Thursday 11 January 2024

Mutations in CACNA2D1 plus KDM6B -- Gabapentin and Calcium Folinate? Perhaps PQQ? Perhaps BHB?


A little research can sometimes be eye opening

I was recently sent genetic results from several parents and surprisingly some have multiple potentially highly causal genes. Some are mutations that are extremely rare and one was unique.

Today I am looking at one case with two genes highlighted in whole exome sequencing (WES), one is a calcium ion channel and the other is a gene extremely close to the one causing Kabuki syndrome.  Interestingly, two possible interventions did very quickly appear.

The report states:


Variants of uncertain significance (VUS) identified

Based on current evidence, the clinical relevance of the detected variants remains unclear.

Kabuki syndrome is caused by mutations in KMT2D or KDM6A.

KDM6A and today’s gene KDM6B both target trimethylation on lysine 27 of histone H3 (H3K27me3), a mark associated with gene silencing. By removing this mark, they activate gene expression. So, mutations in either gene will cause a cascade of effects on numerous other genes.

The old post below suggested the use of HDAC inhibitors to correct the mis expressed genes. In particular, BHB from the ketogenic diet was discussed.

Notably, histone deacetylase inhibition rescued structural and functional brain deficits in a mouse model of Kabuki syndrome.


Ketones and Autism Part 5 - BHB, Histone Acetylation Modification, BDNF Expression, PKA, PKB/Akt, Microglial Ramification, Depression and Kabuki Syndrome


The calcium channel involved today is not one we have previously looked at, but it is the target of the very well-known drug Gabapentin. This drug is used to treat epilepsy and neuropathic pain. The child does have abnormal EEG and seizures, plus autism, ADHD and absent speech.

Mutations of the KDM6B causing autism were first described only in 2019. In 2022 mutations in this gene were found in several patients with cerebral folate deficiency (CFD), one of the authors is our old friend Dr Ramaekers.

We know a lot about CFD, thanks to our reader Roger, Dr Frye, Dr Ramaekers, and now Agnieszka and Stephen. Over in the US one of the founders of an autism organisation told me her son was diagnosed in adulthood with CFD, when he finally had a spinal tap.

Interestingly, Agnieszka has pointed out a novel way to potentially increase folate in the brain using an OTC supplement called PQQ.


Protective effects of pyrroloquinoline quinone in brain folate deficiency


Folate deficiency resulted in increased expression of inflammatory and oxidative stress markers in vitro and in vivo, with increased cellular ROS levels observed in mixed glial cells as well as a reduction of mitochondrial DNA (mtDNA) content observed in FD mixed glial cells. PQQ treatment was able to reverse these changes, while increasing RFC expression through activation of the PGC-1α/NRF-1 signaling pathway.


These results demonstrate the effects of brain folate deficiency, which may contribute to the neurological deficits commonly seen in disorders of CFD. PQQ may represent a novel treatment strategy for disorders associated with CFD, as it can increase folate uptake, while in parallel reversing many abnormalities that arise with brain folate deficiency.


PQQ is a relatively common OTC supplement that looks helpful in older people and those with mitochondrial dysfunctions (most older people, plus many with autism).  It can also improve sleep.  The common 20mg dose seems to be based on what was used in a clinical trial in Japanese adults. Japanese drugs are dosed to reflect the size of Japanese people. American women on average weigh 40% more than Japanese women.

PQQ is present in mother’s milk, so it is not some scary artificial compound.

CFD looks like another nexus point where may different genetic variants produce a downstream meeting point.  This means numerous different underlying autisms will share a common beneficial therapy. It will not be a cure, but it should improve the outcome.

The only way to access I/V calcium folinate looks to be via confirmation of very low levels in spinal fluid, so a spinal tap would be necessary.  This is not easy, as Agnieszka has found out.  For some people oral calcium folinate is not sufficiently potent to reverse CFD.


Mutations of the KDM6B gene causing autism were first described only in 2019. In 2022 mutations in this gene were found in several patients with cerebral folate deficiency (CFD).


Genetic variants in the KDM6B gene are associated with neurodevelopmental delays and dysmorphic features

Lysine-specific demethylase 6B KDM6B demethylates trimethylated lysine-27 on histone H3. The methylation and demethylation of histone proteins affects gene expression during development. Pathogenic alterations in histone lysine methylation and demethylation genes have been associated with multiple neurodevelopmental disorders. We have identified a number of de novo alterations in the KDM6B gene via whole exome sequencing (WES) in a cohort of 12 unrelated patients with developmental delay, intellectual disability, dysmorphic facial features, and other clinical findings. Our findings will allow for further investigation in to the role of the KDM6B gene in human neurodevelopmental disorders.


Layman’s guide to the KDM6B gene


12% of people with CFD studied in the paper below had mutations in KDM6B. So clearly all people with a mutation in this gene should be tested for CFD vis a spinal tap.


KDM6B Variants May Contribute to the Pathophysiology of Human Cerebral Folate Deficiency

Cerebral folate deficiency syndrome (CFD) was defined as any neurological condition that was associated with low concentrations of 5-methyltetrahydrofolate in the cerebrospinal fluid. Previous clinical studies have suggested that mutations in the folate receptor alpha FOLR1 gene contribute to CFD. In this study, we identified six genetic variants in histone lysine demethylase 6B (KDM6B) in 48 CFD cases. We demonstrated that these KDM6B variants decreased FOLR1 protein expression by manipulating epigenetic markers regulating chromatin organization and gene expression. In addition, FOLR1 autoantibodies were identified in CFD patients’ serum. To the best of our knowledge, this is the first study to report that KDM6B may be a novel CFD candidate gene in humans.

The way to confirm CFD, with certainty, is via a spinal tap.  This can then open the door to intravenous therapy with calcium folinate.

There is a blood test which then would lead to oral calcium folinate therapy.  This is now very common in children with autism in the US. It improves speech.

The problem is that some people need the more potent intravenous therapy and without a spinal tap there is not enough proof to get the therapy.



The CACNA2D1 gene encodes voltage-dependent calcium channel subunit alpha-2/delta-1. 

Different types of mutation will have different effects and varying degrees of severity.

Some mutations in this gene are associated with a condition called “Developmental and Epileptic Encephalopathy 110”.

Developmental and epileptic encephalopathy-110 (DEE110) is an autosomal recessive disorder characterized by profound global developmental delay and hypotonia apparent in infancy followed by onset of seizures in the first months or years of life. Affected individuals achieve almost no developmental milestones and show impaired intellectual development, poor or absent speech, inability to walk or grasp objects, peripheral spasticity, and poor eye contact. Brain imaging shows hypoplastic corpus callosum and cortical atrophy.

CACNA2D1 is also a novel Brugada Syndrome susceptibility gene.

Brugada syndrome may be a major cause of sudden cardiac death in men under 40. People with Brugada syndrome on average die between the ages of 26 to 56 years, with an average age of 40 years. If treated appropriately, patients can have a normal lifespan.

A pediatric cardiologist should be consulted.

Fortunately the Alpha-2/delta proteins are believed to be the molecular target of the gabapentinoids gabapentin and pregabalin, which are used to treat epilepsy and neuropathic pain.

This means that an obvious path to investigate is whether the drug gabapentin has a positive effect. Mutations could produce either gain of function of loss of function.

Gabapentin binds to a the α2δ subunit. This binding does not directly block or open the channel, but it influences its overall activity.

The exact mechanism of action is still not fully understood, but it is believed that gabapentin:

·       Reduces the release of certain neurotransmitters involved in pain signaling, such as glutamate and substance P.

·       Alters the trafficking and function of the calcium channels themselves.

·       Therefore, gabapentin's action is more complex than simply "blocking" or "opening" channels. 

Gabapentin is not guaranteed to help in this case, but certainly might do.


The take home is really that if you invest thousands of dollars/euros/pounds in genetic testing, it is well worth your time spending some time on the internet looking up any flagged genes.

People expect too much from the geneticist writing the report.

Double check these things yourself.  Take your findings to an open-minded neurologist, who reads the research literature.

Be aware that the same mutation can be present in one or even both parents, with no noticeable negative effect, but be disease causing in their child. Genetics is often about the probability of something happening, rather the certainty. 

Look at partially-effective or sometimes-effective interventions in the research. For example, one reader is looking at mutations in NF1 plus ZMYND11. She might want to try Lovastatin.  NF1 causes an increase in RAS, which is a pro-growth signal, this leads to RASopathies which can cause intellectual disability (ID). Lovastatin reduces RAS and it was trialled to reduce ID in NF1 - the results were mixed. It probably matters at what age you start trying to reduce RAS.


  1. Peter, this is so interesting! It sounds like there are two ways to diagnose CFD, a blood test or spinal tap; and that the treatment is calcium folate, but the method of administration differs. Is that correct? And if that’s the case, why is it that only those diagnosed via spinal tap are prescribed the IV version? Curious because my son takes leucovorin/calcium folate orally but I’m wondering if he’d respond better to the IV version. He technically hasn’t been diagnosed with CFD, but he’s been taking it for 2 years at the recommendation of his developmental pediatrician.

    1. Most drugs are more effective when given directly into the bloodstream.

      It is far more convenient and much cheaper to use tablets and so that is what is done.

      Giving intravenous medication to many children with autism would be a big struggle.

      The only 100% reliable test for CFD is via a spinal tap. This is an invasive procedure and so generally avoided.

      The blood test is looking for antibodies to one of the 3 transport mechanisms. It is not measuring folate inside the central nervous system. It has a very high positive result.

    2. Hi Peter,

      Do you know if the IV version of leucovorin can be given subcutaneous? (Or maybe Dr. Ramaeker might know) I'm curious if an insulin pump with a basal rate of leucovorin would work. Sort of like how a diabetic gives themselves shots everyday with insulin. Maybe kids with FRAA can get leucovorin similar to insulin.

    3. Stephen, it is intramuscular or intravenous.

    4. Thank you Peter. Probably easier to create an inverse vaccine.

  2. Thank you for another informative post.
    Not related to this post , i came across info that calcium lactate will help with cough and fever. It helped me and my nearly 5 year old child. Do you know if it has any long term side efects? , thanks

  3. Hi Peter, Can you give me an example of how you utilize ? My son's report mentions the Gene, Transcript, mode of inheritance, the DNA variation, predicted (substitution) effects - but the variant does not show up. Am I navigating this incorrectly? For ex: - (CSMD1 Gene, Transcript is NM_033225.5 with DNA variation c.1349C>T and the predicted effect is p.Thr465lle in the report) - but filtering with 033225.5 or 1349C or Thr465lle on does not show results. I see results for 033225.6 but not 033225.5 - but how to interpret lack of results?

    1. The official reference sequence for the CSMD1 gene was represented by the NM_033225.5 accession number.

      On Genecards there is now only NM_033225.6, where there are 300 mutations listed. This is likely the new reference sequence. Maybe ask your lab?

      On a different database NM_033225.5 is listed and 37 variants are described.

      It would best to ask the geneticist for the RS number of the mutation. If there is no RS number then it must be a very rare mutation.

      An RS ID, or Reference SNP cluster ID, is a unique identifier assigned to a specific variation at a particular location in the human genome. These variations are known as single nucleotide polymorphisms (SNPs), which are the most common type of genetic variation among people.

      The RS ID is used to track and reference these variations across different databases and research studies. It provides a standardized way to identify and communicate specific genetic variations, making it easier for researchers to share and compare information.

      For single gene autisms you will find the name of the gene and a list of the mutations, by RS number, that are known to be causal.

      CSMD1 is an autism gene and a schizophrenia gene, but it is mainly seen as a cancer gene. It all depends on the mutation. This is a very large gene, so it has a large chance to contain a "spelling mistake". Many such mistakes are of no consequence.

      On SFARI gene your mutation is not listed.

      Genetics is very much a work in progress. Very few children with autism had their genes screened and shared the results.

      Have you been told his mutation in CSMD1 is causal/pathogenic for autism?

    2. Thank you Peter - I will ask for RS number. It is mentioned as Variant of uncertain significance.

    3. Hi Peter, Have an upcoming Genetics appointment where I plan to request a re-analysis of my son's Whole Exome sequencing test from ~3 years ago. I will ask for the RS number for the mutations identified. I need them to specifically rule out or confirm a suspected genetic syndrome (shows a variant of PTHS but report is non-committal on whether it is PTHS). You mention channel mutations elsewhere in your blog, if the original WES results did not indicate a channel related mutation, do you think it would be there and it did not get mentioned as the analyst/reporting software is looking for Autism / MR causal mutations? Is there a list of areas / mutations that I should explicitly request be looked at or analyzed ? Thank you.

    4. There are hundreds of known mutations in the TCF4 gene; many are classed as pathogenic for Pitt Hopkins Syndrome (PTHS) and some are of unknown significance. When someone is diagnosed with PTHS their mutation eventually gets added to the list, so the list is always growing.

      You can diagnose these syndromes based on their known features. Your son might be the first one to be recorded with that mutation.

      All these syndromes are spectrums; sometimes facial features vary and indeed IQ can vary.

      You can ask for a list of all the mutations they found, not just those associated with autism or ID. The list is unlikely to be more than 20 to 30. Then you just look each one up.

  4. Thank you, the possible use of PQQ in CFD is quite interesting, I will read up on that.

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  6. Torasemide is another nkcc1 drug here is a link on proprionic acid, Torasemide, and asd.


  7. Hi Peter,

    Have you seen this product from Dr Borger? ? It’s doing the rounds of the internet at the moment and it seems to have a similar effect to Folonic acid i.e. increased speech in some, bad/aggressive behavior in others.
    One another note, we are patients of Raemarkers at the moment and he was saying that if both parents have Folate Receptor Antibodies then Folonic acid will have almost no effect. If only one parent then lowered effect. If both parents without then high effect.

  8. I am guardian for 63 year old sister (who has auditory hallucinations and autistic symptoms since very young. With help of an independent (private practice in US) psychiatrist I have her on liquid calcium folinate (with B12) by Designs for Life. Dose is about 9 mg DFE, 3 times a day, 30 minutes before or after eating. (see Google Scholar & research on folate absorption.) Now it seems worth trying PQQ, administered with a reduced calcium folinate dose, to see if she shows more benefit to mood and cognition and overall health. Assuming no adverse effects, this "experiment" should run at least 6 months. She had the Folate Receptor Alpha Auto Antibodies test in 2018 and it wasn't conclusive. We do know her use of calcium folinate has improved mood and cognition since it was started in 2019. Efforts to reduce or increase dose cause worsening behavior. I am looking for any research that might indicate HOW to administer PQQ (as liquid so it can be titrated) and liquid calcium folinate in the same 24 hour period. If oral PQQ had a long time to peak (in serum?) could it be given first and four hours later give folate? It would be nice to avoid multiple experiments to find out some optimum schedule. My sister has stage 3a kidney disease and weighs 230 pounds. Her arthritis keeps her from moving around much. NOTE: Methyl folate oral at almost any dose caused extreme distress. DEPLIN 15 mg (before the calcium folinate) caused a week of psychosis and aggression. We never had a doctor to guide any of this process, only doctors who were willing to be flexible and support trying new things. For people in care of government funded programs (homes) all over-the-counter medications require a doctor's "order".

    1. Changing the dose of calcium folate is known to cause worsening behavior in some individuals. It doesn't necessarily mean that the dose is too high or low! A possibility is that calcium folinate affects serotonin and dopamine in the brain, and that it takes some time before the new balance sets in.
      For arthritis you might consider curcumin or fisetin. Both are known to promote health in general. It wouldn't surprise me if any of them could help with the other health and behavioral issues.
      Good luck!


    2. I do try to follow the rule of "stay low, go slow" on changes, but some behaviors are not long tolerated in group homes so it gets tricky to make any changes. My sister was put on curcumin 3 times a day by a rheumatologist and exhibited extreme anger several days later. The OTC curcumin product had black pepper (piperine) added to increase availability (?) but it also inhibits CYP450 enzyme(s). That could have affected metabolism of Vraylar -- another med she needs at a specific dose to stay in equilibrium. Nothing works in an ideal way when someone is older, has kidney impairment and takes a dozen different meds. It's pharmaceutical Jenga.

    3. Regarding the CKD, some people find that a low carb diet helps (and also low omega-6 fats, e.g. veg oils), and that the high(er) protein that may go along with that does not do any harm. Of course, changing her diet may not be easy or possible ...


  9. Hi Peter, Started Miccil about 20 days ago (0.5 mg twice a day) and just bumped to 1mg starting yesterday. It is hard to say if we are seeing anything different. We also don’t see any diuresis. The child is almost 13 years old weighing 105 pounds. Given the lack of diuresis wondering if my child is metabolizing this differently / non-responder. Plan on continuing with 1mg for 2 weeks if possible for now. Do you think the dosage is too low?

    1. Yes, I think your dose was low.

      I would trial 2mg for 2 weeks, before giving up.

      A sizeable minority of people seem not to experience diuresis from Bumetanide. Does this mean none of the drug reaches the brain? Nobody knows for sure.

      Do any people who experience no diuresis respond with improved cognition and behavior? We have no definitive answer, because many people do not share their results.

    2. My daughter was a non diuretic non cognitive bumetanide responder. I’ve often wondered if the diuresis was the key here. It something seems plausible in my non professional mind.
      Interestingly her school has commented that she never goes to the toilet at school despite drinking plenty of water. No idea where it’s all going.
      Is there any condition that could explain this?

    3. Bumetanide has to be metabolized in the liver before it can enter the bloodstream. Quite possibly some people cannot metabolized it.

      I think the logical follow on would be to then try Azosemide or Torasemide and see if this is metabolized, produces some diuresis and hopefully some cognitive benefit.

      There are people who drink either too much or to little. Whatever is drunk has to go somewhere. She might have an issue with using the toilet at school and so avoids using it.

  10. Peter, curious how the mom you mentioned managed to get a spinal tap diagnosis for her adult son? Did she share those details?

    1. She was very angry/frustrated it took so many years.
      Normally you need a helpful neurologist. It is possible, but you have to persevere. Even in the UK it is possible, the US is much better as long as you can pay. I doubt it would be covered by insurance, because it is not a standard investigation for autism.

    2. Agnieszka has a list of tests to ask for if you can get a spinal tap.

    3. Hi Tanya,

      I recall you've mentioned catatonia before. If this is the case, then spinal tap is a standard diagnostic step as per recently published UK guidelines (catatonia in autism is mentioned):

      Here is a report from the US about an adult with autism who developed catatonia and then was diagnosed with metabolic disorder by spinal tap.

      Unfortunately it's easier to find such papers than a real help, but at least there are now official guidelines published which can be a starting point to obtain these tests.

  11. Peter,

    Did you ever do an organic acid urine test for Monty? I guess my son has high propionic acid levels...


    1. Stephen, we never did that test. High propionic acid is common in autism and is thought to be caused by gut dysbiosis. Propionic acid does cross the BBB and will worsen autism. The mouse model of propionic acid induced autism is reversed by NAC.

      It looks like diet, extra fiber and a butyric acid producing probiotic should help. Vancomycin which you mentioned previously might help, or Rifaximin which is very popular for gut dysbiosis.

      Rifaximin is cheap and even OTC in some countries, but expensive in the US.

    2. I'm curious if this would work.

      The Therapeutic Effect of Bismuth Subsalicylate in a Propionic Acid-Induced Autism Model

      If not I'll see if Bumetanide will work.

      Torasemide Improves the Propionic Acid-Induced Autism in Rats: A Histopathological and Imaging Study,side%20effects%20after%20further%20studies.

  12. Hello Peter, for Pseudotumor cerebri, do you have any advice on alternative treatment? Diamox has many sideffects, thank you.

    1. Reducing the amount of fluid in the brain is normally achieved using a diuretic. Diamox seems to be the fist choice, but loop diuretics are also used (furosemide, bumetanide etc). Loop diuretics are usually well tolerated. Some people have no side effects from Diamox.

      Interestingly research shows that oxidative stress plays a key role in this condition. Taking NAC would seem a logical step to take.

    2. Remove the child tonsils/adenoids


  13. My 3 year old just tested positive for the binding antibodies. Does anyone have any insights on the best path forward from here? Hopeful for a treatment but even though to someone like me who reads Peter's blog the test seems almost mainstream, no one in my local autism parent groups seems to have heard of it, and no nearby doctor has anything about it on their website. We are in Indiana.

    1. Probably the easiest way is through Dr. Rossignol



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