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Thursday 24 June 2021

Betaine (TMG) and Gene Therapy as potential alternatives to Bumetanide Treatment in Autism?


Betaine (also known as TMG, or trimethylglycine) is a methyl derivative of glycine, first isolated from sugar beet and hence its name.

Today’s post was prompted by our reader, and Covid home-school instructor, AJ.  He raised the question of whether betaine can be used like Bumetanide to normalize chloride levels in neurons.

I am combing this idea with news from Genoa in Italy, where they have developed gene therapy as an alternative to Bumetanide and in their words :-

“This sets the stage for the development of a gene therapy approach to overcome the shortcomings of bumetanide treatment.”

The interesting thing is that neither of these ideas come from autism research.  The idea to use Betaine was stumbled upon and was then written up in a Norwegian case study about Creatine transporter deficiency.  The Italians are trying to improve cognition in brain disorders and their model of choice was Down syndrome. 

As we have seen time and again, elevated chloride within neurons is a common feature of many types of brain disorders from some idiopathic autism, to Down syndrome, to adult conditions such as Parkinson’s disease.  Today we learn that it is may well be a feature of Creatine Transporter Deficiency.

I have been rather wary of writing about any kind of gene therapy, because it seemed either too far ahead of its time, or just absurdly expensive.  There are some new $1+ million treatments.

This may be about to change given that the Biontech (AKA Pfizer vaccine), Moderna, Janssen (Johnson & Johnson) and Oxford AstraZeneca vaccines for Covid 19 are all based on gene therapy.

The Biontech people are really clever and were already trying to treat various kinds of cancer and other condition using gene therapy, before they developed their highly successful Covid vaccine.

The Italians in Genoa used an adeno-associated virus (AAV)-mediated RNA interference (RNAi) to target and reduce neuronal NKCC1 expression, rescue neuronal Cl-  homeostasis, GABAergic transmission, and cognitive deficits.   The benefit was still there 6 months after the injection.

Don’t worry if the above paragraph makes little sense. Just read on.

The same type of adeno-associated virus (AAV) vector is the platform for gene therapy delivery used in the Astra Zeneca, Janssen and the Russian Sputnik covid vaccines.

The virus is just the delivery system (vector) to get some genetic code into cells.

The Oxford-AstraZeneca COVID-19 vaccine uses a chimpanzee adenoviral vector. It delivers the gene that encodes the SARS-CoV-2 spike protein, to our cells.  Our cells then transcribe this gene into messenger RNA, or mRNA, which in turn prompts our cellular machine to make the spike protein in the main body of the cell. The mRNA molecule behaves essentially like a recipe.  Then our cells present the spike protein on the cell surface, prompting our immune system to make antibodies and mount T cell responses.

Biontech and Moderna are pioneers of mRNA vaccines, which bypass one step in the above process. They do not require our cells to make the messenger RNA, or mRNA.  They have already made it for you.

 

Gene therapy for autism?

Single gene autisms are all potential candidates for gene therapy.

The problem is that most autism and all Down syndrome is polygenic, there can be hundreds of miss-expressed genes.

But the researchers in Italy show us that even polygenic autism and Down syndrome can benefit from therapy targeting a single gene.  You just have to select the right one.

The problem is the price. Covid vaccines are made in huge quantities and are cheap.

Customized gene therapy is ultra expensive, in part because each therapy has to be approved individually.

 

An NKCC1 Gene Therapy?

The Italians have already made the NKCC1 Gene Therapy.  The question is will it ever going be available to humans with Down Syndrome, Autism or even Parkinson’s disease?

Restoring neuronal chloride homeostasis with anti-NKCC1 gene therapy rescues cognitive deficits in a mouse model of Down syndrome

A common feature of diverse brain disorders, is the alteration of GABA-mediated inhibition due to aberrant intracellular chloride homeostasis induced by changes in the expression and/or function of chloride transporters. Notably, pharmacological inhibition of the chloride importer NKCC1 is able to rescue brain-related core deficits in animal models of these pathologies and some human clinical studies. Here, we show that reducing NKCC1 expression by RNA interference in the Ts65Dn mouse model of Down syndrome (DS) restores intracellular chloride concentration, efficacy of GABA-mediated inhibition and neuronal network dynamics in vitro and ex vivo. Importantly, AAV-mediated neuron-specific NKCC1 knockdown in vivo rescues cognitive deficits in diverse behavioral tasks in Ts65Dn animals. Our results highlight a mechanistic link between NKCC1 expression and behavioral abnormalities in DS mice, and establish a molecular target for new therapeutic approaches, including gene therapy, to treat brain disorders characterized by neuronal chloride imbalance.

 

This sets the stage for the development of a gene therapy approach to overcome the shortcomings of bumetanide treatment.

This highlights a causative role of NKCC1 upregulation in learning and memory deficits in adult Ts65Dn mice, thus also validating brain NKCC1 as a target for ameliorating cognitive disabilities in DS. Furthermore, our neuro-specific knockdown approach points to neurons as major players in the NKCC1- dependent cognitive impairment in DS mice. Nevertheless, we cannot exclude that other cell types which also express NKCC1 (e.g. glial cells) could still play a role in the overall cognitive impairment that characterizes DS.

Despite the very large and fast-increasing literature both on animal models and patients indicating positive outcomes upon bumetanide treatment, there is not yet a strong demonstrated direct link between NKCC1 inhibition, restoration of Cl- homeostasis and full GABAergic inhibitory signaling, and rescue of brain deficits.  Moreover, bumetanide has strong diuretic activity, triggering ionic imbalance, and potential ototoxicity 25,26.  This hampers its use for clinical applications in lifelong treatments4,27 and may strongly jeopardize treatment compliance along years of treatment.  Moreover, bumetanide was given systemically in most studies, and the suboptimal brain pharmacokinetic profile of the drug28 raises questions on its mechanism of action29.  Here, we demonstrate that adeno-associated virus (AAV)-mediated RNA interference (RNAi) to target (and reduce) neuronal NKCC1 expression rescues neuronal Cl- homeostasis, GABAergic transmission, and cognitive deficits in the Ts65Dn mouse model of Down syndrome. This sets the stage for the development of a gene therapy approach to overcome the shortcomings of bumetanide treatment.

 

“Thus, our results indicate the efficacy of long-term AAV9-mediated neuro-specific NKCC1 knockdown in rescuing cognitive deficits in Ts65Dn mice.”

 

“Besides establishing a causal link between NKCC1 upregulation and cognitive impairment in DS, our data also provide a proof-of-concept for a neuro-specific RNAi gene therapy approach to restore hippocampus-dependent cognitive behaviors in adult animals specifically in the brain, and without affecting peripheral organs (e.g., the kidney). This is particularly relevant in the context of the current clinical trials repurposing the strong diuretic bumetanide to treat brain disorders with impaired chloride homeostasis3.  Importantly, we achieved a comparable degree of long-term cognitive rescue with two different amiR sequences against NKCC1, underlining the specificity of our approach.”

  

Gone Fishing




If a trip to Italy for gene therapy is not realistic, this takes us back to AJ’s idea, which is to use Betaine.  The correct version is TMG or glycine betaine, and confusingly not Betaine HCl.

Fish love the taste of betaine.

Betaine was first isolated from sugar beet.

I recall from my time at the sugar factory, when I was 18, that once you have sliced up the sugar beet and extracted as much sugar as possible you are left with the pulp.  This pulp is dried, molasses is added back and then it is made into pellets.  The pellets are fed to cattle and horses.  They taste pretty bad in my opinion.

To humans it tastes bad because of the beet molasses by-product.

The molasses by-product from sugar cane tastes great to humans.  That is why they make rum in the Caribbean, and not in England or Canada.

Brown sugar from a sugar beet factory is made by adding sugar cane molasses to white sugar from beet.  It is a cheat really.

Cows love sugar beet by-products.

It turns out that fish love betaine HCl.

Betaine HCl is an excellent natural attractor that stimulates a strong, prolonged feeding response from carp and many other coarse fish.

Betaine HCl is now used to induce feeding in the fish farming industry

As our reader Tyler has highlighted, Betaine HCl, that fish like and is available is a cheap supplement is not the same as the Betaine used in the medical case study. Confusingly, the original Betaine (TMG, or called glycine betaine) gave way to a class of compounds all called betaines. One of these betaines is betaine HCL.

In most cases, in the medical literature when they refer to Betaine, they mean glycine betaine, also known as TMG.

Betaine HCl is used to increase acidity in your stomach. The effect of betaine compounds other than glycine betaine/TMG on NKCC1 is unknown.


Glycine Betaine (TMG) and NKCC1

It seems that betaine reduces your level of NKCC1 RNA. 

In your DNA are the instructions to make the NKCC1 transporter. To go from these instructions to actually making the transporters you need RNA.

In some autism there are too many NKCC1 transporters, so put simply there was too much NKCC1 RNA. So, if you can find a substance that reduces NKCC1 RNA, you might well solve the problem.

The caveat is that the substance must not also increase KCC2 RNA.  This appears to be what taurine does.

Here, finally, is AJ’s paper:


Treatment experience in two adults with creatine transporter deficiency

Background

Creatine transporter deficiency (CTD) is an X-linked form of intellectual disability (ID) caused by SCL6A8 mutations. Limited information exists on the adult course of CTD, and there are no treatment studies in adults.

Methods

We report two half-brothers with CTD, 36 and 31 years at intervention start. Their clinical phenotypes were consistent with CTD, and intervention was indicated because of progressive disease course, with increased difficulties speaking, walking and eating, resulting in fatigue, and malnutrition. We therefore performed treatment trials with arginine, glycine and a proprietary product containing creatine and betaine, and then a trial supplementing with betaine alone. Results In the older patient, glycine and arginine were accompanied by adverse effects, while betaine containing proprietary product gave improved balance, speech and feeding. When supplementation stopped, his condition deteriorated, and improved again after starting betaine supplement. Betaine supplementation was also beneficial in the younger patient, reducing his exhaustion, feeding difficulties and weight loss, making him able to resume his protected work.

Discussion & conclusion

We report for the first time that betaine supplement was well tolerated and efficient in adults with CTD, while arginine and/or glycine were accompanied by side effects. Thus, betaine is potentially a new useful treatment for CTD patients. We discuss possible underlying treatment mechanisms. Betaine has been reported to have antagonistic effect on NKCC1 channels, a mechanism shared with bumetanide, a medication with promising results in both in autism and epilepsy. Further studies of betaine's effects in well-designed studies are warranted.

 

The mechanism of betaine’s assumed favorable effect is unknown. We do not know whether betaine influences the cell creatine content in itself or its effects are more aspesific. However, we would like to present some hypotheses. First, betaine may have effect in CTD by modulating GABA-transmission. Betaine has been reported to have an antagonistic effect on NKCC1 channels, which also influences GABAergic neurotransmission. Inhibiting NKCC1 is a mechanism shared with bumetanide, a well-known diuretic medication that in recent years has been found to influence GABAergic transmission, and thereby it has been found promising in treatment of several brain conditions, including autism, and epilepsy. NKCC1 inhibition by bumetanide has also been tried with success in other rare neurodevelopmental disorders fragile X syndrome and tuberous sclerosis. Second, betaine’s properties as an osmolyte may be of importance, as betaine has similarities with creatine in being an osmolyte. Osmotic properties are thought to be one of the central mechanism behind bumetanide’s efficacy in treating brain disorders. Thus, it could be speculated that the lack of intracellular creatine in CTD may result in inefficient osmolyte regulation, and that betaine supplementation replaces the lacking creatine and thereby improves the neuronal adaption to salinity changes, edema or cellular dehydration. Betaine has osmolyte properties that even makes it act as a “chemical chaperone” increasing the stability of cell and membrane proteins. Fourth, it is possible that betaine has some effect through modifying methylation. Methylation of GAA by GAMT to form creatine is a rate-limiting step in the creatine synthesis by neurons. Betaine could stimulate this by donating methyl groups to SAMe, which donates a methyl group to GAA to form creatine. This might reduce the burden when body demands more methyl groups for creatine synthesis. Similar mechanisms may be responsible for a beneficial effect of both betaine and s-adenosyl methionine (SAMe). However, as creatine and GAA share the same transporter, one would not expect GAA to enter the GAMTexpressing cells in patients suffering from CTD. Still, it cannot be excluded that there is some rest function in the creatine transporter, and that increased endogenous synthesis improves the condition slightly. Furthermore, it is possible that CTD increases the need for methylation agents in general, as creatine supplementation has been found to reduce the need for other methylation agents [34]. Thus, it is likely that betaine may have a positive effect in CTD by improving methylation capacity for other reactions than those directly involved in creatine production. Betaine’s effect on muscle may be also of importance, as animal studies have shown that muscles growth improves with betaine [35], which potentially could have had a positive impact on our patients fatigue and weight loss. To summarize, betaine has several properties that make it likely that it will have a beneficial effect in CTD, especially the properties as an osmolyte, a down regulator of the NKCC1 channel and an influencer of GABAergic transmission. These properties are similar to the properties of bumetanide, a promising new medication for treatment of autism and epilepsy, which are common symptoms of CTD. Further research is needed, however, to elucidate the role of betaine in CTD.

If you read the detail of the old paper that is referred to in the above paper, you see that betaine is not blocking the NKCC1 channels as suggested, but it seems to be reducing the number of them.  The net effect may be the same, but the process is very different.

 

Expression and regulation of the Na+/K+/2Cl− cotransporter NKCC1 in rat liver and human HuH-7 hepatoma cells

The expression of sodium potassium chloride cotransporter 1 (NKCC1) was studied in different liver cell types. NKCC1 was found in rat liver parenchymal and sinusoidal endothelial cells and in human HuH-7 hepatoma cells. NKCC1 expression in rat hepatic stellate cells increased during culture-induced transformation in the myofibroblast-like phenotype. NKCC1 inhibition by bumetanide increased α1-smooth muscle actin expression in 2-day-cultured hepatic stellate cells but was without effect on basal and platelet-derived-growth-factor-induced proliferation of the 14-day-old cells. In perfused rat liver the NKCC1 made a major contribution to volume-regulatory K+ uptake induced by hyperosmolarity. Long-term hyperosmotic treatment of HuH-7 cells by elevation of extracellular NaCl or raffinose concentration but not hyperosmotic urea or mannitol profoundly induced NKCC1 mRNA and protein expression. This was antagonized by the compatible organic osmolytes betaine or taurine. The data suggest a role of NKCC1 in stellate cell transformation, hepatic volume regulation, and long-term adaption to dehydrating conditions.

 

Aha!  Glycine Betaine and Taurine – not so fast 

You have to check the effect on both NKCC1 and KCC2.  One lets chloride into neurons and the lets it out.  You want to block NKCC1 and not KCC2, otherwise you undo all the good you have done.

Both glycine betaine (TMG) and taurine are already used as autism supplements at low doses.  The paper below suggest that Taurine is not a good idea for people with high levels of chloride within neurons.

 

Taurine inhibits K+-Cl- cotransporter KCC2 to regulate embryonic Cl- homeostasis via with-no-lysine (WNK) protein kinase signaling pathway

GABA inhibits mature neurons and conversely excites immature neurons due to lower K(+)-Cl(-) cotransporter 2 (KCC2) expression. We observed that ectopically expressed KCC2 in embryonic cerebral cortices was not active; however, KCC2 functioned in newborns. In vitro studies revealed that taurine increased KCC2 inactivation in a phosphorylation-dependent manner. When Thr-906 and Thr-1007 residues in KCC2 were substituted with Ala (KCC2T906A/T1007A), KCC2 activity was facilitated, and the inhibitory effect of taurine was not observed. Exogenous taurine activated the with-no-lysine protein kinase 1 (WNK1) and downstream STE20/SPS1-related proline/alanine-rich kinase (SPAK)/oxidative stress response 1 (OSR1), and overexpression of active WNK1 resulted in KCC2 inhibition in the absence of taurine. Phosphorylation of SPAK was consistently higher in embryonic brains compared with that of neonatal brains and down-regulated by a taurine transporter inhibitor in vivo. Furthermore, cerebral radial migration was perturbed by a taurine-insensitive form of KCC2, KCC2T906A/T1007A, which may be regulated by WNK-SPAK/OSR1 signaling. Thus, taurine and WNK-SPAK/OSR1 signaling may contribute to embryonic neuronal Cl(-) homeostasis, which is required for normal brain development.

 

So, it is likely only Glycine Betaine (TMG) may be of potential benefit, in the case of lowering chloride.

 

Glycine Betaine in the broader research

 

Betaine in Inflammation: Mechanistic Aspects and Applications

Betaine is known as trimethylglycine and is widely distributed in animals, plants, and microorganisms. Betaine is known to function physiologically as an important osmoprotectant and methyl group donor. Accumulating evidence has shown that betaine has anti-inflammatory functions in numerous diseases. Mechanistically, betaine ameliorates sulfur amino acid metabolism against oxidative stress, inhibits nuclear factor-κB activity and NLRP3 inflammasome activation, regulates energy metabolism, and mitigates endoplasmic reticulum stress and apoptosis. Consequently, betaine has beneficial actions in several human diseases, such as obesity, diabetes, cancer, and Alzheimer’s disease.

 

Betaine is a stable and nontoxic natural substance. Because it looks like a glycine with three extra methyl groups, betaine is also called trimethylglycine . In addition, betaine has a zwitterionic quaternary ammonium form [(CH3)3N+ CH2COO−] (Figure 1). In the nineteenth century, betaine was first identified in the plant Beta vulgaris. It was then found at high concentrations in several other organisms, including wheat bran, wheat germ, spinach, beets, microorganisms, and aquatic invertebrates. Dietary betaine intake plays a decisive role in the betaine content of the body. Betaine is safe at a daily intake of 9–15 g for human and distributes primarily to the kidneys, liver, and brain. The accurate amount of betaine intake generally relies on its various sources and cooking methods. Besides dietary intake, betaine can be synthesized from choline in the body. Studies report that high concentrations of betaine in human and animal neonates indicate the effectiveness of this synthetic mechanism.

  

Boosting amino acid derivative may be a treatment for schizophrenia

Many psychiatric drugs act on the receptors or transporters of certain neurotransmitters in the brain. However, there is a great need for alternatives, and research is looking at other targets along the brain's metabolic pathways. Lack of glycine betaine contributes to brain pathology in schizophrenia, and new research shows that betaine supplementation can counteract psychiatric symptoms in mice.

 

 

Supplement treats schizophrenia in mice, restores healthy “dance” and structure of neurons Repurposed drug works by building cells’ skeleton and transportation network


 

 

Conclusion

Early on in the Covid saga, I saw interviews with both the Moderna researchers and the Oxford (AstraZeneca) researchers. Both claimed that they designed their vaccines over a weekend.  This was made possible by the Chinese releasing the DNA code of the virus.

When you think about gene therapy for autism and Down syndrome, the same likely applies; much could be achieved over a weekend.

The expensive and time-consuming part is the testing and approval process.

In the Covid pandemic the approval process was modified to allow for emergency use.  Perhaps this should also be the case for all gene therapies?

What use is a $2 million therapy for autism or Down syndrome?

In theory, if you gave your gene therapy prior to birth or shortly thereafter, it might be fully curative.  Realistically, by the time you get the therapy it is just going to be beneficial and you will still need other ongoing therapies.

Note that gene therapy normally applies to just one gene.  In Down syndrome people have a third copy of all, or just part, of Chromosome 21.  This results directly in the miss-expression of hundreds of genes from that chromosome.

The gene that encodes NKCC1 is on Chromosome 5, which has nothing directly to do with Down syndrome.

The NKCC1 transporter is over-expressed in Down syndrome as a down stream consequence of the disorder. It is caused by the “faulty GABA switch”, referred to in earlier posts.

The Italian gene therapy to lower chloride in neurons and so raise cognition, has numerous applications, in people currently of all ages, so there is a big potential market.

Why not gene therapy for all single gene autisms?  It could be a highly productive use of the researcher’s weekends, for a year or two.

The issue is who would pay for the $20 to $30 million approval process, for each gene?

Maybe some of the billions in profit from clever Covid vaccines could be used for pro bono gene therapy?  Highly unlikely.

Biontech, who are the brains behind the Pfizer vaccine, do have plans to develop gene therapy for other medical conditions.  I think these will be ultra expensive,

That brings me back to Glycine Betaine (TMG), is 10g a day of this supplement really going to reduce the expression of NKCC1 transporters in neurons and so lower chloride within neurons?  It seems to work in creatine transporter deficiency, is all we can say.  

Glycine betaine, at much lower doses, has been used by DAN and now MAPS doctors for decades. They use it as a “methyl-donor”.  There is a combination of real science and hocus-pocus surrounding DNA methylation. 

 DNA Methylation and Susceptibility to Autism Spectrum Disorder


 

 



 

48 comments:

  1. Hi Peter,

    Thanks for the great post!

    I actually started 125mg X 2 of Betaine soon after I posted about my finding, and no issues at all and my wife and I think we are seeing some mild improvement in speech complexity. It's likely too soon to tell, and the dose I'm using is low (which I will likely titrate up to 250mg X 2 soon).

    Also, I wasn't aware of the efforts to address NKCC1 via gene therapy. That would be a very interesting option for those who would otherwise be Bumetanide responders.

    Have a great day Peter!

    AJ

    ReplyDelete
  2. Only concern with trialing betaine is do we have to look out for this

    Cerebral edema associated with betaine treatment in classical homocystinuria.

    Not sure if this just reflects people certain disease state but the use of high dose betaine links to brain Edema.

    Otherwise I'm definitely going to give this a go. Are you considering trialing betaine with Monty as an add on

    ReplyDelete
    Replies
    1. The issue seems to relate to increased Methionine levels. This was discussed in the Norwegian case study. In their cases they monitored Methionine and the level did not increase substantially.

      In people with Cystathionine beta-synthase (CBS) deficiency, the most common form of homocystinuria, it appears that an unusual reaction is possible and methionine levels increase substantially and this causes the edema.

      Ideally you would follow the example of the Norwegians and measure Methionine.

      The research paper above states that up to 15 g a day is safe in adults. The Norwegians are using 10g.

      I may well try betaine as an add on therapy.

      Delete
    2. Peter... In short, could Betaine be a substitute for Bumetanide? If 10 to 15g is safe for adults, what would be safe for children up to 10 years old or 30 kg? 5g?

      Delete
    3. I would follow AJ's example and start with a low dose and gradually increase it. The effect will not be identical to bumetanide.

      Methionine supplemention was reported by Johns Hopkins to be beneficial, for no known reason and Betaine may also produce this benefit.

      So you may see multiple effects, including the "bumetanide effect" to some degree, we do not know for sure. But it does look interesting.

      Delete
  3. One big advantage of beta one is that it’s easily available—no need to find a open-minded doctor. On the downside, the doses are much larger. Given the taste issues, getting my son to take 5 or 10 mg a day seems challenging!

    ReplyDelete
  4. Hi Peter

    Talking about future gene therapy Spectrum News released this article about a potential future therapeutic target CUL3 mutation that leads to neuronal migration problems.

    Since problems with Neuronal Migration are what most scientists believe lead to most autism in the first place, if they can do this in early development even maybe shortly after birth, it could potentially prevent autism.

    The main downside I can see is Neuronal Migration is possibly a done deal after a certain critical age so what can be done after to address this specifically is unclear.

    https://www.spectrumnews.org/news/autism-linked-mutation-may-disrupt-brain-cell-migration/

    ReplyDelete
  5. I also found this paper interesting, given the purported links between gluten sensitivities and autism: https://www.frontiersin.org/articles/10.3389/fimmu.2019.02484/full.

    ReplyDelete
  6. I also found this paper interesting, given the purported links between gluten sensitivities and autism: https://www.frontiersin.org/articles/10.3389/fimmu.2019.02484/full.

    ReplyDelete
  7. Peter please correct me if I am wrong, but Betaine HCL is not the same as Trimethylglycine. This is a mistake I made many years ago when I used to use TMG with my son (no noticeable improvements, but I didn't use a lot of it). I first bought Betaine HCL after reading about methylation issues in autism in a now very old paper on the subject and did not realize until later I bought the wrong supplement (I was looking for Trimethylglycine). Is this all confusing? Of course it is so I could be wrong, but you may want to check out what I am saying and correct things in case I am right about this.

    Betaine HCL is used for digestive purposes and the other Betaine known as TMG is what you are looking for in terms of the supplement to take and they have nothing to do with each other besides the name.

    ReplyDelete
    Replies
    1. Tyler, thanks for this.

      The original betaine, the one from sugar beet is TMG. It is also now called glycine betaine.

      Accoding to Mount Sinai “betaine supplements are manufactured
      as a byproduct of sugar beet processing. They are available in powder, tablet, and capsule forms.”

      The term betaine has been broadened to refer to a class of compounds and hence the confusion.

      Which betaine was used in the case studies?

      The constituents were trimethylglycine (betaine) 46%,
      creatinine 36%, creatine 11%, GAA 4%, taurine 4% and no cyclocreatine. Betaine (Fig. 1) is a methyl group donor, commonly used for treating homocystinuria. Betaine donates a methyl group to homocysteine, forming dimethylglycine and methionine.

      “Trimethylglycine (TMG) is a popular molecule in the betaine family – consisting of the amino acid glycine with three methyl groups attached. TMGs main uses are as either a primary (reducing homocysteine into L-methionine for cardio-protection) or secondary (affecting folate and SAMEe metabolism) methyl donor. For clarity it’s important to remember that the terms “TMG” and “betaine” are used interchangeably

      While still in the same molecular family, betaine hydrochloride [HCl] is a very different molecule. It is a protonated form of TMG, meaning it will give up or transfer a proton and a chloride ion in an aqueous solution (liquid). This makes betaine HCl a very good dietary source of hydrochloric or gastric acid.”

      The conclusion is that you need to use TMG, glycine betaine to replicate the case study.

      Delete
  8. Hello! Is DMG a weaker version of TMG? What experience do you have with DMG? Thanks

    ReplyDelete
    Replies
    1. DMG and TMG are related but different. They are widely used by DAN and MAPS type doctors.

      This post is based on the observation that 10g a day of TMG was very beneficial in Creatine Transporter Deficiency. The clinicians suggest that TMGs effect on lowering chloride may be what helped. That would make this therapy equivalent to taking Bumetanide with the advantage of no diuresis (and no prescription).

      I have been using bumrtanide for 8 years. I have not tried TMG.

      Delete
  9. Hi Peter
    About TMG and its NKCC1 effects I heard quite lot and therefore I tried to give to my son about 1g daily since 3 months ago. Now I read this article and tried more - 4g a day (he is 30kg) and effects seems to be more adverse or neutral than better. He is non typical ASD with lowered cognition and more aspects mainly social interraction. But he is verbal he speeks a lot. Dou you have any clue when betaine could have adverse effects instead of positive?
    I'm aaking because we still don't know the ground of his disability and still looking by observing effects of diverse drugs and supplements.

    ReplyDelete
    Replies
    1. TMG and DMG are widely used by the DAN/MAPS doctors, you can seen on google that side effects are quite common, often the problem is hyperactivity.

      If the DAN/MAPS doctors are correct TMG/DMG should have numerous effects relating to methylation and so gene expression.

      TMG will reduce homocysteine and may increase methionine, both of these may have consequences, in some people.

      If you just want to block NKCC1, the simple option is Bumetanide, which we know works.

      Delete
  10. I was rather disappointed with TMG & DMG saw no real difference at all. Having said that it was only for a few weeks each then combined the two for a few more weeks. If my son was a potential responder of TMG how long would I need to wait to see some improvement? & would taking DMG & TMG combined reportedly make any difference?

    ReplyDelete
    Replies
    1. Ross, the DAN doctors said that giving 1g a day of DMG or TMG improved speech. The actual evidence is not very extensive.

      The clinicians treating Creatine Transporter Deficiency, a very specific genetic disorder, gave 10g a day in adults.

      Blocking NKCC1 slowly reduces elevated chloride in neurons; for Bumetanide this usually takes 2 weeks, but sometimes even longer. If the NKCC1 blocker is too weak or too low dose, it will never reduce elevated chloride sufficiently.

      The only actual evidence of blocking NKCC1 refers to TMG. We do not know the effect, if any, of Betaine HCl, or DMG.

      You might expect that if a child took half the adult dose of TMG, the effect would be similar. If the clinicians are correct and NKCC1 is the mode of action in this case, then I think you would need to make a trial for a month.

      Delete
  11. Peter,
    Do you think breaking the 10g of TMG into 2 5g doses/day would make a difference in effectiveness of this one-month trial therapy?
    Nancy

    ReplyDelete
    Replies
    1. Nancy, I just looked it up and TMG is very slowly eliminated from the body and the more you use it the longer it stays in the body. That means that you can split the dose without it making a difference.

      Delete
  12. Peter,
    I tried tmg alone withy son long ago for 3months (low doses) and didn't see any changes

    But he is responding to bumetenide seeing some gains in cognition and writing skills

    Do you think adding TMG now would increase bum. Benefits ?

    The max dose per pill tmg I could find is 675mg so that's lots of pills for 10g?

    Is there any other sources to get tmg in larger dose ?

    Any other add-ons for bumetenide?

    ReplyDelete
  13. Riza, in sound countries they sell TMG as a bulk powder.

    The best add-on for bumetanide seems to be a higher dose of bumetanide. Any drug/substance that is an OAT3 inhibitor will slow the excretion of bumetanide and increase its level in the body. You can search for OAT3 in my blog.

    Try giving bumetanide twice a day, if you are currently giving it once a day.

    ReplyDelete
  14. Could it be assumed that if a child is not a responder to TMG they are unlikely to be a responder to Bumetanide since they both target the same things?

    ReplyDelete
    Replies
    1. Ross, until we have more reports from people who have tried both high dose TMG and Bumetanide, it is too early to conclude anything. All we know is that low doses of TMG seem to provide no benefit in people who are bumetanide responders. The clinicians suggested that the benefit of high dose TMG was due to its bumetanide-like effect; this needs to be confirmed.

      Delete
  15. Another reason why the FDA may be reclassifying NAC as a drug after the last 62 years of NAC being an over the counter supplement:

    https://www.sciencedaily.com/releases/2021/06/210630125421.htm

    Looks like Big Pharma wants to come out with derivatives of NAC which they can sell for big bucks and then when you go to your doctor and say you want NAC, your doctor says "well there is a better version and it only costs 10x as much". Of course the other possible reason for NAC being made prescription only is because it is critical in countering some of the most damaging and fatal side-effects of the COVID-19 vaccines and I can leave everyone else to speculate why that would be happening. I will say, in India the WHO scientist who said nobody should be using Ivermectin and banned it temporarily while they had a mass vaccine rollout and then a ton of COVID-19 deaths is facing a life sentence or the death penalty for stopping the use of Ivermectin. Once India started distributing Ivermectin widely again, COVID-19 deaths dropped to negligible levels. So far the CDC has pretty much tried to ban every cheap and effective drug in treating COVID-19 now and there are just too many coincidences at this point not to assume malice. The most risky period for vaccinated people will be 6 months to 2 years after vaccination when ADE or antibody dependent enhancement could very well kill a vaccinated individual if exposed to any coronavirus, so having NAC widely available could be a lifesaver for a lot of people who got conned into taking the vaccine and need to get by the next couple of years in avoiding a potential cytokine storm. I just cannot believe half the country or more still cannot figure out what is going on by now when the evidence seems pretty clear at this point in that the government seems to be deliberately taking actions to harm and kill as many people as possible.

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  16. Roger, there is a lot of confusion about betaine compounds.
    The betaine used to lower homocysteine is TMG. When TMG breaks down in your body it produces DMG, lowers homocysteine and raises methionine. Clealy TMG was good for you.

    The supplement people take to increase acidity in their stomach is Betaine HCl. The only part that does anything to do with acidity is the HCl part (hydrochloric acid HCl). What then happens to the betaine part? People wanting to raise stomach acidity could use many things, apple cider vinegar being a common choice.

    DAN doctors think some people are “under-methylated”. TMG and DMG are so called “methyl donors” and that is the reason they use these substances.

    Researchers are trying to find a bumetanide equivalent without the diuresis and so TMG raised their interest.

    Parents are struggling to get hold of bumetanide, or any equivalent, because of unresponsive doctors. When finally Bumetanide gets approved for autism, this problem will go away.

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  17. Here is an interesting paper that caught my eye which shows how a GABA receptor on microglia acts as a receiver from GABA released from inhibitory synapses on neurons which then instructs the microglial cell to then go prune the inhibitory synapse which expressed GABA.

    Press Release:

    https://www.sciencedaily.com/releases/2021/07/210706115334.htm

    Paper:

    https://www.cell.com/cell/fulltext/S0092-8674(21)00753-4?_returnURL=https%3A%2F%2Flinkinghub.elsevier.com%2Fretrieve%2Fpii%2FS0092867421007534%3Fshowall%3Dtrue

    Also quite interesting is that when they removed the GABA receptor from the microglia of mice, there was an overabundance of inhibitory synapses and the behavior of the mice were similar to what happens if you dose them with a strong GABAergic substance. Also interesting is that once the mice reached adulthood, the global brain inhibition turned to excitation which the researchers speculated as a compensating factor. With respect to autism one can wonder if the opposite situation could be happening as often autistic children are extremely hyper when they are young and as then as they mature they often become slower, more lethargic, and more hypotonic.

    It would be quite counter-intuitive if excess GABA receptors on microglia or else the microglia being activated some other way caused the microglia to excessively prune inhibitory synnapses, thereby leading to the excitation/inhibition imbalance throughout the brain. I have no evidence of this for autism, but it is an interesting hypothesis that came to mind while reading this paper.

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  18. Dear Peter, sorry to post on a completely different topic but I’m in great need of your advise ( and that of other readers). We are from South Africa and have a 5 year old autistic son. My husband has come across ‘MERT’ therapy and believes it’s a game changer- and so we are looking to come to the US for this. Do you have any experiences with this? It’s quite pricey we would need about US20,000

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    Replies
    1. Tinotenda, MeRT (Magnetic e-Resonance Therapy) is a form of repetitive transcranial magnetic stimulation (rTMS).

      One of the best known rTMS researchers is the very kind-hearted Professor Manuel Casanova.

      He has his own autism blog.

      https://corticalchauvinism.com

      I suggest you write to Manuel and ask him for advice on MeRT. He might tell you to start with something less expensive and closer to home.

      I have no experience of rTMS or MeRT. I would be very cautious about paying $20,000 for any autism therapy, unless you are very wealthy.

      Delete
  19. Hi,
    We are entering week 4 of betaine anhydrous (TMG) and things have been increasingly positive. I have not seen such flexibility and calmness since vagus nerve stimulation and back when we did Pantogam. (both did not last). BCAAs and nicotinaimde have been a staple improving mood about 80%. The TMG seemed to add the additional 20%.

    Then yesterday irritation and then rage began to creep back in. We are in that mid-summer heat and humidity that always takes its toll mood wise though L histadine has helped a lot during these periods.
    Is it possible that after 3 weeks of positives that 10 mg daily of TMG is having a negative effect or is it likely something else responsible?
    I'd like to continue giving it as it makes no sense to me that adverse effects would only begin now.
    Thanks for your advisement.
    Nancy

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    Replies
    1. Nancy, low doses of TMG are a common DAN/MAPS autism therapy. I found this comment:

      "Care must be taken when supplementing betaine (TMG), as approximately 20% of ASD children experience increased hyperactivity. If the side effects with TMG are intolerable, DMG (dimethylglycine) can be used as an alternative."

      The effect you are seeing from TMG are unlikely to be the bumetanide-like effect discussed in this post, because that would require a very much higher dose.

      Summertime makes some people with autism irritable and aggressive.

      The negative effects you are now seeing could be due to TMG or the summer.

      You could pause TMG for a week and see what is the effect.

      You could replace TMG with DMG (dimethylglycine) and see how that works.

      You could also consider repeating your trial after the summer.

      Delete
  20. I erroneously wrote 10 mg. I am giving 10 grams, 5g in the AM and 5g in the evening. I also neglected to add that I had switched brands of powder for about 3 days. I wonder if there are differences in quality? I have resumed what we started with.
    Last night appeared to be somewhat better.

    ReplyDelete
    Replies
    1. OK Nancy, so you may well be seeing the bumetanide-like effects.

      It is hard to know will supplements, sometimes they are not reliable regarding quality.

      The only way to know if the TMG is causing negative effects is to make a pause.

      In my case summer is always the most difficult time of year, so I do not trial something new. I suspect your son is similar.

      Delete
  21. Hi, Peter I recently came across an article on depression and anxiety, I wanted you to take a look at these two drugs (supplements) TIANEPTINE (stablon) and NAG (N-acetate-glucosamine), do you think it might be useful? In our case, thank you and excuse us if we stress you, your mission on earth, I think it is made to help us with a lot of advice and guidance, thank you for existing.

    ReplyDelete
    Replies
    1. Dragos, Tianeptine has very many biological effects. It is effective for ADHD but also pain in fibromyalgia. It is worth trying, but because it has so many effects you need to see if your son gets any side effects.

      Our reader Tyler gives his son NAG, because the research shows it can improve myelination. He says there is no immediate effect, so it is hard to know if there is a benefit.

      I would think Tianeptine will have an effect straight away. You just need to decide if the overall effect is good for him.

      Good luck

      Delete
  22. Hi Peter,

    I’m going to be trying NAC for my 7 year old (he weighs 60 pounds) what dose do you suggest we start at? And how many times per day? I also purchased TMG, Carnitine and sulforaphane but I’m going to try NAC first. I also just purchased EPA\DHA supplement to hopefully help with his hyperactivity. If we can calm him down he would be close to neurotypical, his hyperactivity really holds him back. Thank you!!

    ReplyDelete
    Replies
    1. In the clinical trials they were giving the children 3 doses of 900 mg NAC per day spaced out evenly. As with all medications its a good idea to start lower and work your way up. So, you could start with 2 doses per day for the first week and add a 3rd dose on week 2.

      One aspect of NAC which I don't see brought up is the impact that effervescent vs pill form can have. I use the Pharmanac brand effervescent tablets and it has had far more of an impact than the pill form. I originally tried this brand since it was the same brand used in some of the clinical trials.

      NAC usually takes somewhere between 3 days - 2 weeks to show case an affect, and it usually takes about 2-3 weeks for the impact to reach a plateau.

      Delete
  23. Dear everyone, I read (on fb) the information that TMG can only be drunk for a month! Is that right? Thanks

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  24. Hi Peter, in switching brands of TMG (betaine anhydrous) due to running out and having to use a different brand temporarily, I realized what I thought was 1 tsp for 1500 mg was actually 1/2 tsp. Ling story short, I have been giving 9 mg 2x a day, total of 18 g.
    So when I lowered it to 10 daily using a different brand, I am seeing mood issues come back. Mood and flexibility and overall "presence" have been amazing on the bigger dose with now the correct 10 g noticeable the other way.
    Can I use such a high dose?

    ReplyDelete
  25. not 9 mg but 9 grams 2x a day. I am not used to writing grams!

    ReplyDelete
    Replies
    1. It is not surprising that the effect is dose dependent.

      The best way to check for safety is by looking at kidney and liver function. There are established tests for this.

      I do not think anyone has looked at such high doses of TMG. They may well be entirely safe. I would run these very common blood tests and check; then you will know.

      Clearly you also need to keep an eye out for any visible negative side effects, as with any other intervention.

      The other type of betaine, betaine HCl, has hydrochloric added to it and this will increase stomach acidity. You would expect that type to have side effects at very large doses.

      Delete
    2. Hi Peter, from your research and experience, is there any "cleansing cycles/therapies" you think is recommended for kids with autism due to number of supplements they take? My daughter's autism consultant recommended desmodium recently (although as i search around milk thistle seems to be better options). Have you considered any of these for Monty in the past?

      P.S. i remember even from my bodybuilding experience, when i was off spring i used to go through kidney/liver cleansing for a month of two.

      Delete
    3. Timur, if kidney and liver function are normal, no action should be needed. Not all drugs/supplements are well tolerated by everyone. If you encounter side effects, stop the therapy.

      Some children take a very large number of supplements (20+) and it is not a surprise if this causes problems.

      Your body has built-in, highly effective, "detoxifying" machinery. As long as you do not damage it, or overload it, you should have nothing to worry about.

      Delete
  26. My 4 year old autistic son, and myself with high functioning autism, both have snp's in BHMT gene so we've been taking Betaine for about a month, and I've seen very noticeable improvement in speech complexity in my son, and I have seen a reduction in auditory processing issues in myself. My son went from 1-2 word phrases only, to 4-6 word phrases and several times has spoken multiple sentences.

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    Replies
    1. That is great news. Thanks for sharing.

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    2. What dose are the two of you taking?

      Delete
    3. We've been taking 3 grams daily, as a powder, I'm tempted to increase it. We're on many different supplements and vitamins to target different possible aspects of Autism, but Betaine and folinic acid have been the two I definitely noticed a change with.

      I greatly appreciate the work you are doing! I think one of the issues hindering progress in the advancement of Autism treatment is that there aren't enough subsets in treatment testing to target specific issues, bio or genetic markers. We may never move forward if we're just looking for one thing to cure all or the majority of Autism cases.

      Delete
    4. It may be worth mentioning that we also have some snp's that seem related to calcium and potassium channel signaling.

      Delete
    5. Both potassium and calcium channels are heavily implicated in autism.

      For calcium channelopathies it likely will take a prescription drug.

      For potassium, some people get a benefit from a potassium supplement and some benefit from the potassium channel effects of the OTC painkiller Ponstan.

      Delete

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