intervention for autism, a devastating neurodevelopmental disorder with no treatment currently. Genetic sequencing has revealed extensive overlap in risk genes for autism and for cancer, many of which are chromatin remodeling factors important for transcriptional regulation, suggesting the possibility of repurposing the anti-cancer drugs targeting epigenetic enzymes for autism treatment. ASDDR LLC and Yan Lab at SUNY-Buffalo propose to jointly investigate the hypothesis that histone deacetylase (HDAC) inhibitors are able to restore the expression of key autism risk factors and induce long-lasting rescue of autism-like behavioral and synaptic deficits. Combined behavioral, biochemical and electrophysiological approaches will be used to address two specific aims. 

Aim 1. To discover HDAC inhibitors that can alleviate autism-like behavioral deficits in autism mouse models. Yan lab screened a number of drugs and found that a brief treatment with the highly potent and class I-specific HDAC inhibitor, romidepsin (Istodax, an FDA-approved anti-cancer agent) at the very low dose, led to dramatic and prolonged rescue of the social deficits in the Shank3-deficient mouse model of autism. To determine whether this pharmacological agent can serve as a tool compound for autism drug development, its therapeutic efficacy and safety will be examined in two different models of autism, Shank3-deficient mice and BTBR mice.

Aim 2. To identify the molecular targets of HDAC inhibitors as benchmarks for the treatment of autism. For the discovery of effective drugs to treat autism, the molecular pathways on which HDAC inhibitors act to alleviate the autism-like behavioral deficits in Shank3-deficient mice need to be understood. We will reveal the potential benchmark, such as actin regulators and NMDARs, as molecular targets of romidepsin. This phase I preclinical study will provide great promise for the discovery of new and effective pharmacological agents to treat the social interaction deficits, a core symptom of autism.

Public Health Relevance

This project is to discover novel, mechanism-based therapeutic strategies for autism. The corporate and academic partners propose to jointly investigate the hypothesis that histone deacetylase (HDAC) inhibitors are able to restore the expression of key autism risk factors and induce long-lasting rescue of autism-like behavioral and synaptic deficits.

2. Study Press Release 

Using an epigenetic mechanism, romidepsin restored gene expression and alleviated social deficits in animal model of autism 
 “The advantage of being able to adjust a set of genes identified as key autism risk factors may explain the strong and long-lasting efficacy of this therapeutic agent for autism.”
BUFFALO, N.Y. — Of all the challenges that come with a diagnosis of autism spectrum disorder (ASD), the social difficulties are among the most devastating. Currently, there is no treatment for this primary symptom of ASD. New research at the University at Buffalo reveals the first evidence that it may be possible to use a single compound to alleviate the behavioral symptoms by targeting sets of genes involved in the disease.

The research, published today in Nature Neuroscience, demonstrated that brief treatment with a very low dose of romidepsin, a Food and Drug Administration-approved anti-cancer drug, restored social deficits in animal models of autism in a sustained fashion.

The three-day treatment reversed social deficits in mice deficient in a gene called Shank 3, an important risk factor for ASD. This effect lasted for three weeks, spanning the juvenile to late adolescent period, a critical developmental stage for social and communication skills. That is equivalent to several years in humans, suggesting the effects of a similar treatment could potentially be long-lasting, the researchers say.
Profound, prolonged effect
“We have discovered a small molecule compound that shows a profound and prolonged effect on autism-like social deficits without obvious side effects, while many currently used compounds for treating a variety of psychiatric diseases have failed to exhibit the therapeutic efficacy for this core symptom of autism,” said Zhen Yan, PhD, professor in the Department of Physiology and Biophysics in the Jacobs School of Medicine and Biomedical Sciences at UB, and senior author on the paper.

The study builds on her previous research from 2015. That work revealed how the loss of Shank 3 disrupts neuronal communications by affecting the function of the NMDA (n-methyl-D-aspartate) receptor, a critical player in regulating cognition and emotion, leading to deficits in social preference that are common in ASD.
In the new research, the UB scientists found they could reverse those social deficits with a very low dose of romidepsin, which, they found, restores gene expression and function using an epigenetic mechanism, where gene changes are caused by influences other than DNA sequences. Yan noted that human genetics studies have suggested that epigenetic abnormalities likely play a major role in ASD.
To pursue these promising findings, Yan has founded a startup company called ASDDR, which was awarded a Small Business Technology Transfer grant from the National Institutes of Health last summer for more than $770,000.
Epigenetics in ASD
Many of the mutations in ASD, Yan explained, result from chromatin remodeling factors, which are involved in dynamically changing the structure of chromatin, the complex of genetic material in the cell nucleus that condenses into chromosomes.
“The extensive overlap in risk genes for autism and cancer, many of which are chromatin remodeling factors, supports the idea of repurposing epigenetic drugs used in cancer treatment as targeted treatments for autism,” said Yan.
She and her colleagues knew that chromatin regulators — which control how genetic material gains access to a cell’s transcriptional machinery — were key to treating the social deficits in ASD, but the challenge was to know how to affect key risk factors at once.
“Autism involves the loss of so many genes,” Yan explained. “To rescue the social deficits, a compound has to affect a number of genes that are involved in neuronal communication.”
To do so, the team turned to a type of chromatin remodeler called histone modifiers. They modify proteins called histones that help organize genetic material in the nucleus so gene expression can be regulated. Since many genes are altered in autism, the UB scientists knew a histone modifier might be effective.
Loosening up chromatin
In particular, they were interested in histone deacetylase (HDAC), a family of histone modifiers that are critically involved in the remodeling of chromatin structure and the transcriptional regulation of targeted genes.
“In the autism model, HDAC2 is abnormally high, which makes the chromatin in the nucleus very tight, preventing genetic material from accessing the transcriptional machinery it needs to be expressed,” said Yan. “Once HDAC2 is upregulated, it diminishes genes that should not be suppressed, and leads to behavioral changes, such as the autism-like social deficits.”
But the anti-cancer drug romidepsin, a highly potent HDAC inhibitor, turned down the effects of HDAC2, allowing genes involved in neuronal signaling to be expressed normally.
 “The HDAC inhibitor loosens up the densely packed chromatin so that the transcriptional machinery gains access to the promoter area of the genes; thus they can be expressed,” Yan said.
The rescue effect on gene expression was widespread. When Yan and her co-authors conducted genome-wide screening at the Genomics and Bioinformatics Core at UB’s New York State Center of Excellence in Bioinformatics and Life Sciences, they found that romidepsin restored the majority of the more than 200 genes that were suppressed in the autism animal model they used.
“The advantage of being able to adjust a set of genes identified as key autism risk factors may explain the strong and long-lasting efficacy of this therapeutic agent for autism,” Yan explained. She and her colleagues will continue their focus on discovering and developing better therapeutic agents for autism.  

Full study:-  


HDAC Inhibitors
HDIs have a long history of use in psychiatry and neurology as mood stabilizers and anti-epileptics. More recently they are being investigated as possible treatments for cancers, parasitic and inflammatory diseases. 
HDAC inhibitors have effects on non-histone proteins that are related to acetylation. HDIs can alter the degree of acetylation of these molecules and, therefore, increase or repress their activity.
“To carry out gene expression, a cell must control the coiling and uncoiling of DNA around histones. This is accomplished with the assistance of histone acetyl transferases (HAT), which acetylate the lysine residues in core histones leading to a less compact and more transcriptionally active chromatin, and, on the converse, the actions of histone deacetylases (HDAC), which remove the acetyl groups from the lysine residues leading to the formation of a condensed and transcriptionally silenced chromatin. Reversible modification of the terminal tails of core histones constitutes the major epigenetic mechanism for remodeling higher-order chromatin structure and controlling gene expression. HDAC inhibitors (HDI) block this action and can result in hyperacetylation of histones, thereby affecting gene expression.[5][6][7] The open chromatin resulting from inhibition of histone deacetylases can result in either the up-regulation or the repression of genes.”


Pitt Hopkins Research
We saw that transcription factor TCF4 (the Pitt Hopkins gene) is also lacking in some MR/ID and schizophrenia. We saw in an earlier post that TCF4 can be upregulated by PKA (protein kinase A) and that this can be achieved using a PDE4 inhibitor as used to treat asthma and COPD. So in theory Daxas should help.
The lack of the TCF4 protein in Pitt Hopkins causes a cascade of other genes to be miss-expressed. The logical thing to do is to correct that miss-expression. 
The Shank3 research is not the first to suggest that HDAC inhibition as a potentially viable therapy. In 2016 the same idea was suggested for Pitt Hopkins and while this is a rare condition, milder dysfunctions of the same TCF4 gene are seen as common in MR/ID and indeed in schizophrenia. So HDAC inhibition may be a viable therapy for many people.

HDACi meds may reverse effects of Pitt Hopkins


In a paper published this week by the journal Cell Reports, Sweatt and his colleagues at the University of Alabama at Birmingham (UAB) report that mice deficient in Tcf4 exhibit impairments in social interaction, vocalization, learning and memory characteristic of PTHS.
The impairments were “normalized” when the mice were given small-molecule drugs called HDAC inhibitors, which alter Tcf4-associated gene expression in the brain. The finding suggests that “broadly acting, epigenetically targeted therapeutics … might be particularly beneficial in PTHS patients,” the researchers concluded.
“We are quite excited by these findings, said Sweatt, a Vanderbilt University-trained pharmacologist who formerly chaired the Department of Neurobiology and directed the McKnight Brain Institute, both at UAB.
“Pitt-Hopkins Syndrome is an orphan disease that has not been extensively studied,” he said. “Having identified one potential avenue for possible therapeutics is an important step forward.”

“Nearly one-quarter of the genes dysregulated in the Tcf4(+/−) mice are also regulated by HDAC inhibition. The strong negative correlation between Tcf4(+/−) and CI-994 DEGs (R2 = 0.72) suggests HDAC inhibition is a viable avenue for correcting a large percentage of transcriptional dysregulation associated with Tcf4 haploinsufficiency.”


Which HDAC Inhibitor?
It should be noted that Romidepsin inhibits both HDAC1 and HDAC2.
There are HDACs numbered 1 through 10.
HDAC inhibitors vary in potency. Below is a chart comparing different HDI drugs in the activation of HIV expression.

In vitro activation of HIV expression by HDAC inhibitors in an in vitro latency model.

  
The role of diet
I know that many readers of this blog like dietary interventions and do not like drugs.
In cancer I think diet can be preventative rather than therapeutic or curative. Once cancer takes hold you need very potent therapies.
In dementia it looks like diet can be preventative and therapeutic.
In mild ADHD and mild autism it looks like dietary intervention can be sufficient.  
Many flavonoids have mild epigenetic properties. They are unlikely to be potent enough to halt the cascade of changes seen in a runaway cancer, but they may well be chemoprotective, i.e. they prevent cancer developing in the first place.
Since in some autism we only need a relatively mild  effect perhaps flavonoids do have some potential, depending on which genes are miss expressed.


Food containing high amounts of epigenetically active flavonoids

Description
Class
Flavonoid
Ǿ mg/100 g
Sources of data
Grapefruit, raw (not specified as to colour) (Citrus paradisi)
Flavanones
Hesperetin
1.50
aUSDA Database for the Flavonoid Content of Selected Foods: e.g. [193]


Naringenin
53.00

Flavonols
Kaempferol
0.40


Quercetin
0.50
Onions, red, raw
Flavones
Apigenin
0.24
aUSDA Database for the Flavonoid Content of Selected Foods: e.g. [193, 194, 195, 196]


Luteolin
0.16

Flavonols
Kaempferol
0.70


Myricetin
2.16


Quercetin
39.21
Soybeans, mature seeds, raw (all sources)
Isoflavones
Daidzein
62.07
bUSDA Database for the Isoflavone Content of Selected Foods: e.g. [197, 198, 199, 200, 201, 202]


Genistein
80.99
Spices, parsley, dried (Petroselinum crispum)
Flavones
Apigenin
4503.50
aUSDA Database for the Flavonoid Content of Selected Foods: e.g. [196]


Luteolin
19.75
Strawberries (including frozen unsweetened strawberries)
Flavonols
Fisetin
16
[203]


Kaempferol
0.49
aUSDA Database for the Flavonoid Content of Selected Foods: e.g. [204, 205]


Myricetin
0.35


Quercetin
0.46
Cacao beans
Flavan-3-ols
(+)-Catechin
88.45
aUSDA Database for the aFlavonoid Content of Selected Foods: e.g. [206]
(−)-Epicatechin
99.18
Tea, black, brewed, prepared with tap water
Flavan-3-ols
(+)-Catechin
1.51
aUSDA Database for the Flavonoid Content of Selected Foods: e.g. [196, 207, 208, 209]


(−)-Epigallocatechin 3-gallate
9.36

Flavonols
Kaempferol
1.41


Myricetin
0.45


Quercetin
2.19
Tea, green, brewed, decaffeinated
Flavan-3-ols
(−)-Epigallocatechin 3-gallate
26.05
aUSDA Database for the Flavonoid Content of Selected Foods:

Flavonols
Kaempferol
1.00


Myricetin
1.00


Quercetin
2.77


A good example is EGCG 
In earlier posts on EGCG, being trialed in Spain on Down Syndrome and Fragile X; I was intrigued by the its long-lasting effects: 

For most of the tests (21 of 24) there were no differences between the groups. 
However, in three tests people who'd taken EGCG did better. This improvement lasted for six months after the study ended


Another example is Sulforaphane (sometimes)
It appears that some people taking sulforaphane experience disease changing results, which are likely caused by the epigenetic effects of inhibiting HDAC. 


Summarized Case Reports

A.    Three participants who took SF did not appear to improve during the study. Their parents reported lack of a noticeable effect and were not aware whether their young adults had been taking SF or placebo.

B.     One participant no longer uses SF. However, he improved dramatically while taking it during the study and remained “improved” after the study, suggesting to the study team a possible “epigenetic switch” might have been triggered.
“W is doing fantastic. He really turned into the most relaxed and fantastic child (on sulforaphane). Definitely something great. Helped him a lot. His friends, family, and members at his home all noticed a wonderful change. He is off the sulforaphane and has been since the end of his study in 2012.”
  
Perhaps Butyrate?  


As interest in the gut microbiome has grown in recent years, attention has turned to the impact of our diet on our brain. The benefits of a high fiber diet in the colon have been well documented in epidemiological studies, but its potential impact on the brain has largely been understudied. Here, we will review evidence that butyrate, a short-chain fatty acid (SCFA) produced by bacterial fermentation of fiber in the colon, can improve brain health. Butyrate has been extensively studied as a histone deacetylase (HDAC) inhibitor but also functions as a ligand for a subset of G protein-coupled receptors and as an energy metabolite. These diverse modes of action make it well suited for solving the wide array of imbalances frequently encountered in neurological disorders. In this review, we will integrate evidence from the disparate fields of gastroenterology and neuroscience to hypothesize that the metabolism of a high fiber diet in the gut can alter gene expression in the brain to prevent neurodegeneration and promote regeneration.

  

In general, these data suggest that BT can enhance mitochondrial function in the context of physiological stress and/or mitochondrial dysfunction, and may be an important metabolite that can help rescue energy metabolism during disease states. Thus, insight into this metabolic modulator may have wide applications for both health and disease since BT has been implicated in a wide variety of conditions including ASD. However, future clinical studies in humans are needed to help define the practical implications of these physiological findings.

Conclusion
Clearly HDAC inhibitors are beneficial in some cancer and some autism.
In cancer the dose required is so high there almost inevitably will be some side effects, particularly in people already in poor health.
Hopefully when Dr Yan moves on to trial Romidepsin in her second mouse model, the BTBR model, she will be as successful as with the Shank3 model.
Ultimately, I assume she will trial her low dose Romidepsin as a single dose in humans. I am sure plenty of people will be interested in that, including all the Pitt Hopkins families. Hopefully someone will trial Daxas in Pitt Hopkins (upregulate PKA which then upregulates TCF4).
Dietary HDAC inhibitors include butyrate and sulforaphane. They are much weaker than Romidepsin. Would a very large dose of sulforaphane/butyrate have the potency of a small dose of Romidepsin?
To be effective in autism the HDAC inhibitor would have to freely cross the blood barrier, clearly drugs used to treat brain cancer tick this box.
Vorinostat/Zolinza also looks interesting.
We should not overlook Valproic acid, another HDAC inhibitor. This epilepsy drug can cause autism when taken during pregnancy, but is taken by some children with autism. Unlike Romidepsin and Vorinostat, which are hugely expensive, Valproic acid is cheap.
Continued use of Valproic acid can cause side effects, as seen in the comments section of this blog. A short sharp shock with valproic acid might be different.
I am sure Dr Yan chose Romidepsin for its potency. A small dose of Romidepsin is likely much more effective than a bucket load of broccoli sprouts (sulforophane).  
Just how low a dose is Dr Yan talking about? Recall that Professor Catterall’s  low dose of clonazepam (to modulate alpha3 subunits of GABAa receptors) was so low in humans it has none of the well-known drawbacks of benzodiazepine use (addiction, tolerance etc).
Dr Naviaux’s use of Suramin was long thought to be impractical in humans due to side effects, but now this appears not to be the case.
Back to Dr Yan:- 
Social deficits in Shank3-deficient mouse models of autism are rescued by histone deacetylase (HDAC) inhibition
Treatment with the HDAC inhibitor romidepsin lastingly relieves autism-like social deficits in Shank3-deficient mice. The level of global H3 acetylation (Fig. 1a) in the frontal cortex of Shank3+/ΔC mice was significantly lower than that from wild-type (WT) mice. 
 A systemic administration of low-dose romidepsin (0.25 mg/kg, intraperitoneally (i.p.), once daily for 3 d), a highly potent and brainpermeable class I-specific HDAC inhibitor (with nanomolar in vitro potency25) approved by the US Food and Drug Administration (FDA) for cancer treatment26–28, significantly elevated the level of acetylated H3 in Shank3+/ΔC mice, while it had little effect in WT mice. These data suggest that Shank3-deficient mice have an abnormally low level of histone acetylation, which can be restored by romidepsin treatment. 

This dose looks like about one tenth of the used in mice in cancer trials.
In humans, Romidepsin is for intravenous infusion only. Each 10 mg single-use vial of Romidepsin/Istodax costs about $2,800.
Vorinostat/Zolinza costs about $3,800 for 30 capsules.
If the autism effects of a potent HDAC1/2 inhibitor can last for several years in humans, as suggested by Dr Yan, and if the dose is a tenth of the cancer dose, the cost would not seem to be such a barrier.
The open question is the safety profile of Romidepsin at a single low dose in otherwise physically healthy children.

Risk vs Reward
While nobody wants side effects, one has to consider the risk versus the reward. In some single gene types of severe autism it is clear what the outcome with no intervention will be; perhaps that looming outcome warrants taking a bigger risk than someone with mild autism struggling with social difficulties? But then again, perhaps an HDAC1/2 inhibitor might improve social functioning so someone with Asperger’s, or indeed schizophrenia, does not commit suicide?






36 comments:

  1. Of all the papers I have read on HDACs, comparing them to different colored Post-It notes has to be the most elegant description I have seen to date.

    On another note, polyamines strongly influence chromatin structure as well and overall polyamine levels can be significantly increased via diet, while individual ratios of one type of polyamine or another is largely a function of enzymes which interchangeably convert putrescine to spermidine to spermine and vice versa.

    Also, I found this product on another discussion board which I had never seen before which might be an alternative to eating very large amounts of wheat germ or green peas (both high in spermidine):

    https://www.amazon.com/Spermidine-Free-Base-5-Grams/dp/B0771SJFSQ/

    I have never used this product myself and it is certainly not cheap as well as the fact I have no idea if it is intended to be used orally or intravenously, but at least it is an option assuming the product quality is sufficient.

    ReplyDelete
    Replies
    1. Tyler, I clarified my question in the previos post,last comment
      Valentina

      Delete
  2. Thanks for yet another interesting post Peter!

    HDAC inhibitors are definitely one lane of treatments worthy investigation, but the research on them vs CNS related dysfunctions is still limited. This post also points out that choosing wrong might potentially have a bad effect for a long time. You just don't know what else you are up- or downregulating.

    PDE4 inhibitors and PKA upregulation does look like a way to upregulate the SATB2 gene (I only have indirect indications for this) and if my current trial fails I will have to try this route (again). Except for Daxas/roflumilast we have read about Ibudilast, a pan-selective PDE4 inhibitor and pterostilbene, selective for PDE4D, so plenty to choose from. I think also cinnamon has a positive effect on PKA(?)

    Sulforaphane has epigenetic effects on (pro-) bone formation, as does vitamin K2

    /Ling

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    Replies
    1. Maybe you alread mentioned this somewhere Peter, but my eyes just fell on a PDE4 inhibitor approved for atopical dermatitis in children 2+ years:
      Crisaborole 2% ointment
      https://www.ncbi.nlm.nih.gov/pubmed/29248522

      It probably doesn't have much effect elsewhere, but who knows?

      /Ling

      Delete
    2. Ling, it is a very new drug and I think it only approved in the US. I think you would need an oral drug to affect the brain. Atopic dermatitis affects 20% of school kids so it is a huge market. You would think that since many kids have asthma somebody would get a PDE4 inhibitor approved for this application.

      As you can see in the paper below, side effects are big issue, but this is highly dose dependent.

      Recent Advances Using Phosphodiesterase 4 (PDE4) Inhibitors to Treat Inflammatory Disorders: Animal and Clinical Studies
      https://www.researchgate.net/publication/303398752_Recent_Advances_Using_Phosphodiesterase_4_PDE4_Inhibitors_to_Treat_Inflammatory_Disorders_Animal_and_Clinical_Studies?enrichId=rgreq-a02395fdfb6993f68960174c147b2f04-XXX&enrichSource=Y292ZXJQYWdlOzMwMzM5ODc1MjtBUzozNzk5NDQ5ODgzNjQ4MDBAMTQ2NzU5NzM1MjIxMQ%3D%3D&el=1_x_3&_esc=publicationCoverPdf

      At one fifth of the COPD dose we saw sensory gating was resolved with no side effects, using Roflumilast. I think this should be trialed in adults with ASD.

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  3. Hi Peter, Ling and All

    This is indeed very interesting. Ling, I’ve had the same concerns about long term undesired effects if wrong HDAC inhibitor is used.

    With regards to valproic acid, one well known adverse effect is ammonia level increase. While in most people this does not lead to visible symptoms, it seems like ammonia is associated with neuronal chloride regulation:
    “we show that ammonia rapidly compromises astrocyte potassium buffering, increasing extracellular potassium concentration and overactivating the NKCC1 in neurons.”
    https://www.ncbi.nlm.nih.gov/pubmed/24240184

    “These findings suggest that long-term exposure to ammonia increases the expression of AE3 through the activation of PKC, resulting in an increase in [Cl-]i in neurons and a reduction of inhibitory postsynaptic potentials.”
    https://www.ncbi.nlm.nih.gov/pubmed/9739146

    Peter, do you think it is not of concern with the “short sharp shock with valproic acid” that you proposed?

    Also, in these studies they reduce ammonia harmful effects with bumetanide or acetazolamide, but can we think opposite: if ammonia production could be somehow decreased, can we expect improvement in neuronal chloride regulation? And better effects of bumetanide or Diamox, not only in valproic acid users? Or is it oversimplified?

    Most studies are on toxic ammonia levels, but I wonder how this relates to lower levels and autism/neuronal chloride regulation relation.

    Treatments used for valproic hyperammonemia are: carnitine, arginine, carglumic acid.

    Do you have practical conlcusion on butyrate use? How can I compare its impact on HDAC with beta hydroxybutyrate? BHB is also HDAC1 inhibitor highly regarded by ketogenic diet/longevity connection believers. While I don't know how to compare their strength, I can easily compare the price :/

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    1. Agnieszka, I think it is fair to say that several different mechanisms may lead to elevated neuronal chloride which will lead to symptoms diagnosed as autism. So elevated chloride is just another "Gargus" nexus for autism.

      Does being a Bumetanide/Diamox responder mean you have elevated ammonia? I do not think it necessarily does, but it does mean you want to avoid elevating ammonia.

      Long term use of valproate in a bumetanide responder might lead to ammonia becoming a problem, so we might expect those people who respond well to valproate for a while to later regress, as you found.

      My short sharp shock with valproate might be so brief that AE3 expression did not change and so chloride levels might not change. This would have to be studied, but I think a researcher will seek a drug that is easier to patent/restrict access to. I think Romidepsin is a good choice for Dr Yan, I think it would not suit her to show that all you might need is to drink a bottle of valproate syrup.

      My long term use of an HDAC inhibitor, was broccoli/sulforaphane. After years of use, it no longer seems to have any effect. Is that because after all this time an epigenetic switch has finally made a permanent change? That would be nice, if it was true.

      Butyrate clearly has many potential beneficial effects, but it seems some of these effects can be found only at a low dose. So this requires a personalized approach.

      Interestingly one of the many benefits of exercise comes from β-hydroxybutyrate.

      Exercise promotes the expression of brain derived neurotrophic factor (BDNF) through the action of the ketone body β-hydroxybutyrate.
      https://www.ncbi.nlm.nih.gov/pubmed/27253067

      Exercise seems to have the potential to cause permanent changes, which is one reason for young people to do aerobic exercise for their future long term health.

      I have no idea of the relative potency of sodium buytrate vs β-hydroxybutyrate.

      On a simplistic level, the dose of β-hydroxybutyrate used for the KD is far higher than the “suggested” sodium butyrate dose. I wonder what has happened to long term users.

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    2. LOLA (L-Ornithine L-Aspartic) therapy can help reduce ammonia.

      Delete
    3. Tyler, that is interesting. I see that when given intravenously to humans/animals with very high ammonia it does indeed work. I suppose when taken orally by humans with only slightly high ammonia it might well help.

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    4. Thanks Peter and Tyler,

      It seems to me that ammonia testing should be a part of routine diagnostics in autism then. Some readers commented here about high ammonia levels in their children, but I can't find it now. I haven't heard about LOLA before.

      β-hydroxybutyrate has interesting properties and other ketone bodies as well:
      https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4176946/

      "Ketone bodies as signaling metabolites"

      There are children with epilepsy treated with ketogenic diet are later weaned off KD without loss of seizure control, perhaps due to epigenetic switch?

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    5. I would like to know if the people who think their child has elevated ammonia have actually done the standard test to measure ammonia in blood. This is an inexpensive test, about $10.

      How will measuring anything in your urine tell you, with certainty, what is the level of ammonia in your blood? Elevated ammonia in your brain will do no good at all, but it can only get there across the BBB from your blood.

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    6. Just to clarify my comment, I meant blood ammonia level and this comment:

      https://epiphanyasd.blogspot.com/p/polypill-for-autism.html?showComment=1523557220273#c4329947534977054070

      The test is inexpensive, but where I live it turned out that it's done mostly in a hospital setting as the test needs to be run immediately after sample is taken.

      I don't know if urinary ammonia is relevant.

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    7. Agnieszka, it looks like "high ammonia" is sometimes a DAN-type concept regardless of what the blood test shows. The favoured lab in the US does indeed have a urine test to diagnose high ammonia. If you google ammonia and autism you will find a mixture of science, pseudo-science and nonsense "I did not test ammonia in his blood because his urine smelt so bad I knew it was ammonia" and then spends a decade trying to "lower ammonia" and presenting her ideas at conferences.

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    8. Agnieszka, first line treatment here for high ammonia, hepatic encephalopathy, mainly due to alcoholic fatty liver, is Lactulose, also used for constipation.

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    9. Thanks, I forgot about it.

      I suspected valproic acid induced hyperammonemia rather than alcoholic fatty liver... but it seems that lactulose is also used in such situations. My main concern was that ammonia may increase chloride neuronal concentration, which is something to avoid at least in some autism and probably it may happen even without sky-high ammonia levels.

      Interestingly, Joan Fallon patented lactulose for autism.

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    10. Peter,

      It seems doctors here in India are more thorough and sensible as the neurologist we met wrote blood ammonia, lactate, pyruvate and host of other tests and said these are the basic tests that every individual coming with a developmental issue should undergo.

      My sons blood ammonia levels were normal.

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    11. Kritika, in many countries including the UK a child with autism would only get to see a neurologist if there were seizures. So big city healthcare in India is more advanced, and this is not just for autism. Free care is not always the best care.

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  4. Peter, What do you think of using domperidone, less than 30 mg a day as 5TH3 antagonist? It decreases dopamine release and it is used in schizophrenia Also mint and boldine are 5TH3 antagonists.
    Valentina

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    1. Valentina, I think that this might just create new problems, but it is thought that this anti-nausea drug can work as an anti-psychotic. Mint has many effects.

      "The prevailing theory is that antipsychotics work by blocking D2 receptors. We see that domperidone between 30 and 160 mg/d could function as an antipsychotic."
      https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3962062/

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    2. Valentina, you could look at Mirtazapine, which did seem to greatly help one reader of this blog. It is normally used as an antidepressant, but it affects 5TH3, H1 and other receptors. At different doses you get different effects, so at 5mg it is a strong sedating antihistamine. At 30mg it is an antidepressant and much less sedating.

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    3. Peter,when I think clearly,giving to my son anything that works as an antipsichotic,with direct or undirect antidopaminergic effect, after having defeated dyskinesias,is really crazy.Next week I will get sarcosine and will see.Incredibly I had luck with bumetanide which it is a great thing!
      Valentina

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  5. Kritika and others interested: we stopped the Bimuno experiment because my son developed that pesky red rash above upper lip (just below nostrils). I’m convinced this is an infection type of rash. He had the rash before after trying certain probiotics (back then I was wondering if verapamil was to blame). So - I’m starting to think there is some pathogenic bacteria (or even virus?) that just loves supplemental prebiotic and certain strains of probiotics. Why not? Who ever studied with absolute certainty otherwise? There still is so much unknown about the microbiome. So anyway - now on to killing some weeds for a few days to get him back to baseline and never again with prebiotics or probiotics. Permanently crossed off list. He will just get his pre and pro bios his natural, usual way - healthy diet, fresh air and lots of time outdoors. On a positive note: no gut pain/gas whatsoever from b-GOS so that claim holds water in our case at least.

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    1. Thanks Tanya,probiótics and prebiotics inulin and pectin are also crossed for us, like antipsychotics and antibiotics.
      Valentina

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    2. Thanks Tanya,

      So Bimuno goes out too. I was reading that a trial is underway to test the efficacy of bimuno in ameliorating autistic symptoms in a group of kids. I think we already have the results for our kids.

      So crossing off my list herbs, probiotics, prebiotics, vitamins and minerals. EPA?

      Peter, any updates on EPA?

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    3. Kritika, anyone trialing EPA is very welcome to share their experiences.

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    4. Hi Peter, Kritika and community,

      I've had my daughter on EPA (and I took her off DHA) for about 2 weeks now, and maybe its too early to try to see results, but we've noticed an improvement in the complexity of her speech and an improvement in her behavior.

      In terms of behavior, we had been getting some feedback that she was being a bit more fussy / cranky / tempermental prior to EPA, and I attributed this to having recently started Methionine (it's the only thing I could think of).

      I stopped Methionine at the same time as I started EPA, and it has been a positive. The improvements are noticeable, but at the same time, not a "WOW" either (at least not yet). I will continue to use EPA as it has been beneficial in my eyes, and I'm hoping to see better results after 2 - 3 months.

      In terms of Bimuno, I've been using this for a year, no negatives at all, and no obvious positives. I found it when I looked through clinical trials for ASD to see what was being trialed. Since it doesn't hurt, and may be good for my daughter's GI (although she never had GI issues), I keep using it.

      My black garlic also just arrived (sat in customs for a while for no reason ....), and I will likely start using that on the weekend.

      I'm sending in my OAT test and my Amino Acids test hopefully tomorrow. For those of you in Canadian provinces with LifeLabs, for the Amino Acids, go to LifeLabs for and look for a test called Metabolic Screen Amino Acids, $60 CDN and you need a doctor's requisition. You can find it on their website. I actually got a copy of the reference intervals from the lab they outsource it to, it looks pretty comprehensive, easily worth $60.

      Also should be getting my genetic test results in the next week ... the results could lead me down completely new avenues.

      AJ

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    5. AJ, you really get me curious about that genetic test too. Remember, IF you get an answer now, you will might have to learn if it is a gain-of-function or loss-of-function before you can decide what to do. And do keep us posted!
      /Ling

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    6. Hi Ling,

      You're exactly right, that's what I'm really curious about. The other possibility is that there is a Copy Number Variation (CNV) such that my daughter is getting a different "dose" of the relevant protein than what she should be getting. If there is a CNV, and she is getting too much of something (which I think is easier to deal with than a loss-of-function mutation, depending on downstream effects), then I'd just have to find an antagonist / inhibitor of the relevant protein.

      I will definitely share the findings with the community, and would appreciate any input anyone has if there is a relevant finding.

      Have a great day Ling!

      AJ

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  6. My favourite epigenetic intervention is parental care (maternal care is the name most often used though).

    "Here we report that TSA and methionine treatment reversed the effect of maternal care on open-field behavior. We identified >900 genes stably regulated by maternal care. A fraction of these differences in gene expression is reversible by either the histone deacetylase inhibitor TSA or the methyl donor l-methionine. These results suggest that early-life experience has a stable and broad effect on the hippocampal transcriptome and anxiety-mediated behavior, which is potentially reversible in adulthood."
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1413873/

    Co-sleeping anyone?

    /Ling

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    1. Ling, nurture is very important and we saw in Prof Nelson's epic study in Bucharest that lack of nurture/emotional bonding causes long term harm, measurable MRI brain variations and even autistic behaviours.

      https://epiphanyasd.blogspot.com/2016/12/quantifying-benefits-of-stimulation.html

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    2. Ling,

      Co-sleeping....yes. For six years and elaborate co sleeping with singing and gentle rhythmic patting on the back. It was fairly common here but now so out of fashion.

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    3. Valentina, yes same here with antibiotics - except for xifaxan - that was the only beneficial one, but only for one course. And antipsychotic only last resort, crisis situation. When all else fails. And probably in combination . The neuroscientist PhD md we consulted with said pediatricians really shouldn’t be prescribing these medicines

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    4. Tanya, there is a study of Shangai Mental Health Centre of donepezil in combination with lithium for acute mania.Good to know.Valentina

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  7. I am having trouble posting here - my replies go to bottom instead of to the right thread. Hmm should I take the hint ;)

    My son’s ammonia in urine was always high - common in those with homozygous mthfr a1298c . Ornithine never brought it down, neither did citrulline - but we didn’t try ornithine and aspartic combo. The only thing that moved the market lower was lowering his dietary protein

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  8. Thank you Valentina!! I hope this reply goes through - having a hard time posting

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  9. Agnieszka, maybe it was useless to consider, we’ve gone down many rabbit holes I will admit - but the doctor we were working with at the time checked it as an overall treatment plan for his gut issues. His blood ammonia was always normal when we were with that particular DAN doctor who checked.
    Info on urine ammonia testing from mayo:
    https://www.mayomedicallaboratories.com/test-catalog/Clinical+and+Interpretive/62657

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