Autism Research Connections #4.5: A Conversation with Dr Naila Rabbani

Dr. Naila Rabbani was interviewed by Seth Bittker over email in March 2018 on her recent research on blood and urine-based biomarkers for autism spectrum disorders. Dr. Rabbani is an expert in Translational Medicine at University of Warwick.  Her work focuses on disease mechanisms, particularly the study of damage to the proteome by glycation, oxidation and nitration. © 2018

Image result for naila rabbani

SB: Dr. Rabbani, congratulations on your paper “Advanced glycation endproducts, dityrosine and arginine transporter dysfunction in autism – a source of biomarkers for clinical diagnosis.”  What do you see as its most important findings?

NR: Thank you. I believe the most important finding is the emergence of the basis of a blood-based diagnostic test for autism that could be made widely available. This requires validation in a further large group of children with and without autism.

SB: There has been a lot of discussion of a need for discriminatory biomarkers for ASD.  A lot of biomarker research has focused on neurological biomarkers such as electroencephalography or eye-tracking.  Other research, such as yours, focuses on levels of compounds in blood or urine.  Can you comment on the relative utility of these two types of biomarkers?

NR: A blood-based test or urine-based test may be performed in the clinical chemistry departments of well-equipped hospitals and test results may be interpreted without the need for a specialist in child development.

SB: It is interesting to compare some of the findings in your research to similar studies.  You find lower arginine in urine in ASD with statistical significance.  West, et al. found decreased homocitrulline in the plasma in ASD with great statistical significance (West et al., 2014).   Is there a connection between West’s findings on homocitrulline and your findings on arginine?

NR: Actually, we found increased arginine in plasma and urine of children with autism, compared to those without. Renal clearance of arginine was lower in children with autism than those without. This relates to dysfunctional handling of arginine by the kidney. There is no link of homocitrulline to arginine. We used the most robust analytical method for mass spectrometry metabolite analysis – a technique called stable isotopic dilution analysis. The studies by West et al. should preferably be repeated using this method for surety of the outcome they found.

Our approach of studying spontaneously modified or “damaged” proteins and amino acids is more powerful than previous studies of unmodified proteins and amino acids. For proteins, this is because the combination of changes in protein modifications is likely reflecting changes in processes that drive ASD or are markers of them. For damaged amino acids, these are released from modified proteins by proteolysis. Unlike unmodified amino acids, when released they are not re-incorporated into proteins and accumulate – providing the basis of the diagnosis of ASD.

SB: Some research suggests that ASD is associated with methylation deficits and sulfation deficits (James et al., 2004; Adams et al., 2011).  Is there a connection between the markers you have found in ASD and methylation or sulfation deficits?

NR: No connection that I am aware of.

SB: A lot of research has shown ASD is associated with increased autoimmune activity (Mostafa et al., 2014; Careaga et al., 2013 for example).  Do you see some of the biochemical markers you have found as potentially a result of autoimmune activity?   Do you see some of the biochemical markets you have found as increasing the risk of development of autoimmunity?

NR: Dityrosine is formed mainly by the enzyme DUOX which has an important role in host immunity. Increased DUOX may be associated with autoimmunity in some cases.

SB: There is a lot of research on generic abnormalities and genetic polymorphisms that increase risk of ASD.  Some of this research is on genes that affect neuronal development.  Your findings do not seem to be directly connected to neuronal development.  Is there a connection?  If not, does this suggest development of ASD in some cases is dependent upon two hits (susceptibility to neuronal dysfunction in combination with metabolic dysfunction)?

NR: We were encouraged in that our findings support previous evidence from genetic studies – particularly those linked to amino acid transport proteins, arginine transport proteins particularly. Genetic polymorphism of arginine transport proteins affects all cells in the body, of course, and a non-invasive way to assess change in amino acid transport is to measure renal clearance which, with normal glomerular filtration rate, is influenced by amino acid re-uptake from renal tubules mediated by amino acid transporters. The decreased renal clearance of arginine and Nω-carboxymethylarginine (CMA) in children with ASD is consistent with a change in function of arginine transporter proteins as may occur in amino acid transporter genetic polymorphism.

Also, although we studied modifications of plasma protein in part of our study, the major protein in plasma – albumin – exchanges with albumin in cerebrospinal fluid. So some of the modifications we find of protein in plasma may have occurred whilst that protein was in cerebrospinal fluid in the ventricles of the brain. The increased CMA free adduct found in plasma filtrate may have originated in part from proteolysis of increased CMA-modified protein in the brain. Further studies are required to follow up the mechanisms that underlie these interesting changes found in ASD.

SB: Does your research on advanced glycation endproducts and ASD suggest anything about diets that should be researched further in cases of ASD?

NR: Advanced glycation endproducts (AGEs) in the diet are absorbed into the body as glycated amino acids. These are not incorporated into our proteins; they are rather excreted by the kidney. This likely poses limited challenge to body function with the changes found of renal function in ASD. The changes of AGE content of plasma protein reflects endogenous AGE formation – formation of AGEs in the body. For AGEs in the diagnostic algorithm, CML and CMA were increased in ASD and a further AGE, 3DG-H, was decreased in ASD. So, there is not a general increase in AGEs but rather an increase in AGEs formed by a specific precursor – glyoxal, and decrease of AGE formed by a different precursor – 3-deoxyglucosone (3-DG). We interpret this as likely increased glyoxal exposure from lipid peroxidation in ASD and increased activity of enzymes that metabolize 3-DG in ASD – which may be a protective response to increased 3-DG. The latter is a metabolic state called dicarbonyl stress. This complexity of response likely makes the characteristics of the changes specific to ASD and may account for the high accuracy of the diagnostic test based thereon. Again, further studies are required to follow up the mechanisms that underlie these interesting changes found in ASD.

SB: Are there other environmental factors that might contribute to the biochemical gestalt your research suggests is characteristic of ASD?

NR: We have not studied this and therefore have no comment.

SB: Was it difficult to get funding for this research?

NR: The study was part funded by grants obtained by our collaborators at the University of Bologna and part-funded by our own research group resources at the University of Warwick. Innovative research of this type is always difficult to fund via peer review system of obtaining grant funding. We hope now the advances made have been published and publicised by the media, funding bodies and reviewers supporting studies of ASD will be aware that there are initial-stage data and findings of potential diagnostic importance to build on and funding for further investigations will be forthcoming in due course.  Our innovative research will continue, I would like to repeat and validate our blood and urine test and translate it to benefit parents, grandparents and autistic community, who have been so desperately waiting for blood test like this for potentially a rapid diagnosis of their loved ones.

SB: Do you have any suggestions on how those who fund ASD research should determine which studies that they fund?

NR: I encourage all research funding bodies and funding application reviewers to maintain an openness to new developments, findings and ideas that may make a step-advance in diagnosis and understanding of ASD.  If something new appears that may not fall within the current strategy of funding bodies, then I encourage sufficient flexibility of strategic considerations to adapt to change and support follow-up of the new findings in service and benefit of the ASD community.

SB: Is there anything else that you would like people to understand about this research?

NR: I would like to thank the ASD community for the many correspondences of support and encouragement given to me and my colleagues since our recent publication. We will be working to take this initial step further for improved diagnosis to support clinical care and services for ASD. I will be setting up a www page to provide further information on our current study and further advances and for those who may like to support our further research.

This is the first-time low-level damage to proteins and amino acids in blood and urine have been studied in autism. The research found a link between ASD and damage to proteins in blood plasma by oxidation and modifications by glyoxal and sugar. The changes found suggest a link to low level inflammation, lipid peroxidation and a protective response to potentially damaging sugars (dicarbonyl stress). The most reliable of the tests used is protein in blood plasma.

Anyone wishing to make a donation to the ongoing study of autism can do so via the link ( Please make sure to specify ‘Autism Research’ in the My Own Preference box.



Autism Research Connections #3.5: A Conversation with Dr. Rosemary Waring

Dr. Rosemary Waring was interviewed by Seth Bittker on April 17, 2016 on sulfation deficits in autism, Epsom salts, the nature of autism, and prospects for future research.  Due to the poor quality of the audio recording, this interview will only be made available in transcript form. © 2016

Rosemary Waring

SB: This is Autism Research Connections [#3.5].  It is April 17, 2016.  I am your host, Seth Bittker, and our guest today is Dr. Rosemary Waring.  Dr. Waring has had a long and productive career as a research scientist focusing on metabolism and toxicology.  While she is technically retired, I have seen her name on a number of recent papers and she is an honorary reader in human toxicology at the School of Biosciences, University of Birmingham.  Dr. Waring is arguably the world expert on sulfur enzymes and the role of sulfation in inflammatory disease.  Specifically of great interest to our audience, Dr. Waring was the first to identify the sulfation deficits which often accompany autism.  Dr. Waring welcome to Autism Research Connections.

RW: Well thank you very much.  It is an honor and a pleasure.

SB: Thank you for being here.  Dr. Waring, what is sulfation?

RW: Sulfation is addition of a sulfur atom and four oxygen atoms, and makes everything more water soluble.

SB: Wonderful. And when the body does not have enough sulfate available what are some of the metabolic consequences?

RW: Well unfortunately sulfate is one of the major building blocks.  If you don’t have enough sulfate, you run into problems with sulfation of the gut mucins – these are proteins which line the gut.  You can also run into trouble because it is more difficult to form heparin sulfate and sulfated synovial fluid.  For instance you can get rheumatoid arthritis because you need sulfation for the joints to move and you can also have problems with sulfating neurotransmitters.

SB: Wow.  That is quite a picture.  Dr. Waring if I might highlight to our audience, you have published a series of remarkable papers starting in the mid-1990s on autism.  One is Biochemical Parameters in autistic children published in 1996.  Another is Sulphur Metabolism in Autism published in 2000 and I know you coauthored a number of other papers and perhaps I’m missing a few.  These are really significant findings.  Can you tell us what are those findings from your research regarding sulfation and autism?

RW: Well when we looked at children with autism we found that generally they had really quite low levels of sulfate in plasma and quite high levels in urine where the sulfate was leaking out.  Hence obviously this had a lot of knock on effects.

SB: That makes total sense.  What would some of those knock on effects be?

RW: For a start most people who we were looking at had gut problems, and they seemed to me to have so-called ‘leaky gut’.  They were much more susceptible to food intolerances I think because the food proteins get in through the gut wall to cause reactions much more easily.  You really rely on sulfating mucin proteins in the gut for their function.  If they are not sulfated, then you can get holes in between the mucins.  Obviously this will lead to a lot of problems.  So you can get gut problems, a number of children we looked at had neurotransmitter problems with high levels of serotonin and dopamine for instance, and many of them came from family backgrounds with rheumatoid arthritis.

SB: These are really important points.  So you are saying if somebody has leaky gut, they really should be looking at whether they have low levels of sulfate.  Is that correct?

RW: Well I think it is one of the main factors.  You see normally you rely on the gut to be non-permeable except for what you actually want to absorb.  The lining on the gut is renewed pretty regularly – about every three days, but if it has holes obviously that is not going to work in the same sort of way, and this is where the real trouble comes I think.

SB: That makes total sense.  So what happens to people who have reduced sulfation capacities in a world of chronic low grade exposure to xenobiotics?

RW: Well people are more susceptible to drugs.  For instance they tend to be more susceptible to what we in the UK call Paracetamol, what you call Tylenol.  And you also often find people are more susceptible to amines for example in cheese, chocolate, bananas, citrus fruit, and some parents of autistic children  have found their children have terrible reactions to these foods.  So it just creates all kinds of problems getting rid of quite ordinary things.

SB: This is terrific insight.  When we suspect there are issues with sulfation what are some of the most effective markers for flagging it?

RW: What I always suggested to people is that they try to increase the sulfate to see if that improves the problem.  I feel if you put more sulfate in and things get better, that tells us it was needed.

SB: That is a terrific low-tech, very practical solution.   I guess I also wonder does it make sense to look at markers of blood sulfur or not really?

RW: Well you can do it.  I think one of the problems is some of the markers for instance neurotransmitters  can be quite difficult to look at.  You could do spinal fluid couldn’t you, but that would be a bit much.  I really think it’s easier to put in more sulfate to see if things feel better.  Because you get such different things reacting to sulfate it is impossible to say if you see X, then Y.

SB: That is wonderfully practical advice.  I know a lot of children with autism have issues with sulfate.  Do you have an estimate of what percentage of the children – I should say individuals because many are not children – with autism have abnormalities of sulfation?

RW: Well we looked a series of about 150 children with autism – this was just a random sample really – and about 60% had issues with sulfate – a significant issue.

SB: …Do you recall what the definition used of issues with sulfate is?

RW: Well these were children who had a lot of gut problems.  They are the ones that had the lowest levels.  We did find low levels in all of them, but I think they were lowest in the children with gut trouble.

SB: Very fair.  So is the sulfate deficit in autism due to excess exposure to chemicals – xenobiotics, inadequate consumption of sulfur containing compounds, or is genetic predisposition?

RW: Well I think it is all three potentially because we know one of the problems with sulfate is that is not that easy to absorb huge amounts of it.  You usually rely on cysteine and methionine – the amino acids sulfur and most of the work is done by cysteine dioxygenase and sulfite oxidase.  It is a design fault because these enzymes are easily inhibited.  You can have people who have an ineffective enzyme for a start.  You can have people where an enzyme is being inhibited by various factors.  You can people who don’t have much sulfate in the diet.  You can have people who seem to have an overload of xenobiotics who have depleted their sulfate.

SB: It makes complete sense.  You mentioned in some instances that it may be a defect of genetics.  Does this indicate that as a species we are not adapted to some of the exposures we are getting?

RW: Well I think some people are less able than others.  There is a huge spectrum.  For instance drinking alcohol, now we all know somebody who perhaps will drink half a pint of beer and is really pretty drunk at the end of it, and other people who drink half a bottle of whisky and you would never know they had been drinking.  There is huge variation with response to alcohol and huge variation with response to drugs.  I think that what makes autism quite difficult to explain to lay people is to say that there is this variation.

SB: That makes sense.  Your research also shows that sulfation abnormalities seem to be connected to a number of other degenerative and autoimmune diseases such as rheumatoid arthritis and ulcerative colitis.  What typically comes first in your view, sulfation deficits leading to immune dysfunction or immune dysfunction leading to sulfation deficits?

RW: Oh it is immune dysfunction leading to sulfation deficits.  When we looked at human cysteine dioxygenase which is the principal enzyme to form sulfate ions, it is inhibited by pretty much all the cytokines in the inflammatory process, where TNF-alpha is the major one.  So if you have any inflammation as in rheumatoid arthritis for instance you get a high level of TNF-alpha and you knock down the level of sulfate.  So you get what you do see in RA which is a sort of spiral where you get worse and worse.

SB: I see.  That is a really good point.  So putting this back in an autism context if somebody has a low sulfate and potentially autoimmunity associated with it, you can maybe add a sulfation agonist to the diet and that may help some but it may not get at the underlying dysfunction.  Is that what you are saying?

RW: Yes.  As a matter of fact I think nobody has ever tried it, but some children [with autism] have high levels of TNF-alpha.  Now in rheumatoid arthritis you prescribe the antibodies to TNF-alpha which have been really very successful.  Would it work in autism?  I don’t know, we have never tried.

SB: It’s a great question, and I hope somebody looks at that.   I think there are some people who have done intravenous immunoglobulin in autism as a similar sort of thing and some work and some don’t.  Do you have any comments or thoughts on that?

RW: I don’t know, I have never tried it myself.  I am not a clinician.  I have talked to some people who tried it and they say sometimes it helps.  I think the real trouble is we don’t know who would benefit.

SB:  That’s a very good point.  Are there certain markers that you think people ought to look at?

RW: I don’t know.  It would be interesting to look at those children who had those treatments and had who benefited and see what the markers were opposed to those who did not benefit.

SB:  Wonderful.  We have been talking a lot about this sulfur deficit in autism – or sulfate deficit excuse me – I should say.  What therapies are most effective at remedying the sulfation deficits that we often see in autism?

RW: Well I think one of the most helpful ones is Epsom salts.  You can have a bath in it.  You have people who wash their hair with Epsom salts and that works perfectly, and you can actually put it against the skin with a plaster to hold it on.

SB: That is fascinating and I am glad you brought this up.  So for our audience we are talking about magnesium sulfate – Epsom salts.  I believe you.  You know this stuff but can you give us some information on what evidence there is that this actually works?

RW: Well we have looked at it in normal people.  I have never looked at it clinically in autism.  In normal people if you have an Epsom salt bath you do get much higher levels of sulfate in plasma, with  more coming out in urine.  You don’t see much change in magnesium because in normal people magnesium is very tightly controlled.  So there shouldn’t be much change and you don’t see any.  In children with autism I  know people who have talked to me or phoned have told me it is very helpful.  I don’t think people will phone all the way from Australia to the UK unless they mean it.  It has to work.

SB: That is wonderful.  I have seen I believe some of your research on Epsom salts effectiveness posted on a website.  Is this in the medical literature or is this more tests you have run if not formally published?

RW: Oh there were certainly tests that we formally published.  I had a lot of trouble finding a journal that would take it, and rather left it for a bit and did a little more work.  And then a colleague of mine in London used magnesium sulfate and was able to show that it crosses the skin.  Now if anybody asked, I would have said it doesn’t, but it certainly did.  So that was interesting.  We have written a review that I guess is out either now or very shortly in Xenobiotica.  I think it is there electronically.  It is all about transport across the skin – across biological membranes, but in fact when we looked at the literature we found that quite a few other people had shown that Epsom salt cross the skin.  So it is an interesting finding.

SB: It is.  It is wonderful and it is very important.  [Are] there other things that people who have issues with sulfation or sulfation deficits might want to consider doing in terms of I think you mentioned cysteine and methionine – or is that something that is too complicated?

RW: Well if the sulfur oxidation was faulty this might not work terribly well, and all amino acids are toxic anyway.  You could certainly try to take a small amount.  I know some people take methylsulfonylmethane (MSM) and say that is helpful.  You don’t know what the problem is in any given individual, and so the thing to do is to try a small amount and see what works.

SB: That makes a lot of sense.  You made an interesting point there about amino acids being toxic.  I imagine we are talking about a dose effect, correct?

RW: Yes.  You certainly don’t want to take any more than about 200 mg of any amino acid and you would not want to do it often.

SB: Very fair…  On the Epsom salts, if the Epsom salts go through the skin, and it sounds like there is very good research that they do, what is to keep them from going through the skin and you getting an overload?

RW: Well you probably do because people who have tried it have usually told me that the best thing to do is to have baths perhaps two or three times a week for three weeks and then stop for three weeks and then start up again.  I would assume because they are talking about headache here we are talking about some kind of overload.  Not of sulfate because we get rid of that quite easily but probably magnesium.

SB: That is fascinating.  I honestly did not expect that answer.  I was thinking… let’s say you go into the ocean would you also get the same effect or is it just with concentrated magnesium sulfate?

RW: Well I don’t know.  We have never looked.  I would think you would get the same effect.  I think it goes across better if the water is warm.  Perhaps you could get it in the Pacific Ocean?

SB: Very interesting. If you are absorbing sulfate is this also affecting levels of other sulfur containing compounds such as things like cysteine and methionine?

RW: We don’t know.  We have never looked.  I would think not.  Sulfate is pretty much a sort of end product.  You can have quite high levels in plasma without any ill effects.  It just comes out through the kidneys.

SB: Wonderful.  Are there drugs or other compounds that we should avoid providing to those with autism who also have associated sulfation deficits?

RW: Oh.  It depends on the drug.  I would certainly be happy to not use Paracetamol.  Your Tylenol… You would have to look at the metabolism of the individual drug and see whether it did affect sulfate.  Offhand I can’t think of any specific ones.  No family of drugs.

SB: That is very interesting, and with respect to Paracetamol or Acetaminophen, I think there is some research indicating that people who have higher rates of use have higher rates of autism.  Does your research on this give that associated research some additional credence?

RW: That’s what people have told me…. It is the impression I have got.

SB: That’s wonderful.  Are there research groups working on extending your sulfation research in autism today?

RW: Well yes.   There is Jim Adams in Arizona.  Isn’t he at Arizona State?  He has done a lot of work on autism and certainly replicated our sulfate results and as far as I know he is continuing that line.

SB: I am glad you brought his work up.  Just as an observer of all this it does seem to me when Jim Adams not only reproduces your work on low sulfate in the blood but also has a high sulfur multivitamin which provides benefit to many with autism in some ways he is validating what you have already found.  Would you agree?

RW: Yes.  I think highly of his work.  I think he has done some very good stuff, and it is in line with a lot of work in the literature.  Funnily enough there is a mouse model for autism and this mouse has low levels of sulfate as well.  It is a basic problem.

SB: It really makes sense.  Dr. Waring should we be talking about autism as a single disease entity or autisms?

RW: Well it isn’t a single disease.  I think it is one of the problems in the field that a lot of researchers tend to take what looks like a rather heterogeneous set of children and then try to get sense out of it.  Certainly… with 150 children, obviously that is a very small number, we were able to go into three main groups.  So the problem is that we have got other groups out there.

SB: And what were those three main groups?  Do you recall?

RW: Well about 60% had gut problems and they came from families with autoimmune dysfunction of some sort – rheumatoid arthritis, ulcerative colitis, eczema, asthma, all that stuff.  They seemed to be very different from other groups.  About 15% of the children had what I think are damaged hypothalamus pathways.  They tended to be very thirsty – to drink gallons of liquid if they could get it – obviously had problems with vasopressin levels.  This is damage pre-birth.  Then there was a last group, about 20 plus percent, who I think had frontal lobe damage.  They tended to have problems with handedness and had very vivid dreams.  They seemed to be much more autistic than the children with gut problems.

SB: That is really interesting stuff.  So when you split these children up into these different groups were there any markers with which you could do it or are you doing this all based on observation?

RW: We asked the parents/carers to fill in two sided A4 questionnaires.  You can’t make it too detailed because otherwise [they won’t fill it in] but it was quite enough to divide children into one of the three groups.  I always thought if we had a much larger group and a more detailed questionnaire you could get more detailed grouping, but they were very different biochemically and in autistic features.  So I have always felt that autism is a catch-all term.  It is a bit like saying you have an infection but this could be caused by any number of things.

SB: That makes complete sense.  So I believe if I am understanding you correctly you would only think of suflate as a potential therapy in the group that has 60% gut problems or would you be thinking of it in all groups?

RH: Well it did work in all groups, but the ones that responded the best were the ones with gut problems.

SB: That makes sense.  I know that you are not a clinician, but from your other work in biochemistry are there other types of over-the-counter supplements or treatments that people in that gut problems group should be given or at least have as a trial?

RW: I can’t think of anything off hand that is really for gut problems, and I think that is one reason why [they] are so intractable and people usually respond by going on a casein free / gluten free diets which seem to work in some children.

SB: That is a great point, and this is something that I know there still seems to be some debate about. On the casein free / gluten free diet, how does that connect in with your sulfur research?

RW: Well we looked at this in some detail but not in a very large number of children.  One of the problems with casein is that it is  obviously broken down in the gut by peptidases like all proteins.  In fact casein is not that easy to breakdown and some people are much better at it than others.  One of the basic problems is that you have a sequence in which you sulfate a protein called gastrin which is obviously in the stomach, and the sulfated gastrin then causes release of sulfated enzymes in the pancreas which lead to breakdown of carbohydrates, lipids, and proteins.  But if that initial sulfation and further sulfation of the enzymes does not occur, then you don’t get breakdown of proteins into amino acids or tiny peptides.  You get quite long peptides and if you then have a leaky gut as well, I think these peptides get into the blood stream and that gives you the cause of the problems.

SB: This is a wonderful portrait you have just painted.  It makes complete theoretical sense, and I have seen lots of anecdotal reports.  It works.  I actually happen to be one of those that believe that it works.  At least from my reading research is mixed on whether it is effective or not – say in a double blinded trial.  Do you have any thoughts or comments on that?

RW: Well actually I think it is very difficult to do a true double blinded trial on something like that.  I think that is a basic problem.  Again I find myself saying, try it out, it should work in two to three weeks max.  If it seems to help then go on with it.  If it doesn’t, then don’t bother with it.

SB: This is one of the wonderful things I am learning about you. You are intensely practical Dr. Waring, and that is a wonderful characteristic for those of us who are trying to understand some of your research and apply it.

RW: Well it is no use having things that are too complicated.  I think it is quite difficult if you have children with autism to go on a really complex diet, and then there is the rest of the family to consider and so on.  I think it is only worth it if they seem to benefit.

SB: That’s wonderful.  What other autism research that you have seen in recent years do you feel holds significant promise?

RW: What has been really exciting in recent years is the rise in the number of studies that have shown that children with autism have high levels of cytokines – the markers released in inflammation.  Now what we don’t understand is why they have got these levels.  Often the siblings have medium high levels.  Well there is obviously a family problem.  Why are they up?  Everybody has raised level of cytokines with an infection, but why don’t they go down to normal?  There is obviously a basic problem with the immune system.  In studies in children on autism – the little one we did with about 100 children, mothers were 6.4 times more likely to take antibiotics during pregnancy.  Now that is a huge difference isn’t it, given that they really don’t like to give anything if you’re pregnant.  So that is quite serious.

SB: That is a very big clue.  So it almost sounds like are you saying there is immune dysfunction and that is related back to the gut presumably.  Is that correct?

RW: Yes.  I think there is immune dysfunction, and I think this means that the brain tissue of the fetus as it is developing is not laid down as it should be.  Certainly in the mouse model they have less sulfated heparin for instance and less sulfation of brain markers as well.  So as I see it, what happens is you get some sort of infection in the mother.  For some reason it does not die away.  Perhaps she has persistent chronic infection.  We really don’t understand that much about the immune system.  This then leads to trouble when the baby’s brain is being formed.

SB: That makes complete sense.  If it was that particular case, why would the baby’s immune system be affected longer term?

RW: Well the bottom line is that we don’t know.  There is nothing that really explains it.  Why the baby’s immune system doesn’t drop back down to normal?  But they don’t.  They appear to have chronic inflammatory conditions which they don’t clear properly.

SB: Makes sense.  Dr. Waring is there anything else you would like out audience to know?

RW: Well what I would like to see for autism long term is to look at the epigenetics.  These are where environmental factors don’t affect the original DNA but what they affect is the reading of that DNA.  So they turn the reading of that DNA on or off.  Now I think if we can show epigenetic effects as well as other things, this will really give us a clue on what environmental triggers to avoid.

SB: This a very important point.  What kinds of studies would show epigenetic affects?

RW: You could take a group of children with autism and look at epigenetic variations of some common proteins – methylation in particular.  Anyway you look at the DNA and just see what the differences are in methylation patterns.

SB: That’s fascinating.  Is there anything else you think we should be looking at?

RW: Well not really.  A better understanding of inflammation in general would probably be the most helpful thing.

SB: That makes complete sense.  Dr. Waring thank you for this interview, and thank you for being with us on Autism Research Connections.

RW: Well I’ve enjoyed it.  It has been a pleasure.  Thank you very much for asking me.

Autism Research Connections #3: A Conversation with Dr. Derrick Lonsdale

Dr. Derrick Lonsdale was interviewed by Seth Bittker on April 12, 2016 on autism, thiamine deficiency, thiamine supplementation, and TTFD. Dr. Lonsdale observed that thiamine deficiency is often present in autism. He sees autism as typically a metabolic disease that combines genetic risk with environmental stress and in some cases marginal malnutrition. © 2016

A podcast of the interview is available here.  A transcript of the interview appears below.

SB: This is Autism Research Connections #3.  It is April 12, 2016.  I am your host, Seth Bittker, and our guest today is Dr. Derrick Lonsdale.  Dr. Lonsdale has had a long and productive career in medicine.  At one point Dr. Lonsdale was head of Biochemical Genetics at the Cleveland Clinic, but he gave up this position and decided to become a pediatrician focused on nutrient based therapies at the Preventive Medicine Group.  Dr. Lonsdale is arguably the world expert on thiamine deficiency and supplementation of thiamine analogs in those with autism and other conditions.  He has written a number of papers on these topics, and in many cases he has significantly improved the lives of his patients in circumstances when other physicians had given up.  Dr. Lonsdale, welcome to Autism Research Connections.  Thank you for being here.

DL: Thank you for asking me.

SB: Dr. Lonsdale, as I alluded to in the introduction, I understand that you were head of Biochemical Genetics at the Cleveland Clinic, a very prestigious position, but you left there in 1982 to join the Preventive Medicine Group specializing in nutrient based therapies.  What drove you to make this decision?

DL: Well it’s an interesting question.  I was a pediatric oncologist for six years, and I gave that up to study the inborn errors of metabolism.  Those that could be treated could only be treated by dietary means.  So I was interested in picking up these metabolic disorders, and one day a six year old child came to my attention because he had had episodes of cerebellar ataxia.  That is just sort of like being drunk if you will, and he had had every test under the sun with the question of whether it was encephalitis or whether it was a tumor.  All the tests had been negative.  Well to cut a long story short, we found out that he was suffering from what became the first published case of vitamin thiamine dependency – not deficiency.  This is a situation where the linkage or bonding of the cofactor thiamine to the enzyme is disturbed genetically speaking.  So the bonding requires enormous doses of the cofactor, and he required as much as 600 milligrams a day of thiamine whereas the recommended daily allowance of it is between 1 and 1.5 milligrams.  So it was a humongous dose, but what was so interesting about him was that he never got an attack of ataxia unless there was some kind of stress factor – an infection like a cold, on one occasion a light head injury, and on another occasion even an inoculation.  So that the stress factor was an important part of the initiation of these attacks, and that case drove me on a research way of going.  I spent a great deal of time in the library and studied thiamine in all its different aspects and that went on right through my career, and I am still studying it after retirement.  So that is about the answer to your question.

SB: I mean that is terrific.  We are sure glad you embarked on that journey.  So if we were to look at somebody with thiamine deficiency, what are some of the macroscopic symptoms of thiamine deficiency when you see them?

DL: Well thiamine deficiency causes the well-known disease called beriberi.  It is one of the standard vitamin deficiency diseases and it is a disturbance of glucose metabolism.  So that if we take our cue from beriberi, we have known for many many years – from the very earliest discovery made in this disease that if the patient had a normal blood sugar, the patient would respond to doses of thiamine.  If he had an elevated blood sugar, he responded poorly, and if he had a low blood sugar, sometimes he did not respond at all.  So this probably represents the staging of the disease – the seriousness of the disease – as it progresses.  Also we find that the triglycerides in the lipid profile are usually elevated.  It is almost diagnostic of sugar intake.  It is sort of interesting that in 1973 a book was written called “Sweet and Dangerous” and in that book a professor of nutrition at one of the major London hospitals had discovered that cardiovascular disease was due to sugar and not due to cholesterol.  So he was way way way ahead of his time and he found that many diseases were due to intake of sugar, but never came to a conclusion as to why sugar caused so much disease, and I think the answer to it is that because sugar has been known since 1936 to induce thiamine deficiency.  So it may be that thiamine deficiency is an extremely common phenomenon in America because of the high intake of sugar.

SB: That is a fascinating connection.  So you mentioned this issue with sugar and the high triglycerides.  What are some of the other metabolic markers that people would see in cases thiamine deficiency – and maybe not the most extreme cases of true beriberi but maybe more modest deficiency?

DL: Well the trouble is that you get some of the standard laboratory tests such for example as CRP – C-reactive protein and other inflammatory markers are increased because thiamine deficiency does produce inflammation, and I think that what is happening is that because thiamine deficiency simply does not exist in the lexicon of the American physician for many reasons.  Particularly because they say well the food industry has been using vitamin enrichment.  So we don’t have to worry about vitamin deficiency diseases.  So when these inflammatory markers are elevated, it is ascribed to some other cause.  For example we now know that rheumatoid arthritis is as much of a mental disease as a physical disease, and the inflammation is conducted from the brain, and the brain is defective in thiamine deficiency.  So you keep getting back thiamine deficiency as a cause of many different conditions.

SB: That is an excellent summary.  From some of your papers I also understand that thiamine deficiency is involved in you mentioned inflammation, maybe relatedly things like oxidative stress.  You mentioned the high triglycerides.  It looks like there is an issue with fatty acid metabolism when you have thiamine deficiency.  Is that correct?

DL: Yes and even worse than that, it has recently been found that it is absolutely essential for alpha oxidation.  Now alpha oxidation is a preparation for what is called beta oxidation and beta oxidation is one of the ways in which energy is produced in the energy producing mechanism.  So we now know that thiamine presides over sugar metabolism.  We have known for a long time that it presides over aspects of protein metabolism because it is necessary for the oxidation of the three branched chain amino acids – leucine, isoleucine, and valine.  So it actually presides over diet in general because carbohydrate, protein, and fat make up the entire diet.

SB: That is great color, and just to highlight for some of our listeners there probably [are] some parallels between what you are saying is common biochemical indicators of thiamine deficiency and autism.  Would you like to comment on that Dr. Lonsdale?

DL: Well as you know I did a pilot study in which I studied ten autistic children and eight of them improved as a result of giving a derivative of thiamine known as thiamine tetrahydrofurfurl disulfide and this is actually a prescription item in Japan by the name of Alinamin.  It is also referred to as Fursultiamine and Allithiamine, and it has got various trade names, but they are all the same substance, and I gave these children big doses of it by rectal suppository because of the taste factor.  The taste is so horrible that I couldn’t get them to take it by mouth.  Well when we finished the study – and the whole idea of the study was as a pilot study to go on and then we wanted do an interinstitutional study, but the FDA wouldn’t let us do it because we had to get a separate IND if the TTFD was given by suppository, and we couldn’t do that.  It was just impossible.  So all I can say is that this needs to be further researched because if TTFD is a valuable nutrient in autism, then it certainly should be researched further.

SB: Right.  That is outrageous, and your study as far as I could see was very impressive.  It was just as you mentioned it is not like a double blinded trial.  It is just a case series, but it did highlight that many of these kids did have thiamine deficiency as you mentioned and many of them improved.  When you are looking at measuring thiamine deficiency what measure are you using, and what is the standard criteria that you would use for determining whether somebody does have thiamine deficiency?

DL: Well there is a very accurate study called red cell transketolase.  Thiamine and magnesium are cofactors to a very important to a very important enzyme known as transketolase, and transketolase occurs in the brain.  It occurs in red cells.  So you can measure the enzyme activity in the laboratory and then you come out with a baseline figure of how active that enzyme is, and then you add thiamine pyrophosphate to the reaction and repeat the reaction, and if you find that activity of the enzyme accelerates it is simply saying OK this is what we needed thank you.  Now we can reach our maximum efficiency.  So it is an excellent test for thiamine deficiency.  The trouble is that the Mayo clinic for example has published a statement that the only way of measuring thiamine is the level in the blood and that is totally inaccurate because that level can be completely normal in the presence of thiamine deficiency.  The baseline transketolase activity can also be normal with thiamine deficiency.  So you can be hoodwinked.  You can miss a fairly severe case of thiamine deficiency by doing the test improperly.

SB: This is a really important point.  So to actually measure it properly is sounds like you do this reading and then you need to provide it, is that correct?  And then you do the reading again to see if there is a difference?

DL: Yes.  Exactly.  If it accelerates, then the enzyme was not saturated with the cofactor.

SB: It sounds like you may have already answered this, but just to be clear is there possibly some other biomarker that you think people should look at to see oh this person likely has thiamine deficiency, and then later on you could do the confirmatory test to provide the thiamine?

DL: Well you don’t need any other test.  This is a definitive test.  It is absolutely the most accurate test that you could possibly imagine.  I have literally done thousands of these tests and when I have given the patient thiamine or TTFD or magnesium or whatever, the transketolase activity becomes normal and the patient’s symptoms disappear.

SB: That’s terrific.

DL: It’s very very accurate and very very closely relates to the clinical situation.

SB: That’s great.  So there [are] various forms of thiamine and you have already described the TTFD form.  What other forms of thiamine are there, and what are their advantages and disadvantages in your view as supplements?

DL: Well that is a good question, and about mid-century Japanese investigators were looking at the biochemicals that are in garlic, and they found that there is an enzyme in garlic which acts on thiamine, and converts it into a disulfide derivative of the vitamin, and because it was found in garlic which is a member of the allium species it was referred to as allithiamine – A-L-L-thiamine – all one word.  They thought originally that it had lost its biologic value, but when they used it in animal studies they found it had a biologic activity, which was increased over the biologic activity of the original thiamine from which it had been derived.  And they did many many studies in animals and in human studies, and showed that I really had extraordinary therapeutic value.  For example they were able to show if they pretreated mice with TTFD which is the most modern of the disulfide derivatives, the mice were partially protected from cyanide poisoning.  They also found that it could prevent carbon tetrachloride poisoning of the liver.  These are amazing experiments which we had ignored.  So the Japanese went on to study these many different disulfide derivatives, and they found that the tetrahydrofurfuryl was by far and away the best, because the previous ones had made the patients smell very strongly of garlic which made it very very unpleasant.  They also made a whole series of what are called acyl (A-C-Y-L) derivatives and these are nondisulfide derivatives and in order to process them they require an enzyme in the body whereas the disulfides are automatically reduced at the cell membrane and they pile the thiamine into the cell where it is needed.

SB: That is great.  I also recall from your search that TTFD in animals…

DL: Well we know it crosses the blood brain barrier.  Benfotiamine which is has become quite a popular one is an acyl derivative and it does not cross the blood brain barrier.  That’s been proved by basic studies done in Belgium – in Liege University, Belgium.

SB: That’s great, and I also recall that TTFD I believe there were some studies you mentioned – or in some of your research where it had protective capabilities against lead and some heavy metals if I am recalling correctly?

DL: That’s correct.  Yes.  It’s interesting because thiamine actually the mechanism is not by any means clear, but it clears the lead through the liver.  It goes through the bialary system.  So the lead comes out in the stool.  Not in the urine, and what people are looking for is lead in the urine and they should be looking for it in the stool if they use thiamine.  Well of course nobody is using thiamine because they don’t know about it, but I would have loved to have got in touch with the people in Flint and tell them that the kids that have lead poisoning all they need is 100 milligrams of thiamine a day and that will ease the lead out of their systems.

SB: That’s well said.  In our discussion a moment ago of the various forms of thiamine I know there are the standard water soluble forms, there is the TTFD form, there is benfotiamine.  Am I correct in thinking that you favor the TTFD based on your research?

DL: Oh absolutely.  I think it is the most superior product.

SB: And also when we look at autism and thiamine – well frequently in autism as you know often times there is a sulfur deficit, and by this I mean often time we see excess sulfur compounds in the urine and too little – or deficit – in the blood.  Is there a connection between thiamine supplementation…

DL: Well that is an interesting question.  I know the researcher in Britain who reported that.  We were unable to repeat it when we did it ourselves.  We were unable to repeat it when we did it ourselves, but I think that the work that was done in Britain is authentic.  I think the answer is this.  If you take an example of a machine like a car, it has an engine. Well in the human body mitochondria are the equivalent of an engine.  Well a car has to make energy and then has to transfer it to the wheels, and that is through a transmission.  So the citric acid cycle and the electron transfer chain represent the mechanism of the engine.  That is where the energy is produced, but then transsulfuration which is what you were talking about about the sulfur – transsufluration – represents the transmission.  It is an energy consuming device.  The citric acid cycle produces the energy and the traunssulfuration, transmethylation mechanism consumes energy, and that enables us to what we call function.  In other words, cells function.  So we function.  Does that make sense?

SB:  It makes complete sense.  And I thought it was interesting the way you have tied this in with the mitochondria.  So when you do have thiamine deficiency – or close to it, is it accurate to say you have some sort of mitochondrial dysfunction or at least it appears that way?

DL: It has been actually proved that autism is a mitochondrial disease.  It has also been shown by Bettendorff in Belgium that thiamine deficiency will damage the mitochondria, and in the early stages – these are in vitro experiments.  In the early stages of thiamine deficiency in the mitochondria they can be repaired simply be reinstating the thiamine.  So thiamine is in an inherently essential component to mitochondrial function.  Of course it is not the only one, but it has a dominating influence.

SB: That is well said.  What percentage of those with autism do you think could have their functional level improved with some form of thiamine supplementation?

DL: Well our pilot study showed eight of ten had improvement.  It is interesting that only – I think either only two or three of them had evidence through an abnormal transketolase, but in spite of that eight of those children improved significantly.  Now we used computer read forms in the study that had been invented by the Autism Research [Institute] that Rimland ran and they had been tested over time as being extraordinarily accurate.  As you know there is actually no biologic marker that identifies autism per se.  So this symptom reporting device was used in the study and even though it wasn’t a controlled study it showed a definite advantage in the use of TTFD in those kids.  We’d have loved to have done a complete study after that between the institutions but as I say we weren’t allowed to do that.

SB: It is really unfortunate that you weren’t allowed to do it.  Why…

DL: They said that we had to take out a new IND if the thiamine was given by rectal suppository or on the skin or whatever.  It is typical FDA.

SB: It is an outrage.  So why do you think there are so many individuals with autism who seem to be thiamine deficit?

DL: Well the only thing that I can do is to guess that it’s the sugar intake because it starts in pregnancy.  Now I came across a case just recently where I was asked to give some help to a boy that has become a screwball like so many of these kids are today.

And so I said to his mother, “Did you have nausea and vomiting in pregnancy?”

“Oh yes,” she said, “Yes.”

I said, “Hyperemesis?”

“Yes.”  Hyperemesis that has actually been shown to be a thiamine deficiency disease.”

I said, “Did your son have jaundice at birth?”

She said, “Yes he did.”

And I said, “Did he have colic?”

She said, “Yes he did.”

Both of those – jaundice and colic – have been shown to be because of oxidative inefficiency.  Actually it is free oxygen radical pathology.  So you start the ball rolling right at the beginning of life, and the mitochondria are injured at birth.

SB: Thank for that additional information.  When you provide TTFD or supplemental thiamine do you ever see cases of it inducing other deficiencies?

DL: Oh yes.  I think we have to remember that when you are talking about something like thiamine that it is a member of a team.  Magnesium is its first cousin.  I never give thiamine without magnesium because they are double cofactors to the enzymes that are represented, and I think that what we call the noncaloric nutrients – the vitamins and essential minerals – really have to be regarded as a team.  And so what I have done for many of these situations, I give large doses of thiamine and magnesium, and then back it up with a well-rounded multivitamin.

SB: That makes a lot of sense.  When you are giving the magnesium what form would you normally give?

DL: My favorite is magnesium potassium aspartate.  Potassium is a very important noncaloric nutrient and aspartate actually enables the ingredients to pass through the cell membrane.  It sort of helps to load the magnesium into the cell.  So it is a very useful way of giving it.

SB: That is very interesting.  So aspartate as I recall is an amino acid, but you are not concerned that would somehow overload that particular amino acid?

DL: Right.  Right.

SB: So you have mentioned magnesium, a multivitamin, and thiamine.  Are there any other supplements that you think are underutilized?

DL: No. I don’t think you can pick on any one in particular.  The research that is needed is to identify which of the nutrients is apt to be missing, but the commonest way in which these nutrients are deficient is through poor nutrition.  Nutrition in the United States is absolutely awful because so many people are eating junk food as a staple.  Good food and good nutrition is at a premium.  It is hard for people to buy it.  So they dodge it by getting cheaper stuff which then causes them to have illnesses which are then being treated as various different diseases.  For example allergy.  And I think that what happens is that a patient goes to his physician and goes, “I have got these palpitations of the heart.”

So you go back in a week later, “Well the palpitations are all right, but now I can’t sleep.”  So now he gives you another prescription to help you sleep and so on.

Well really the palpitations of the heart and the diarrhea alternating with constipation and the abdominal pain and the headaches and the emotional disturbances are all due to the fact that oxidative metabolism in the brain is in poor shape.  And when that happens the brain becomes very very hyperirritable.  So that any impulse coming in like a change in the weather or something like that can impact in the brain and fire off an autonomic nervous system response which is exaggerated in volume.  Am I making sense?

SB: Absolutely.  This is a beautiful – well not a beautiful – a portrait of disease.  So it makes complete sense.  Let’s say there is a kid that is brought into see a practitioner.  I understand you are retired I believe?

DL: Yes.

SB: But when somebody comes in what sort of labs would you think that should be ordered?  When a child comes in and they have autism, what are labs that should be ordered?

DL: Well I think the standard labs you do, but it is the interpretation that matters.  I mean the interpretation of laboratory markers is very misaligned.  For example if I see a cholesterol that is too low, I think that is more dangerous than a cholesterol that is too high, and yet we are being told constantly by physicians get your cholesterol down its dangerous.  Let me tell you a story just for an example.  I knew the laboratory director at the Cleveland Clinic and he told me that patients would come in for surgery and their cholesterol would be normal before surgery, or maybe high, or whatever, and after surgery and they would get complications and they would do repeated lab studies and the cholesterol would go down down down down down, and they would die.  So what was really going was that the stress of the surgery caused them to use their steroid hormones released by the adrenal gland and the adrenal gland requires cholesterol in order to make the steroid hormones.  Because their energy level – their cellular energy level was inefficient – they couldn’t make the cholesterol to make the steroid hormones.  So as the cholesterol ran low, the steroid hormones could no longer be made, and that is when they died.  It is a different way of looking at it.  You have to think in terms of energy metabolism being the background of health and consequently its loss is the cause of disease.

SB: I mean given your views that are well summarized there, I’m almost afraid to ask but I’ll ask anyways, are there any pharmaceuticals that you feel are useful in treatment of autism?

DL: No.  I never used any pharmaceuticals.  They are unpredictable.  They often have paradoxical effects, and you just can’t tell what is going to happen with a pharmaceutical.

SB: OK.  In terms of autism research there does seem to be a lot of exciting autism research going on.  Is there anybody extending the research you have done on thiamine in autism?

DL: Not that I know of.  No.

SB: Well that’s disappointing.  Of course that is not your fault.

DL: I tried to get… Did you ever go to Defeat Autism Now – DAN?

SB: I’m familiar with the group.  I’m very familiar with the group.

DL: Well I tried to present this to their group, and I don’t think they even listened.

SB: I’m very sorry to hear that.  Are there other areas of autism research that you find are promising?  Interesting?

DL: Well not really because I’m out of the swim now, but I do think that we have to understand that autism is not a psychological disease.  It is a biochemical disease, and it is fundamentally due to mitochondrial dysfunction.  I think that what we are looking at is genetic risk – not genetically determined disease – but genetic risk.  And I think that these minor genetic risks can be made to initiate disease when there is a stress factor imposed on the individual who has perhaps marginal malnutrition.

Let me illustrate that in a case that I had many years ago.  A six year old boy had a head injury.  His skull was fractured.  When he went back to school the school nurse said, “I want you to come down and get your eyes tested every two weeks because” – open quotes “people go blind with this kind of injury” – closed quotes.  Where she got that information I don’t know, but it happened to be proverbial, because three months later there was a major change in his vision, and she referred him to an ophthalmologist who found cataracts in both eyes.  Well the ophthalmologist knew of my interest in metabolism, and there is a disease which occurs in children called galactosemia.  And galactose is a sugar that is formed from lactose, and the galactose then has to be broken down to glucose, and it is under genetic control, and if you don’t have the gene then you accumulate galactose, and it causes cataracts in children.  So the question was does he have galactosemia?  He only had actually one allele.  This is a recessive gene, and he only had one of them.  So he was a carrier of the gene. And so I sat down with mom to find out what his diet was like, and she had been giving him lots and lots of milk to drink as a health drink.  That is quite common with many people pushing milk as a health drink.  Now the milk contains lactose which was then converted to galactose and overwhelmed his single gene.  So he got the cataracts.  Now the point is this, if he had never had the head injury, he wouldn’t have had the cataracts.  If he hadn’t had the milk, he probably wouldn’t have had cataracts, and if he hadn’t had the carrier gene, he definitely wouldn’t have got cataracts.  All three had to be represented.

SB: That is very well said.  So we have many different risk factors and they can work in combinations.

DL: Yes.

SB: It’s very well said.  Dr. Lonsdale is there anything else you would like our audience to know?

DL: Well I think anybody with a child with autism has to think in terms of biological phenomena.  It has got nothing to do with Dr. Freud.  It has got nothing to do with traditional psychology.  It is very much of a biochemical disorder, and I think that ADD and ADHD and O[C]D and all of these conditions that we are seeing in children today are really nothing more than variations on a symphonic theme.  Even obesity is related to dysfunction of the brain and that is published material too. We are damaging ourselves wholesale in this country and in Western Civilization in general.  That is the take home message I think.

SB: Thank you for that sobering point and thank you for taking the time with us.  Thank you for being with us on Autism Research Connections, Dr. Lonsdale.

DL: Well thank you for asking me.  I’ve enjoyed being with you.

Autism Research Connections #2: A Conversation with Dr. Robert L. Hendren

Dr. Robert L. Hendren was interviewed by Seth Bittker on March 26, 2016 on autism, methyl-B12, Omega-3s, Vitamin D, over-the-counter therapies, pharmacological therapies, and the shift in the treatment paradigm for autism.  The first part of the interview focuses on Dr. Hendren’s recent methyl-B12 double blinded trial for autism. © 2016

Robert Hendren.jpg

A podcast of the interview is available here.  A transcript of the interview appears below.

SB: This is Autism Connections #2.  It is March 26, 2016.  I am Seth Bittker, your host.  Our guest today is Dr. Robert L. Hendren.  Dr. Hendren is the Director of Child and Adolescent Psychiatry unit at University of California San Francisco.

In addition to autism, Dr. Hendren specializes in the diagnosis and treatment of a number of other neurodevelopmental disorders such as pervasive developmental disorder, bipolar, schizophrenia spectrum and impulse control disorders. In his research, Dr. Hendren studies pharmacology and nutrition in treatment of autism.  He has been involved in a number of placebo controlled trials in autism including those on omega-3 supplementation.  He has also published reviews on biomarkers for autism and complementary and alternative treatments for autism.  In addition Dr. Hendren is the lead author of a remarkable paper that was published just last month – that is February 2016 – in the Journal of Child and Adolescent Psychopharmacology.  The paper is “Randomized, Placebo-Controlled Trial of Methyl B12 for Children with Autism”.  Dr. Hendren thank you for being with us on Autism Research Connections.

RH: Thank you for inviting me.  It is a pleasure to talk with you and to talk with your audience about ways that we hope that we can improve childrens’ resilience to help them push back against autism and other neurodevelopmental disorders.

SB: This is great.  I think we are going to learn a lot.  Prior to – or maybe before we get into your methyl-B12 research I just wanted to give our audience some background.  Back in the early 2000s Dr. Jill James and some other researchers noticed that those with autism typically had a higher ratio of S-adenosyl homocysteine to S-adenosyl methionine than controls.  In addition they noticed that those with autism typically had higher ratios of oxidized to reduced glutathione than controls.  These are issues in the methionine cycle and more broadly issues of insufficient methylation.  These researchers hypothesized if they gave their patients betaine, folinic acid, and methyl-B12, this would address these issue.  In fact they noticed when they gave their patients these supplements they seemed to get somewhat better behaviorally and they could measure that the biochemistry became slightly more normal.  This was not a placebo controlled trial though.  In any case they published these results in a remarkable paper in 2004 where Dr. James was the lead author.  Since that time these compounds and especially methyl-B12 have been compounds of interest in autism, and a number of practitioners and parents have been providing methyl-B12 to their patients and children.

If you flash forward to 2010, Dr. Kiah Bertoglio published a paper on a double blinded cross over trial with methyl B12 in autism.  Dr. Hendren, who is our guest today, was a coauthor on this paper. In this trial they did not find efficacy with statistical significance but they did find some interesting things in subgroups.

If we flash forward again to last month, Dr. Hendren’s double blinded trial of methyl-B12, and Dr. Hendren found that methyl-B12 outscored the placebo with statistical significance based on a Clinical Global Impressions Improvement index.  In other words Dr. Hendren established efficacy for methyl-B12 in autism.  He also showed that improvements in this index were correlated to improvements in biochemistry.  So congratulations on this remarkable work Dr. Hendren.  What aspects of it would you like to highlight to our audience?

RH: Well I will tell you a little bit of a story about how I got into that I think says something about what I think is especially interesting.  When I first became the executive director of the MIND Institute at UC Davis parents encouraged me to leave no stone unturned about how kids with autism could be helped and to keep an open mind but to do good science.  So I called together a group of people that were doing kind of alternative or complementary type treatments and asked them what should be the first trial that I do?  What one do they think is most promising and they said methyl-B12.  I have to say I was skeptical at the beginning.  I was mostly used to more traditional medicine and yet thought this was fine.  We would try to see if it made some difference.  I had known and really liked Jill James and so Jill was willing to do the labs, and so we started a trial.  The first trial that you mentioned that Kiah Bertoglio and I did, didn’t show efficacy but we only had thirty kids in it.  I think so many of these nutraceuticals or nutritional type trials are things that don’t show dramatic improvement right away – they show slow steady improvement over time, and its hard then to do a double blind placebo controlled trial in that amount of time because you have to hold everything else constant but we did the thirty kids and we found a subgroup that seems to be helpful and we applied to Autism Speaks for funding and they funded us to do fifty.  So since then I have moved to the University of California San Francisco and we did the trial with fifty kids and we found significance.  So I think the biggest difference was that we had twenty more kids.  We saw a trend towards improvement in the first trial but we needed more in order to reach that significance, and I think it says in one way that in the first trial we saw improvement in oxidative stress biomarkers and the second one it was methylation biomarkers and those are related as you pointed out, but it says that this is a way to approach autism that will hopefully make the body healthier, make it more resilient, take away some of the products that might be – the metabolic products that might be causing difficulties and methyl-B12 is one that does that and I think there are others that as we get more sophisticated and thoughtful in what we are doing that we can find that can help kids in this gene environment interaction to push back on the environment and become healthier.

SB: Its outstanding work and we are so glad that you – as you said you were so open minded and you explored this.  One question that I did have on the research: if you dig into your most recent paper – I mean you did definitely get statistically significant improvement the Clinical Global Impressions Improvement index and I believe that you had set that as your primary… reference…

RH: Endpoint.

SB: Endpoint.  Primary endpoint.  Thank you, but then there were some other measures such as an Aberrant Behavior Checklist and the Social Responsiveness Scale where it was more of a mixed picture and I wondered if you could comment on that?

RH: That’s a great question and its one that people have brought up in our study.  The CGI – the Clinical Global Impressions scale is based on usually a physician but a clinician gathering all the information they can.  Seeing the child – talking to the parents – and then as a blinded rater – someone that doesn’t know whether they are on active or a placebo – giving an impression of improvement or worsening in their condition.  It is a general impression and isn’t based on particular items in a scale.  It in most studies has been shown to be in most studies the single best predictor – that kind of – somehow the physician’s impression and feel of the kid or the person in the study is a better predictor of someone improving than any kind of blood or rating scale measure that we can use.  So in this trial the two [other] measures we used were the Aberrant Behavior Checklist and the Social Communication Questionnaire and neither of those seemed to capture whatever the improvement was in these kids, but the impression – and I was the one that did all the ratings – was that the child had improved, and I would say the parents had that impression too.  They could get this feel of the kid that doesn’t fully get captured on a rating scale, and there were things on the blood test that showed changes, but I think it is more as they say in advertisements all the time, “Je ne sais quoi.”  There is something that is there.  I guess the thing that impressed me was that when I saw these kids improve it was like a veil had been lifted a little or their eyes seemed cleaner or clearer or they seemed more connected and that is something that doesn’t get captured well on a ratings scale other than the CGI.

SB: Very interesting.  Very interesting, and you must be a great observer.  Did you see cases where you would see a sudden improvement and then a drop off or it wasn’t like well you could see that given when you were doing these two before and end – after points?

RH: I think when we saw somebody seem to improve and then drop off – we didn’t know whether they were on active or placebo – but I think that happens more with the placebo group.  I didn’t find any kids that were as we look back over the records that we then learned were on methyl-B12 and they showed an initial improvement and then it dropped off.  A real rapid response too is often something that is associated with a placebo response. The methyl-B12 -sometimes people noted improvements within a few days, but it wasn’t usually something you would see the next day.  That was more because the way we were giving this was with subcutaneous injections that you know it’s pretty dramatic for the parents to do that, and I think they really hoped and wanted to think that their child was doing better.

SB: That makes total sense.  So I understand research is one thing and clinical practice is another.  What do you think the take away is from this research for physicians who are out there that have autism patients?

RH: Well I think our study needs to be replicated before we could say that this would be a general recommendation that people ought to put into their practice on a regular basis.  I think there are a lot of physicians I know who are comfortable making it part of their practice.  Then I would say it is important to make sure that you find a good lab that makes your syringes and makes up the methyl-B12.  Our pharmacy at first UC Davis and then at UCSF really wanted to be sure that the lab was making the mixture in a consistent way and that it was sterile and the first lab that we found they didn’t approve and then we found one that they did.  So I think that part’s important – finding a good lab if someone feels comfortable doing it.  When people finished our trial in order for them to continue to get the methyl-B12, they went back to their primary care physicians and I sent them our protocol and sent them our first paper and most of them were willing to prescribe it.  I think as a kind of general thing for people that don’t have any familiarity with it, it might be something that they hesitate using regularly, but I hope that we can do another trial, or get funded to do another trial, or somebody that else does another trial that shows benefit, and then it becomes something that is used regularly.  I also think it may help some kids and not others.  So I think there needs to be some abnormality in oxidative stress or something along in that pathway and if we can find a better biomarker of that that was easy to use, it would help us know to target those kids.

SB: That brings up a great point.  I mean your paper it seemed to highlight that those that had high levels of S-adenosyl homocysteine or at least the ratio of S-adenosyl homocysteine to S-adenosyl methionine was high – I believe it indicated – or maybe it was the same ratio but for the glutathione – the reduced to oxidized glutathione actually improved – I think that was what is was.  Should people be looking at some of these markers and if a physician is considering using this and they look at some of the markers and then they make a judgement based on the markers as to whether it makes sense to use it or this is just too preliminary?

RH: I think if someone’s knowledgeable in the area they might find it helpful.  Jill and I talked a lot about – you know homocysteine is not that hard to get.  People could look at it and say is that a marker, but I don’t think she found it in all of her patients.  But it kind of makes sense that homocysteine could be. It is a marker that is used in other disorders of oxidative stress.  Maybe that one would be helpful.  I think we need more study to really be able to say what biomarker would be most useful.

SB: OK.  Very fair.  I know you have also done some trials of omega-3s in autism.  What can you tell us about omega-3 supplementation in autism?

RH: It makes a slow steady difference.  Our study again was limited by the length of time that we could do it.  I think we have done three now.  They have all shown trends toward significance, but they never fully reach significance in part because we just didn’t have long enough that we could keep using them.  I think it makes sense that Omega-3s and they have been helpful in other disorders as well in other disorders ADHD and bipolar disorder and even schizophrenia.  It helps with neuron growth and if that’s the case then it seems like it would be good for kids with autism as well.  There was one positive study in autism done by Amminger, and I think based on all of that there is enough evidence to say that we routinely ought to give kids with autism Omega-3s usually around a gram.  Some of the studies used more than a gram, but when you just go to the health food store and buy a bottle of Omega-3 or fish oil, you will find that it has a lot less than a gram, so you need to find either a better concentration or give multiple pills.

SB: That makes a lot of sense.  So again would you look at some sort say fatty acid panel, or you would say this kid has autism, I think we should do a trial of Omega-3s?

RH: For the kids that I see in the clinic, I suggest to the parents that they give them a gram of Omega-3s, and I don’t base that on any laboratory tests.

SB: Interesting.  That makes a lot of sense.  If it works, it works.

RH: Yeah and there is no real harm in it and it seems like people are using it for lots of things.  So might as well use it this way.

SB: That makes a lot of sense.  I also understand that you have done some work around oral vitamin D supplementation in autism.  Have you found anything there you can tell us about?

RH: We had a hard time enrolling in that trial.  So many parents felt they don’t want to come into a double blind trial when they can just go out and buy it.  We said that we are using really high doses and we are monitoring it carefully.  So this might be a reason to come in.  We had about ten people sign up.  A number actually dropped out.  We had four that actually completed and we are just writing that up.  It is not going to be anything that is profound other than saying that people could tolerate very high doses and there was a slight trend in this small number of cases to looking towards improvement.  The person that has really been a great proponent of that is Dr. John Cannell at the Vitamin D Council.  You can look at his website and he has lots of anecdotes and testimony and case studies of vitamin D benefiting kids with autism.  There have been studies indicating that about 30% of American kids in general have low vitamin D and about 50% of kids on the spectrum have low vitamin D.  So I think all of that makes it another one of those things that I routinely suggest to parents that they give their kids a couple thousand IU of vitamin D3.  Sometimes I get a vitamin D level and I think it is the right thing to do, but sometimes if I’m not drawing blood for anything else I might just suggest that they go ahead with vitamin D.

SB: Thank you for that summary and the results you have gotten so far.  I would like to ask a few more questions or at least one more question about vitamin D, but before I do so I should acknowledge to you as well as my audience that I am a little bit of skeptic on the practice of providing large doses of vitamin D to those with autism, and I have some biases on this issue and my questions probably reflect that.  That being said when you give the large doses of vitamin D, do you see negative reactions or in general it’s positive or neutral?

RH: It is generally positive or neutral.  I think what you are referring to and there are a few thoughtful papers suggesting that perhaps really large doses of vitamin D could make autism worse or even lead to autism.  That is an interesting theory, and it certainly has made me cautious about supplementing at really high doses for long periods of time.  I worry too that if parents are [giving] vitamin D that has A with it – which often comes in fish oil or others – that you can get toxic on vitamin A since it is a fat soluble vitamin.  I have seen one kid that really kind of wrecked his liver and got his abdomen all swollen up with too much.  It wasn’t my patient, but I saw him in the hospital.

SB: Oh my goodness.

RH: I think people have to be cautious especially with the fat soluble vitamins, and saying well if a little bit is good, then maybe I can give him a whole bunch and that really ought to be good.  I think for the water soluble vitamins it mostly just makes expensive urine to give such high doses, but giving them at a good dose – and I think that the vitamin D idea makes some sense to me and the way that it could push toward health.  In our study we started at 6000 IUs and then gradually went down.  When I see patients in practice I usually just go with 2000.  I know in groups where I have asked physicians how much do they take, a lot of physicians raised their hand at 5000 IU and say that they take that much every day and think that it is helpful.

SB: Wow.  OK.  I know that you have also done a review paper and research on biomarkers in autism.  Do you have a couple that you find to be most useful?

RH: I wish that I did.  In the review there was not a kind of really strong conclusion about things that we might use.  We are just writing up a study that we did of four practices of integrative medicine docs who see kids with autism, and we couldn’t get any agreement about [inaudible] and tried to reach consensus about what labs other than just the routine labs what labs should we be doing.  There are obviously the kind of routine ones like a metabolic panel and a CBC and I think if there is any concern it is worth getting iron.  There have been a number of studies saying that ferritin is sometimes low in these kids.  It’s also good to think about lead screening.  There has been some concern about that as well.  And a D3 level that I mentioned previously.  The ones that are kind of in the maybe they are worth getting kind of category or maybe if you see something of concern.  Some people get serum amino [acids] or urine organic acids and wonder about that and helping to screen for mitochondrial dysfunction.  Some people also routinely get lactate and pyruvate to help with that kind of screening.  The ones that become a little less clear are things like even B12.  In our study the kids mostly had fairly normal B12 levels.  It is mostly that the B12 doesn’t get where it is needed or the methylation isn’t occurring there.  Others sometimes get zinc / copper ratios.  Some look at selenium.  Some look at magnesium, but it’s not part of anything that I think would say that these could be routinely used.  We are doing another study now at a school for autism in the Bay area – the Oak Hill School.  We have implemented an outcome study there, and now we are doing a trial of something that seemed innocuous that the family would feel OK about and that is with this sulforaphane you know the concentrated broccoli sprout extract that had one very positive study from a very good group, and it was thought to help with oxidative stress.  So we are doing a trial at the school where we do before and after urinary metabolomics – a way of seeing clusters of metabolic by-products that could cluster in areas like oxidative stress or mitochondrial function or free fatty acids and so we had twenty of the parents sign up which we think is great.  The kids are giving the urine before and after.  We hope that that could eventually become a kind of biomarker panel that might help us screen and then better target our metabolic interventions to help the kids be more resilient and healthier.  We will see whether that pans out or not.  We have a meeting at a place called the Think Tank that occurs a couple of times a year, and I’m presenting some of this there, and I’m hoping that I can get some consensus at some point from all the different practitioners doing these about what things do they get routinely, but I haven’t found it so far.

SB: That’s great and that is interesting news about your sulforaphane follow-up trial.  I think that the way you did your methyl B12 research, and it sounds like you are doing something similar with your sulforaphane research where you look at behavior before – maybe some metabolic markers before – then you do the trial.  Then afterwards you look at the same thing.  That is very powerful.  I’m looking forward to seeing what you produce on that.

I wondered when a child comes into your clinic and you can see the parents have brought him in.  He or she has the classic signs of autism, are there particular labs you normally order at that point or how do approach these things?

RH: So for the labs that I order, it is usually just those routine ones – the metabolic panel, CBC, D3, and if I think it is indicated the ferritin and those are the only ones I order regularly.  I’m wondering about starting to order more regularly the serum amino acid and urinary organic acid, and lactate and pyruvate.

SB: So metabolic panel.  This may be my ignorance.  What are typical things that are in the metabolic panel?

RH: That is the regular things like sodium, chloride, glucose.  You know whenever you go on your yearly test.  It is just kind of a general view of your body’s health but not metabolic in the way that we have been talking about in this conversation.

SB: I see. Makes sense.  Thank you for clarifying that.  So aside from the supplements you mentioned earlier, are there other over-the-counter supplements you find useful in treating these patients with autism?

RH: Yeah I think melatonin is really helpful.  I think people often give it to their kids for sleep, but it may even boost immune function.  So for parents that are so inclined, I say it seems worth just giving your kid melatonin.  There is concern – a small concern – about giving high doses for long periods of time.  I have never seen that kind of borne out, but I do use melatonin regularly for kids that have sleep difficulties.  If they have a lot of GI disturbance, I suggest probiotics and pancreatic digestive enzymes.  For a while every time I would see a kid that had GI issues, and I would send them to the GI specialist, they would always do that – put them on probiotics and pancreatic digestive enzymes.  And I thought well I could do that first and if that helps then maybe they don’t have to go to the GI specialist.  But we were part of a study done by Curemark of pancreatic digestive enzymes where they used fecal chymotrypsin as a biomarker, and while the results of that trial have not been released, they are coming back and doing it again this time without fecal chymotrypsin biomarker and that was at the recommendation of the FDA.  So it would suggest – I’m just kind of guessing – that it showed some benefit since they are going through this again and the FDA is continuing to monitor that with them.  The probiotic study that was done at Caltech with the mouse model of autism that reversed with probiotics I think has made people think about how that might be helpful even in even more of a routine way than just for those kids that have GI difficulties.  As I had mentioned I do the D3 and the Omega-3.  Those in general are the things that I think there is enough evidence that I use routinely.

SB: That’s terrific background and you have sort of given us a scoop there on these trials of the pancreatic enzymes.  It does seem like there is a lot of interesting research around the microbiome – a lot of indications that the microbiome is involved.  The one trial I saw on probiotics in autism the results were… disappointing.  Do you have any thoughts are we giving the wrong probiotics in general?  Is there an effect there that is not being picked up?  Any thoughts on that?

RH: I think that is a good point.  There are a whole bunch of these little things that they put into the probiotic.  The one in the mouse model that showed benefit was B Fragilis.  I don’t know whether there is anything special about B Fragilis, but it is the one that they used and reversed the mouse model.  I think it is thoughtful kind of way that these things might get formulated and used.  That Curemark study – the thoughtful lady that led it and puts together the things in the digestive enzymes.  They are not just the things you buy off the shelf.  She kind of put it together and then patented it.  I think if we could just find better biomarkers, we could begin to start to personalize the treatment for these kids and find ways to show benefits.  The other thing that happens I think in these studies, is that if we don’t screen with the biomarker, and we take all kids with autism, they may not have an active process going on at the time, and so the intervention that we use doesn’t help, and then we wind up with a negative study and we say well this doesn’t work.  It’s more like saying well – I think we are increasingly referring to the autisms, or you’ve seen one kid with autism, you have seen one kid with autism.  So each child kind of has a unique way that they have autism.  I think if we could do a better job of identifying that, I think we could better target our treatments.

SB: Those are great points.  Dr. Hendren excluding your own research, what other autism research that has been published in the last year or so you thought this is really exciting – this is really important?

RH: Well it’s been a little over a year, but Antonio Hardin’s NAC paper is exciting, and I find a number of kids that I see if they have skin picking or OCD type things or they are irritable, I find that NAC is helpful.  I think there another study that will be out soon that is negative for NAC with autism. So it’s not a panacea or it’s not something that we can say it will do great for everybody.  I think the work that comes out of your first interviewee – Richard Frye’s lab – I think is interesting and promising.  The things that he and Dan Rossignol do together and the reviews and things that are moving things forward like the cerebral folate deficiency.  Also looking at folinic acid.  I think those are interesting thoughtful studies, and I think those groups are moving the field forward.

SB: That’s really interesting.  I appreciate you bringing up NAC.  The Hardin study I think showed improvement in irritability.  I understand that part of that study showed a lot of GI issues.  There are other issues with NAC.  What do you think we are doing when we give NAC to a kid biochemically?  Any thoughts on that?

RH: I think it seems to be helpful for oxidative stress as well.  So I think that’s one of the strong ideas of what the mechanism of its benefit is.

SB: Makes total sense.  We talked about your research, about this research that is going on.  Is there research that has not been done yet that is really important that should be done?  Is there something out there that you think – gosh – people should be doing this now?

RH: Well I think looking for biomarkers and I think doing more with metabolomics.  It may or may not work, but we are excited about trying to do that.  It would help us make better cocktails that would kind of fit with particular kids.  So I think that.  I am not thinking right now of anything that is really promising and someone hasn’t been doing the studies of.  I’m not sure what that would be.  Everybody has their favorite.  Nothing comes to mind.  When I talked about being about the MIND Institute, and I asked the group what things should we study?  What they wanted me to do a study of chelation, and I said no, I’m not going to start there.  That is just too controversial and too difficult to know whether you are doing a good trial.  I have had some parents tell me that it helped them.  I think there is an exciting potential in using transcranial magnetic stimulation – TMS – that maybe is showing some benefits.  I have heard real mixed reviews on stem cells, but I have had some people tell me that they thought their kid got quite a bit better.  So there is some research beginning to look at those things, but I think all of it reflects that we are changing our paradigm.  We are not thinking just as neurotransmitters or brain regions and then trying to use conventional medications like anti-psychotics or SSRIs or other things to help people do better because we are only helping their behavior and we are not fixing the metabolic process, but now I think we are moving in the direction that cancer research is and vascular research and the practice of those that are increasingly looking at metabolic processes and ways that they can target those, and I think we will learn a lot from them, and a lot in that way as we move forward in understanding the metabolic process of autism.

SB: That is terrific, and I realized I wanted to ask you one question that I had really neglected, which is that we have talked a lot about over-the-counter supplements for autism.  We really didn’t talk so far about pharmaceuticals with autism, and I know you are an expert on that too.  I understand that there are a lot of…a lot of… side-effects and issues with some of the traditional anti-psychotic medications and things given with autism.  Are there certain pharmaceuticals that you think – wow – we are just underutilizing this – this should be given more to certain kids with autism?

RH: I think that about some of the nutraceuticals, but I think the one that has been kind of a surprise to me was propranolol.  It has been around for a long time.   I have used it for many many years.  I kind of quit using it because I didn’t think it was really helpful, and then the group at the Thompson Center in Columbia Missouri has had a series of studies showing propranolol benefiting some kids.  I think some of the literature around that suggests that kids that have an autonomic over-reactivity – they get flushed and red and sweaty and there is something in the autonomic nervous system that is driving their meltdown or their falling apart and maybe propranolol works for that.  So I have been using it more and I find a few kids do better.  So I guess it is a little bit underutilized but it is not for everybody.  I think people keep trying to find ways to have anti-psychotic medications have less side effects, but that hasn’t totally worked.  Like Risperdal and Abilify cause a lot of weight gain and they worry about increasing glucose and then the risk of type 2 diabetes.  So Latuda has come along – lurasidone.  It was thought that it didn’t increase glucose.  It didn’t increase blood lipids.  The trial of that for autism was negative.  I still have some kids on it, but I find that other kids get a kind of inner restlessness that makes them uncomfortable.  So while we are moving ahead in some ways, we haven’t really figured out a way to have some of those medications that help.  They don’t help reverse the disease process.  They help for kids that have major behavior problems and the parents can’t go out, and they can’t take their kid anyplace, and they are worried about their siblings, and they are thinking of an out-of-home placement, and then this medicine can make a world of difference even though it has a lot of risks.

SB: Thank you for that excellent overview of what is available in term of the pharmaceuticals for autism.  Is there anything else that you would like our audience to know about the research you are doing or other thoughts on autism?

RH: I think the thing that is pushing me – or driving me – or that I find most exciting in this new approach – not necessarily a new approach – but it is a shift in the paradigm is that it is kind of saying is this is whole body disorder – that you can treat the immune system or you can treat the GI tract and it effects the person’s socialization and the way their brain works.  All of these are things that say if we can find ways to help the body to be as resilient as possible, then I think these kids are more able to take advantage of education, speech and language, behavioral programs, interacting with other kids – all of those things – and that is what we are trying to do in this school – the Oak Hill School that I had mentioned.  It is a two part thing: it is getting their bodies as healthy as possible and everything working as well and then having an excellent program for them to be in.  I think that approach is going to move the field forward.

SB: That makes total sense.  Dr. Hendren thank you so much for being with us on Autism Research Connections.

RH: Mr. Bittker it was a pleasure.

Autism Research Connections #1: A Conversation with Dr. Richard E. Frye

Dr. Richard E. Frye was interviewed by Seth Bittker on March 15, 2016 on autism, mitochondrial dysfunction, autoimmunity of the folate receptor, over-the-counter therapies, folinic acid, the microbiome, and other advances in autism research and treatment. © 2016

Dr Frye

A podcast of the interview is available here. A transcript of the interview appears below.

SB: This is Autism Research Connections #1.  This is Seth Bittker, and our guest today is Dr. Richard Frye.  Dr. Frye is Director of Autism Research, Director of the Autism Multispecialty Clinic, Co-Director of the Neurometabolic Clinic, Associate Professor of Pediatrics of Child and Behavioral Neurology at Arkansas Children’s Hospital Research Institute as well as University of Arkansas Medical School.  Dr. Frye has written extensively on mitochondrial dysfunction in autism, autoimmunity of the folate receptor in autism, methylcobalamin and folinic acid in autism, the microbiome, as well as numerous other papers on autism.  Dr. Frye, thank you for being with us today.

RF: Sure it is my pleasure.

SB: Dr. Frye I wonder if you could introduce us to what is mitochondrial dysfunction?

RF: Well we should start out with what is the mitochondria.  So our cells which make up our body have different pieces in them that do different functions, and the mitochondria is the part of the cell that is best known for making all the energy in the cell, and it is a very unique part of the cell.  First of all you don’t just have one mitochondria for every cell.  You can have anywhere from hundreds to tens of thousands per cell in any cell in your body depending on how much energy that cell needs, and what we have found out that sometimes this powerhouse is not working the way it should.  And so you can imagine if you are living in a city and the power and you have a blackout, there are so many things that are going to happen, and things won’t work.  What we find is that mitochondrial problems or what we call mitochondrial dysfunction seems to be pretty prevalent in autism.  Now the idea that the mitochondria doesn’t work and diseases associated with the mitochondria really are pretty new on the playing field of medicine.   Really the first well known or well characterized disorders are really just in the 1980s that described diseases of mitochondrial disease, and for a long time we believed mitochondrial disease occurred when the mitochondria was very dysfunctional, that is just like if you were living in a city and there was a blackout and you had absolutely no power.  So many of the early mitochondrial diseases were described where the mitochondria was not working at all.  So the cells were not working in the body and the body became sick and many of the organs did not work, but what we are starting to find out and we think we see this in autism is that sometimes this powerhouse of the cell, the mitochondria, it’s not that it doesn’t work at all, but it doesn’t work up to the potential that is needed for the body to work optimally.  So you can think of this as if you are kind of in the city you have a brown out where the power goes down a bit, maybe you can’t use your air conditioner, but the lights are still on.  And what we find is that this is analogous to what we see in many children with autism is that this powerhouse of the cell isn’t working up to the capacity that is needed, and this is very important during development – during that time at which we see the onset of autism because the body is growing.  When the body is growing, it needs lots of energy to grow.  It needs lots of building blocks, and if it doesn’t have the resources that it needs, it is going to stall, and we believe that is what happens is that brain development stalls and doesn’t complete through the steps that it needs to fully develop and we end up with the characteristics of autism.  So that is just in a nutshell you know mitochondria and why we think they are important in autism.

SB: That is a wonderful – or very revealing picture there that you have sketched for us.  So when you suspect mitochondrial dysfunction what are the lab tests that you would typically order?

RF: Well it varies, and one of the problems that we have with mitochondrial disease and dysfunction again is it’s really the new player on the block.  So the method for diagnosing it is still not all that well developed.  So usually what we do is some of the standard labs which would be fasting labs first thing in the morning before breakfast where we look at biomarkers such as lactate or pyruvate.   We also look at amino acids and their levels in the body because many times if the mitochondria is not working there will be certain patterns of amino acid abnormalities.  We also look at fatty acid metabolism because fatty acids are also processed and burned by the mitochondria for fuel, and we look at many of these tests and we kind of put them together to see if there is a sign of mitochondrial dysfunction, and then if there is there are many other tests that really have to be done to confirm whether mitochondrial dysfunction is there or not, what is causing it, and to what extent it might be interfering with the life of the child.

SB: That is very well said.  So if you find mitochondrial dysfunction, it seems like you might provide maybe carnitine.  Are there other supplements you would consider providing  or how do you treat it?  How do you typically think about treating it?

RF: Sure.  So every child is different and different doctors that treat mitochondrial disease treat them slightly differently.  A child with autism and mitochondrial dysfunction is unique in many ways.  First of all we know that many children with autism also have what we call redox abnormalities.   That is the system that balances oxidative stress – that is toxic radicals in the body – isn’t working as well as it should be in many children with autism.  We know that when the mitochondria is not working the same thing happens.  We have this elevated level of oxidative stress.  So one of the first things that we do is try to improve the body’s  ability to make the antioxidant glutathione by giving B12 and folinic acid to help the systems that naturally make the natural antioxidants of the body – most notably glutathione.

The next thing we do to treat mitochondrial dysfunction that I find is very useful is something called CoQ10, and many people see CoQ10 for heart health – for general health as a vitamin that they can take, and CoQ10 is known as an antioxidant – is advertised as an antioxidant, but what a lot of people don’t know is that CoQ10 is actually part of the machinery of the mitochondria.  That is the final common pathway of the mitochondria that make energy called the electron transport chain – the different parts of that chain – the so called complexes – actually transfer some of their energy using CoQ10.  So CoQ10 is almost like the wires that connects some of the parts of the electron transport chain.  When there is high levels of oxidative stress, it is used as an antioxidant and can be used up, and therefore it is not available to the mitochondria for the mitochondria to function.  So that tends to be my second thing is to use CoQ10 which can act as both an antioxidant and help the mitochondria work.  Other treatments as you noted are carnitine is extremely useful.  Carnitine is important for fatty acid metabolism.  It is also important for actually removing toxic organic acids from the body and cells and eliminating them from the body.   So carnitine is definitely one of the go to vitamins.  Also many of the B-vitamins are very important.  So we supplement B-vitamins.

Another important supplement would be creatine.  Creatine is creatine monohydrate which a lot of the body builders use.  They use it because what it does is it creates phosphocreatine in the muscles which store energy.  So when the mitochondria isn’t working, what it can do is it can actually store some of this energy and you store your energy in phosphocreatine.  So creatine can go into the cells and particularly the muscles and actually store energy and so it can compensate for certain mitochondrial dysfunction.

The other thing that we find really is helpful for a lot of children with mitochondrial dysfunction is high doses of folinic acid because as you had mentioned in the beginning one of the things we do a lot of research on and have done clinical trials is central folate deficiencies – that is deficiencies in folate that seem to affect the brain but not the body, and central folate deficiencies are very important because of course folate is one of the essential vitamins.  It is important for many of the essential functions of the cell.  It ends up that your brain is protected by the blood brain barrier, and in order for many things to get across this blood brain barrier they need a special carrier to take them, and we find that this carrier that carries folate into the brain sometimes can be dysfunctional.  We find that there are these folate autoantibodies that many children with autism have that block the ability of this carrier to work, but we also know this carrier needs lots of energy, and any part of the body that needs lots of energy is going to be affected if the mitochondria is not working.  And so we find that many children have problems with cerebral folate deficiency or insufficiency and respond to high dose folinic acid.  So in a nutshell those are some of the treatments that we usually start out and use for children with mitochondrial disorders or mitochondrial dysfunction.

SB: Thank you for sketching that out, and I believe I recall that you have done some – I believe it was a double blinded trial with folinic – is that accurate or not the case?

RF: Yes.  That is definitely accurate.  You know our first study  was an open label study that was actually a large case series that I did with Dr. Rossignol where we showed that kids with the folate autoantibody seemed to respond very well to high doses of folinic acid with particularly increases in language.  So we followed that up recently – we completed a double-blinded placebo controlled study where half the children are getting placebo, half getting folinic acid and neither their doctors nor the patients know, and what we actually showed in that trial is that the kids that got folinic acid had over a three month period – demonstrated a very significant increase in language as compared to the placebo.  We also did some analysis to look to see if those kids that responded had the folate autoantibody and we found that those kiddos with autism and the folate autoantibody had a particularly large response to folinic acid.  So we think that not only is it an important treatment just in general for children with autism but particularly those that have this autoantibody that block the ability of folate to cross the blood brain barrier.

SB: Congratulations on that work.  I’m curious about the choice of folinic versus methylfolate or some other form of folate.  Do you have any thoughts on whether folinic is maybe best in some sense for a larger number of children or whether there are other forms that would be useful?

RF: So theoretically the reason why we do folinic acid is that it’s a reduced form of folate.  So one of the major health advances we have made in the United States and other developed countries is fortifying our food with folic acid but one thing a lot of people don’t realize about folic acid is that folic acid is the oxidized form of folate and because it is the oxidized form it’s not readily available for your body to use.  So your body actually has to go through a number of steps to make it activated and reduce it and we know that some people particularly people with autism have a problem with the enzyme that actually reduces it.  So many times just taking folic acid will not be sufficient especially in those that have certain polymorphism in genes that are responsible for reducing folic acid.  So folinic acid is a reduced form of folate.  So it more readily goes into the biochemical pathways that are important for the body and it’s readily used by the body.  The block that we talked about in this transporter can be circumvented by using another transporter that transports folate into the brain and it’s called the reduced folate carrier and that carrier only can transport reduced folates.  It can’t transport oxidized folates.  So we use a form of reduced folate – that is folinic acid.  And we use it at high doses for a number of reasons.  One because the reduced folate carrier has a lower affinity for folate than the folate receptor alpha which is the one that is blocked.  So you need higher doses because of decreased affinity, and in addition the levels of folate in the brain are actually higher than the levels in the blood.  So that is why the folate receptor alpha –the one that is blocked – is so important under normal circumstances and why it takes energy because it actually transports folate from a lower concentration in the blood to a higher concentration in the brain.  The reduced folate carrier is not so good at doing that.  So you have to get higher levels of folate in the blood – reduced folates in the blood for it to get into the brain at adequate levels.  So why do we folinic acid or also known as leucovorin calcium?  So there are a number of reasons.  The most important one is that the studies that have looked at this disorder of problems with central folate have used folinic acid and they have actually measured the levels of folate in the brain before and after folinic acid treatment.  So we know that folinic acid works and we also know the approximate doses that it takes for it to work.  So that is important.  Now theoretically other reduced folates should work also.  So other methylfolates and such should work but we are not so confident about what the doses are that need to be used.  So if you use another type of reduced folate and it works you know that might be great, but if things aren’t working out the way that you want, you are kind of lost as far as knowing if your dosing is wrong or something else is not correct.  So personally I like to stay with something that I know more about.  The other thing about folinic acid also known as leucovorin calcium is that it has been used in oncology as rescue therapy to reduce side effects from many chemotherapy agents.  So we know the safety profile of it also.  In general folates are safe, but it gives you a little more confidence when you know that a compound has been used for many decades and people have had experience with it and there is no major side effects from it.

SB: Wow.  Thank you for that explanation.   So earlier we had spoken a little bit about mitochondrial dysfunction.  We also have the autoimmunity of the cerebral folate receptor.  When you are seeing these thing, do you have any hypothesis on is there an order in which this happens?  So in other words, do you think that you initially have some sort of mitochondrial dysfunction, then that leads to the autoimmunity or is it the autoimmunity that then leads to mitochondrial dysfunction?

RF: It is really a great question.  It is one thing that we are finding out is that a lot of kids with signs of autoimmunity with autoantibodies – not only the folate receptor autoantibodies – but other autoantibodies to the brain seem to also have mitochondrial dysfunction.  For some reason they seem to go hand in hand, and it is not exactly clear why that is, or which came first – you know which was the chicken or which was the egg?  And I think it is an important connection.  We see that the two seem to go together.  It maybe that it starts out with one or the other and it creates a viscous spiral that worsens both of them.  So for example we know that the immune system and immune cells take a tremendous amount of energy to work.  So if you have mitochondrial dysfunction, the immune cells aren’t going to be working up to their capacity, and they can end up becoming dysfunctional and creating autoantibodies.  So those autoantibodies cause inflammation which will then cause oxidative stress and more mitochondrial dysfunction.  So it can be a viscous spiral.

In the same way it may be an immune trigger.  It may be that you have a slightly weak mitochondria and some type of inflammatory event happens which just pushes things over the edge where the mitochondria can’t handle it and they become dysfunctional.  We are not sure, and it may be that there are many ways to get to the same place, and that is something that really future research has to look.   You know I don’t want to [inaudible].

SB: Very interesting.  Very interesting.  So if you also have the microbiome coming into play –and I know you have written extensively on this as well – do you see that as further something that is downstream, upstream, or again we are not really sure how exactly the order works but it something that becomes dysfunctional?

RF: Yeah.   The microbiome is just really fascinating, and the research that is coming out on the microbiome is extremely fascinating.  We know that the microbiome especially early on in life – in the first three years of life – but probably in the first month – there are critical times when the microbiome is developing and influencing the immune system and the metabolic systems.  We know that it helps program the immune system.  We know that it can also interfere with metabolic systems also.  You know there are some compelling studies that suggest that these changes in the microbiome seem to precede the onset of autism sometime suggesting that there is potentially some causative effects of the microbiome, although you can’t rule out that it is an epiphenomenon to something else.  But there is lots of data that suggests that the microbiome has some type of real key relationship to the development of autism.   You know the fact that children with autism tend to have more antibiotic use early on, which can change the microbiome.  They tend to have other changes in diet – GI problems – that may precede the onset of autism.  So I think it is really interesting, and I’m very interested to see what we find out in the near future.  You know there are great studies now that are coming out not so much on autism but in other childhood diseases that show that really modifying the microbiome particularly within in the first month of life is preventative of many particularly immune diseases and allergic diseases.  So there is a lot of data that seems to be pointing that this is really an important component and something that has really been underappreciated for a long period of time, and given the fact that it can modulate metabolic systems and can modulate the immune system, I think it is most likely a very important component to the etiology of autism, and so hopefully studies within the next five – ten years will give us those answers and will help us understand how we can change our lifestyles and our diets and such to manipulate the microbiome to make us healthier.

SB: Very interesting.  Very interesting.   So I know that in autism research recently there has been a big focus on biomarkers for autism and some of the work has gone into things like eye tracking and such.  Do you have any candidate biomarkers you would be willing to tell us about?

RF: Yeah.  I mean we work on more metabolic biomarkers – things like the folate receptor alpha.  We find that many parent and particularly mothers of children with autism that have the folate autoantibody, their mothers also have it too.  We know the same thing with many of the changes in redox metabolism that we see in children with autism, we also see them in moms.  Right now we are also developing some markers of mitochondrial function and dysfunction that seems to be very unique to autism.  We are developing those to help us not only understand how mitochondria are working or not working, but also our goal is to be able to use these tests early on in life and even prenatally to initiate treatment before autism onset occurs.

SB: That is brilliant – a great future.   So I know earlier you had spoken about some of the over-the-counter therapeutics that you sometimes use.  Are there any other over-the-counter therapeutics that you feel are just underutilized in autism that you would like to mention to our audience?

RF: Yeah.   I think there is a lot of potential for a lot of different over-the-counter and other types of treatments.  The issue with a lot of them is that they really haven’t been studied in well-controlled studies to actually tell us how helpful they would be, but we know for example one of the things that has gone through some double-blinded placebo controlled studies and seems to be very helpful  for irritability is n-acetyl-cysteine which helps deplete[d] glutathione and decrease glutamate, the major excitatory neurotransmitter and something that can be purchased from suppliers and can be very helpful.  As you mentioned carnitine.  One of the other things that I use a lot of times is the Autism Nutritional Research Center’s multivitamin that Jim Adams put together, which he is showing two double-blinded placebo controlled studies  to be helpful.  Of course you know it is important when we talk about over-the-counter supplements that people understand that it is very important to get these supplements from reputable laboratories and producers as they can vary in quality quite a bit.  It is just very important that you get good quality supplements, because if they don’t work you don’t know if it is because the supplement was poor quality and if you have a poor quality supplement , it can have things in it that you didn’t expect and that is not always good.  So I think there is a lot of potential for certain over-the-counter supplements that can be useful.  I think every child is different, and it is important to understand that there is not one size fits all.  So I really encourage parents to see a physician that is knowledgeable in the use of both over-the-counter and non-over-the-counter treatments to see what would be best for a child, and really not do things so much themselves, but make sure that they have an experienced [physician] that guides them.

SB: That makes a lot of sense.  You mentioned also about the importance of using trustworthy sources.  Are there a couple of brands or names in the industry that you feel are unusually trustworthy with respect to the contents of their supplements?

RF: Yeah.  There are certain laboratories – of course the USP seal is an independent laboratory that tests supplements that can verify what is in the supplements is good, but there [are] other different pharmacies and suppliers that I think are high quality.  So I think that it is important for parents to talk to their support groups – other parents – to see what their experience is.

SB: Excellent.  Also you mentioned the potential importance of prescription pharmaceuticals in the treatment of autism.  What are some of the prescription pharmaceuticals that in your view are just underutilized in autism treatment?

RF: Yeah.  I mean I think that many of the prescription pharmaceuticals still need to really be tested on kids with autism a little bit better.   I think one of the problems is that really the only big studies that have been done have been done with antipsychotics which have significant side effects that should not be ignored.  So personally when I see a patient I try to find out what symptoms are the key symptoms to treat and then direct treatment toward that whether it be prescription or non-prescription.  I tend to try to avoid antipsychotic medications, but the studies on other prescription medications are very variable, and so I would say that it is important for anybody to see a doctor with knowledge of what has been really researched – what hasn’t been – what the best evidence is so that they get the best care for their child.   And there are a variety of different medicines that can help, and I think that it needs to be kind of customized on a case by case basis.

SB: Makes total sense.  Dr. Frye there is a lot of great autism research going on.  Is there any recently that you have thought my goodness, this is really interesting, it is something we should be watching?

RF: Oh I think there is a lot.  I think as you mentioned the microbiome research is incredible.  I think there is a lot of really great research on the immune system.  We are really finding out how the immune system and many autoantibodies seem to play into possibly not only causing autism but increasing the disability and symptoms in children of autism.  I think those are two really important areas.  Of course I am very excited about the research we have going on on metabolic systems.  I’ll tell you what I like about metabolic systems, and why I think it is so important to research and look at is that these are systems that we can change.  Although you know it’s exciting to look at the immune system, the immune system does not have any really great therapies that are extremely safe.  So until we find really good therapies, I think there is going to be limitations to what we can do to modulate the immune system.   Whereas things like the metabolic systems and the microbiome are two things that we can change pretty easily and safely and maybe have a big impact.

SB: That sounds very exciting.  With respect to the metabolic system research that you are doing – obviously don’t tell us anything you don’t want us to know – but do you feel comfortable telling us anything about that research in terms of what you are doing – what you are looking at?

RF: Well what we are looking at is how these systems may be dysfunctional and why they are and what can be done about it and what type of things can be done to support them.  What we are finding as I mentioned previously is that mitochondrial dysfunction seems to be very much linked with oxidative stress and immune dysfunction, and we are starting to learn how these are all connected and what things we can do to actually either alter these systems to normalize them or to make sure that they are working optimally so that the child and the body can work optimally.

SB: Sound great.  Dr. Frye is there anything else that you would like us to know in terms of things we should be keeping an eye on with respect to autism research?

RF: Well I think that it is very important that we focus – I think as you mentioned before that really I think the focus so far for a lot of research has not been on treatment.  I think there is a lot of treatment on the horizon – either novel treatments that we haven’t used yet or treatments that we believe that work that just have to be tested to make sure that we know how to use them.  I think that understanding what is causing autism and how metabolic and immune systems are dysfunctional are very important areas of research that we have to concentrate on, and of course really the missing piece I think is that we have not been concentrating on how environmental influences may be triggering systems to go awry and cause autism through the microbiome or other systems and what we can do to prevent that from occurring.

SB: Brilliant.  It makes a lot of sense.  So Dr. Frye, thank you so much for being with us.  This has been a truly illuminating conversation.

RF: Sure.  It’s my pleasure.

What is Autism Research Connections about?

Seth Bittker

My name is Seth Bittker.  Research shows that autism is usually fundamentally rooted in biochemistry.  Therefore, therapeutics for autism should be focused on ameliorating the negative aspects of that biochemistry.  So in my view autism biochemistry and relatedly therapeutics targeted to that biochemistry are the two areas of autism research which are likely to have the greatest impact on the lives of those with autism and their families.

Autism Research Connections is an interview series which focuses on research and researchers who are working in these promising areas to highlight their challenges, their thinking, and their successes.  I hope you find the interviews as compelling as I do.