Carnitine supplementation and autism: "side-effects and behavioral outcomes"

'Favourable outcomes' is a term mentioned in amongst the various findings reported by Robin Goin-Kochel and colleagues [1] following their examination of "dose compliance, attrition, and potential side effects of short-term, high-dose carnitine supplementation" in a small group of boys diagnosed with an autism spectrum disorder (ASD). Understanding that the Goin-Kochel study was primarily directed at looking at safety, on the basis of 'high-dose' carnitine supplementation, it appears that an elevation of plasma carnitine and related metabolites was not the only effect noted in their small cohort (N=10).

Tracking back slightly, carnitine is an important compound. Not quite an amino acid, carnitine plays an important role in energy production; as per use of the word 'mitochondria' and it's transporting duties of long-chain fatty acids to the cell powerhouse for energy conversion. You probably won't be surprised to hear that carnitine has a *connection* to some autism (see here and see here). Indeed, Goin-Kochel et al mention the findings reported by Patrician Celestino-Soper and colleagues [2] and their identification of a genetic issue that impacts on 'carnitine biosynthesis' in some people diagnosed with ASD. At least one of the authors on the Goin-Kochel paper has some pretty important knowledge about that finding of trimethyllysine hydroxylase epsilon (TMLHE) gene issues in the context of autism...

Alongside looking for reports of any side-effects from the use of carnitine - "oral suspension or tablets of levocarnitine in 3 divided doses, starting at 200 mg/kg/day and increasing to 400 mg/kg/day, with a maximum daily dose of 6 g" - various behavioural schedules were included in the study protocol. Some were objective measures of autism symptomatology; others were parent-report measures. The use of the Clinical Global Impression Scale (CGIS) also provided a helpful 'clinicians' overview' of before and after supplementation in this open-trial.

Results: a few side-effects coinciding with carnitine use were reported. These included: "heavy odor (4 parents), diarrhea (4 parents), and sporadic vomiting (1 parent)." Such reported side-effects meant that three children remained at the lower dose of carnitine over the experimental period (8 weeks).

Alongside, a few other 'favourable outcomes' were also reported: "calmer behavior (2 parents), more energy (2 parents), increased prosocial behaviors (4 parents), greater awareness (2 parents), better eye contact (2 parents), and improved language skills (2 parents)." These parental reports were accompanied by some 'changes' noted on the various schedules included in the study protocol, including those CGIS ratings. The authors used the study results produced by Geier and colleagues [3] as their comparator; highlighting how both studies had picked up "improvements in overall ASD symptoms... and some language ratings." Importantly too, Goin-Kochel et al talk about how none of their cohort were rated as "worse at post treatment."

Where next? More research please. Bigger participant numbers, more methodologically sound study designs and perhaps also, investigation of the potential pros-and-cons of carnitine supplementation over a longer period of time. By all means keep an eye on those side-effects and perhaps look to the biochemistry as to why such side-effects might appear; indeed look to the biochemistry for potential best-responders to this type of intervention too ("One child had documented TMLHE deficiency and 3 had low carnitine levels" in the Goin-Kochel cohort). But more study is definitely indicated...

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[1] Goin-Kochel RP. et al. Side Effects and Behavioral Outcomes Following High-Dose Carnitine Supplementation Among Young Males With Autism Spectrum Disorder: A Pilot Study. Global Pediatric Health. 2019; 6: 1-8.

[2] Celestino-Soper PB. et al. A common X-linked inborn error of carnitine biosynthesis may be a risk factor for nondysmorphic autism. Proc Natl Acad Sci U S A. 2012 May 22;109(21):7974-81.

[3] Geier DA. et al. A prospective double-blind, randomized clinical trial of levocarnitine to treat autism spectrum disorders. Med Sci Monit. 2011 Jun;17(6):PI15-23.

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"Comprehensive Nutritional and Dietary Intervention for Autism Spectrum Disorder"

It's been a while coming but the paper by Jim Adams and colleagues [1] detailing the effects of a "comprehensive nutritional and dietary intervention for autism spectrum disorder" has finally seen the peer-reviewed light of day. I say 'a while coming' because as per the ClinicalTrials.gov entry for this research (see here), it was seemingly scheduled to start back in 2011 and be completed by 2013, I assume, without taking 5 years to write up and be published. But better late than never I suppose.

Anyhow, the nutritional and dietary intervention scheduled adopted by Adams et al was rather a complicated affair as per the study description. So: "Day 0: Vitamin/Mineral supplementation begins. Day 30: Essential Fatty Acid supplementation begins. Day 60: Epsom salt baths begin. Day 90: Carnitine Supplementation begins. Day 180: Digestive Enzyme supplementation begins. Day 210: Healthy, casein-free, gluten-free diet [HGCSF] begins." Quite a few of those individual intervention elements have been fodder for this blog before (see here and see here for examples); also reflecting other research interests from Adams and colleagues (see here). Talk about a gluten- and casein-free diet is also music to my [research] ears (see here), as is the welcome inclusion of sulfate / sulphate back into autism research proceedings (see here).

Results of that nutritional and dietary schedule are reported for a starting pool of 67 children diagnosed with an autism spectrum disorder (ASD), where 28 participants completed the 'treatment' arm and some 27 participants completed a non-treatment arm (where no new intervention(s) were reported for the 12 months of the study). Additional findings for 50 not-autism controls (I don't like the word 'neurotypical' and its rather sweeping connotations) are also reported. The study duration was a year, and the sorts of measures examined over the course of the intervention were quite comprehensive, covering both behaviour and cognition and also physiological parameters.

Results: it's always refreshing to see a study first and foremost reports any adverse effects based on the tenet 'first, do no harm'. Authors note that: "A few adverse effects were reported for some treatments" and go on to list what happened over the course of each element of the intervention. They talk for example, how: "One parent reported that implementation of the diet [healthy, gluten- and casein-free diet] in a strict manner resulted in increased aggression towards peers, inability to problem solve, and increased spinning behavior, probably due to frustration in regards to removal of favorite foods." Thankfully, most of the adverse effects noted over the study period were relatively minor and certainly not life threatening. Once again, first, do no harm.

With levels of compliance regarding the various study elements also reported as being quite high, the authors report that across the various behavioural assessments - including the CARS, SRS, VABS, and ATEC - significant effects in favour of intervention were found. Based on blinded evaluations using something called the Reynolds Intellectual Assessment Scales (RIAS), authors reported "a significant improvement in nonverbal intellectual ability in the treatment group compared to the non-treatment group." By contrast, blinded use of the gold-standard assessment instrument known as ADOS revealed "no significant change on the ADOS scores for either treatment or non-treatment group." Interestingly, when parents were asked to rate the effectiveness of each part of the intervention, results revealed that: "The highest rated treatments were the vitamin/mineral supplement and the essential fatty acids, followed by the Healthy HGCSF diets, followed by the carnitine, digestive enzymes, and Epsom salt baths."

Adams and colleagues also provide further details on "3 exceptional cases of improvement during the study, all of which occurred in the treatment group." For one participant it appears that the introduction of a carnitine supplement was associated with some quite remarkable improvements in relation to strength and energy levels in particular. The authors note that: "low carnitine seems likely to have contributed to her challenges, and carnitine supplementation seems to have helped." There could be some interesting tie-ups there with regards to previous peer-reviewed results too (see here for example). For another participant it seemed that the introduction of a HGCSF diet *correlated* with the resolution of urination problems, where a dairy-free diet removed the need for "intermittent catheterization" and resolution of associated problems. OK, these examples don't so much focus on the core issues associated with autism, but I'm pretty sure that they were factors that would have influenced quality of life.

There is quite a bit more to see in the Adams paper and I would encourage readers to take the time to read it in its entirety. Despite the fact that not every measure showed significant effects from such an intervention regime, I like the idea that authors didn't just focus on one intervention but rather, in a systematic way, looked at a whole suite of interventions focused on nutritional and dietary factors. I believe this is more 'naturalistic' in terms of what parents/caregivers tend to report. Indeed the authors themselves discuss how: "A limitation of this study is that all participants received all treatments, whereas probably only a subset are likely to benefit from any single intervention (for example, only participants with low carnitine are likely to benefit from carnitine supplementation)." Yup.

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[1] Adams JB. et al. Comprehensive Nutritional and Dietary Intervention for Autism Spectrum Disorder—A Randomized, Controlled 12-Month Trial. Nutrients. 2018; 10(3): 369.

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"Mitochondrial Modifying Nutrients" and chronic fatigue syndrome: a pilot study

"Recent evidence suggests that mitochondrial dysfunction may play a role in the pathophysiology of chronic fatigue syndrome (CFS)" was the starting point for the study results reported by Ranjit Menon and colleagues [1].

Detailing findings -  "open-label trial" findings - following use of a 'nutraceutical combination' in a small number of participants with CFS, researchers produced evidence that further investigations might be needed. The trial protocol for their investigation can be viewed here. The 'combination' under inspection included "primary nutrients: Coenzyme Q10, Alpha lipoic acid, Acetyl-l-carnitine, N-acetyl cysteine, B Vitamins"; many of which have been shown to act on various "mitochondrial targets" in the context that mitochondria might play a role in at least some cases of CFS (see here for example), but not necessarily all (see here). Indeed, I'll draw your attention when other groups have talked about nutraceutical 'intervention' (see here) in the context of mitochondria and CFS previously (see here and see here).

Over the 16 weeks of the trial period, researchers quite regularly assessed various parameters relating to the core feature of fatigue (based on use of the Chalder Fatigue Scale) and various mood, sleep and general health variables. They observed that alongside "a significant improvement in fatigue symptoms across [the] treatment period on the Chalder Fatigue Scale" there were also some potentially important differences noted in other measures too. Not least with "clinician-reported symptom-improvement" in mind.

Obviously the emphasis on the Menon results being an open trial (i.e. not blinded/masked, not randomised, with no control group), and very much, a small open trial, mean that these are preliminary findings and shouldn't yet be informing any research or clinical opinions. The additional fact that no objective 'actigraphic' measure of physical functioning was included for study is something else to bear in mind. Such results *should* support further research; indeed, one would hope that in this new era of interest and 'changing perspectives' with CFS in mind (see here), many more investigations in this thread would be forthcoming.

And whilst on the topic of clinical trials for CFS, the news out of Norway when it comes to the use of Rituximab is not looking too good despite a previously promising start [2]. No-one said it was going to be easy...

To close, I've nabbed a screenshot of a picture from the film Unrest that I think is starting to take on an almost iconic status...

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[1] Menon R. et al. Mitochondrial Modifying Nutrients in Treating Chronic Fatigue Syndrome: A 16-week Open-Label Pilot Study. Advances in Integrative Medicine. 2017. Nov 15.

[2] Fluge Ø. et al. Benefit from B-lymphocyte depletion using the anti-CD20 antibody rituximab in chronic fatigue syndrome. A double-blind and placebo-controlled study. PLoS One. 2011;6(10):e26358.

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Mitochondria support for mitochondrial activity in [some] autism

"This study examined the effect of common mitochondrial treatments on specific mitochondrial components in a group of children diagnosed with ASD [autism spectrum disorder], some of which also were diagnosed with co-morbid mitochondrial disease."

That was the premise of the study results published by Leanna Delhey and colleagues [1] (open-access available here) and follows previous discussions suggesting that mitochondrial disease might not be totally unfamiliar to at least some autism (see here). Including some notable names on the authorship list previously linked to the area of mitochondrial functions in relation to autism (see here), the authors provide some important information about how specific mitochondrial function might be 'supported' by various interventions.

I'm not on this occasion going to venture into all the details discussed by Delhey but I do want to pick out some interesting titbits. First, of the 127 children diagnosed with an autism spectrum disorder (ASD), we are told that "15% of the sample was clinically diagnosed with mitochondrial disease." Bearing in mind this particular cohort might not be totally representative of the autistic population at large, 15% is not an insignificant figure. What this tells us is that as and when a diagnosis of autism is received, screening for a possible mitochondrial disorder should be initiated (yes, an autism diagnosis is a starting point not the finishing line and the diagnosis rarely exists in a diagnostic vacuum).

Next, various supplements were taken by participants, some of which have recognised effects on mitochondrial functions. Of particular note was the use of coenzyme Q10 (CoQ10) and carnitine; both of which have been discussed on this blog previously (see here and see here respectively) with the word 'mitochondrial' also being mentioned. Interestingly, a couple of other supplements are also included in the Delhey paper including fatty acids and folate; some of which I have to say, didn't immediately pop into my mind as being primarily mitochondrial-related (folate is though, still a hot topic when it comes to autism). The authors head into how said supplements might affect specific facets of mitochondrial function. It also reminded me that I really need to brush up on my knowledge of mitochondrial functions...

"This study provides empirical support for common mitochondrial treatments and demonstrates that the relationship between activities of mitochondrial components might be a marker to follow in addition to absolute activities." I'd agree that there is the beginnings of a roadmap for further study based on the Delhey results. That and including important parameters related to the presentation of autism and how it may/may not be affected by treating underlying mitochondrial disorder, and the scene is set for further recognition of how indeed, autism rarely exists in a diagnostic vacuum...

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[1] Delhey LM. et al. The Effect of Mitochondrial Supplements on Mitochondrial Activity in Children with Autism Spectrum Disorder. J Clin Med. 2017 Feb 13;6(2). pii: E18.

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Delhey LM, Nur Kilinc E, Yin L, Slattery JC, Tippett ML, Rose S, Bennuri SC, Kahler SG, Damle S, Legido A, Goldenthal MJ, & Frye RE (2017). The Effect of Mitochondrial Supplements on Mitochondrial Activity in Children with Autism Spectrum Disorder. Journal of clinical medicine, 6 (2) PMID: 28208802

KPAX002 for Chronic Fatigue Syndrome?

My attention was grabbed recently by the paper published by Jon Kaiser [1] (open-access available here) detailing the results of a 'proof-of-concept investigation' examining the use of something called KPAX002 on a small number of participants diagnosed with Chronic Fatigue Syndrome (CFS).

Looking at how KPAX002 - "a combination of low-dose methylphenidate hydrochloride and mitochondrial support nutrients currently under development by K-PAX Pharmaceuticals" - impacted on fatigue symptoms and "concentration disturbance" symptoms, Kaiser reports that there may be more to see from this preparation.

Indeed, after 12 weeks of use: "Treatment with KPAX002 was well tolerated and significantly improved fatigue and concentration disturbance symptoms in greater than 50% of patients with CFS." Further information about the current (and future) trial results can be found here including details on the Synergy trial (see here and see here) representing the next step in the research process. I  might also draw your attention to an interview with Dr Kasier here (thanks to Russell for the link).

Before progressing further, I should perhaps comment on a few methodological issues to bear in mind. This was very much an observational 'pilot' study over and above a thorough clinical trial. Participants knew that they were taking KPAX002 and when it came to scoring behaviours over the trial period, this was done using subjective instruments without any objective representation. That no control group was employed (either asymptomatic nor CFS taking a placebo) should be noted. Also slightly unusually, in the section titled 'Disclosure of conflict of interest' on the paper, the words 'none' appears although in the discussion section, Dr Kasier elaborates that: "as a current employee of K-PAX Pharmaceuticals, the author may be viewed as biased toward the success of this treatment."

Such issues aside, I'm interested in this formulation and findings reported. I was not aware that methylphenidate, more typically indicated as a management option for attention-deficit hyperactivity disorder (ADHD), was something 'suggested' for CFS. Kaiser does make reference to the findings reported by Blockmans and colleagues [2] who reported that under placebo-controlled conditions: "Methylphenidate at a dose of 2 x 10 mg/day is significantly better than placebo in relieving fatigue and concentration disturbances in a minority of chronic fatigue syndrome patients." Various other studies looking at the issue of 'fatigue' attached to various other diagnoses have utilised methylphenidate with varying degrees of clinical success.

The 'mitochondrial support nutrients' included in the KPAX002 preparation are a little more familiar to me. Covering 30+ additional vitamins, minerals and other nutrients including acetyl L-carnitine and N-acetylcysteine, I was interested in the possibilities here. Mitochondria and CFS is a topic that has cropped up on this blog before (see here) in light of the findings from Sarah Myhill and colleagues [2]. As a point of note, Dr Myhill's book 'Mitochondria, Not Hypochondria' received something of an accolade at the recent British Medical Association (BMA) book awards suggesting that views might be changing in this area of the CFS landscape. Although quite a bit more research is required on the topic, mitochondrial issues in relation to CFS is something of a research growth area [3].

Reiterating that the current Kasier results should be viewed with methodological caution, it will be interesting to see what becomes of KPAX002 in relation to [some] CFS. As a point of note, Kaiser also has another entry on the US ClinicalTrials.gov database for KPAX002 in relation to an equally mystifying condition: Gulf War Syndrome (GWS) on the basis of a "high degree of symptom overlap" between CFS and GWS. No doubt KPAX002 will be gracing this blog again in future...

Music: Semisonic - Secret Smile.

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[1] Kaiser JD. A prospective, proof-of-concept investigation of KPAX002 in chronic fatigue syndrome. Int J Clin Exp Med. 2015 Jul 15;8(7):11064-11074.

[2] Blockmans D. et al. Does methylphenidate reduce the symptoms of chronic fatigue syndrome? Am J Med. 2006 Feb;119(2):167.e23-30.

[3] Morris G. & Maes M. Mitochondrial dysfunctions in myalgic encephalomyelitis/chronic fatigue syndrome explained by activated immuno-inflammatory, oxidative and nitrosative stress pathways. Metab Brain Dis. 2014 Mar;29(1):19-36.

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Kaiser JD (2015). A prospective, proof-of-concept investigation of KPAX002 in chronic fatigue syndrome. International journal of clinical and experimental medicine, 8 (7), 11064-74 PMID: 26379906

The autisms, case reports and two 'intervention' options

I'm looking at two papers today which I'd like to think cover the title of this post pretty well dealing with the plurality of autism - the autisms - and the idea that intervention or management-wise, there is no 'one size fits all' when it comes to the autisms.

First up are the findings reported by Ziats and colleagues [1] who presented results for a child - "A 4-year-old male with autism and two episodes of neurodevelopmental regression" - who was also found to have a "mutation in the TMLHE gene, which encodes the first enzyme in the carnitine biosynthesis pathway, and concurrent carnitine deficiency." Supplementation with carnitine (see here) seemed to lead to some interesting changes in the developmental profile for this boy such that: "the patient's regression ended, and the boy started gaining developmental milestones."

Accepting that this was another example of the N=1 and autism (see here) I was rather interested in these results having previously blogged about issues with the TMLHE (trimethyllysine hydroxylase) gene in relation to autism (see here). The source of that previous post was the paper from Patricia Celestino-Soper and colleagues [2] (open-access) who concluded that: "TMLHE deficiency is a risk factor for autism" and quite a bit more should be done to screen for such issues. I wouldn't disagree with those sentiments (see here).

Next up are the results reported by Serret and colleagues [3] (open-access) who presented findings based on two participants "diagnosed with autism spectrum disorders in childhood and presented regression with catatonia features and behavioural disorders after a stressful event during adolescence." Further: "both patients presented mutation/microdeletion of the SHANK3 gene, inducing a premature stop codon in exon 21." Issues with SHANK3 have been reported in relation to autism previously.

Authors reported that: "lithium therapy reversed clinical regression, stabilized behavioural symptoms and allowed patients to recover their pre-catatonia level of functioning, without significant side effects." Further: "These cases support the hypothesis of a specific SHANK3 phenotype" and that lithium might hold some favour in improving clinical presentation in those cases.

Again, I was interested in the Serret findings with the caveat about their also using the case study approach in their paper. Lithium is an interesting compound that has graced this blog a few times in relation to its potential 'anti-suicide' correlating properties (see here) and as a possible management tool when it comes to the presentation of mood disorders comorbid to a diagnosis of autism (see here). Accepting that lithium has its own potential side-effects profile, the idea that cost-benefits might be calculated and if so deemed more benefit and less cost subsequently applied to 'some' autism, is an interesting prospect.

Reiterating my opening paragraph, what the Ziats and Serret papers serve to tell us is that within 'the autisms' there may be many different roads to a diagnosis of autism and that under the diagnostic label of 'autism', genetics, biochemistry and subsequent intervention/management strategies may vary from person to person. As I've said before, receipt of a diagnosis of autism (when it is eventually received) should be a starting point for further inquiry not the 'finishing line'.

That comorbidity - if I can still call it that - might also be a 'target' for analysis and investigation is also an important point raised and further asks more questions about the value of intervening on said comorbidity and the possible knock-on effects on the presentation of more core autism symptoms (see here). Y'know something like what is emerging in the body of research looking at anxiety and autism (see here).

With the body of work linking this, that and t'other to autism I'm starting to think that some further resources might be needed to pull all the available peer-reviewed information together in terms of what factors have been linked to those 'autisms'. I've always been very partial to autism research looking at inborn errors of metabolism (IEMs) as a starting point for investigations (see here) given both the data on overlap and even the idea that some of the various interventions for specific IEMs might hold promise for 'some' autism (see here). Analysis of things like rare genetic variations also being linked to the appearance of autism (see here) ties into the IEM investigations and perhaps represents the next tier of evaluation, bearing in mind the reduced costs of things like whole genome sequencing these days set within the perspective of personalised medicine (see here). Environment, bearing in mind the range of factors this might cover, should also be included in any diagnostic work-up based on the evolving science connecting something like infection to autism onset for some (see here and see here). There are various tests that could be performed covering a whole slew of potential infective agents (see here).

This is just a rough-and-ready idea of where autism research and practice could go with this but much like the pathways to diagnosing and managing bowel issues when comorbid to autism for example (see here), a general diagnostic roadmap is perhaps indicated...

Music: Years & Years - King.

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[1] Ziats MN. et al. Improvement of regressive autism symptoms in a child with TMLHE deficiency following carnitine supplementation. Am J Med Genet A. 2015 May 5.

[2] Celestino-Soper PB. et al. A common X-linked inborn error of carnitine biosynthesis may be a risk factor for nondysmorphic autism. Proc Natl Acad Sci U S A. 2012 May 22;109(21):7974-81.

[3] Serret S. et al. Lithium as a rescue therapy for regression and catatonia features in two SHANK3 patients with autism spectrum disorder: case reports. BMC Psychiatry 2015, 15:107.

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Ziats MN, Comeaux MS, Yang Y, Scaglia F, Elsea SH, Sun Q, Beaudet AL, & Schaaf CP (2015). Improvement of regressive autism symptoms in a child with TMLHE deficiency following carnitine supplementation. American journal of medical genetics. Part A PMID: 25943046




Serret, S., Thümmler, S., Dor, E., Vesperini, S., Santos, A., & Askenazy, F. (2015). Lithium as a rescue therapy for regression and catatonia features in two SHANK3 patients with autism spectrum disorder: case reports BMC Psychiatry, 15 (1) DOI: 10.1186/s12888-015-0490-1

Persistent hyperlactacidemia in cases of autism

The paper from José Guevara-Campos and colleagues [1] (open-access can be downloaded here) is fodder for today's short post, and a topic that has not been seen on this blog for quite a while: hyperlactacidemia (elevated plasma lactate levels) and autism.

Previous mentions of lactate and autism on this blog (see here and see here) were potentially pretty important; specifically, how elevated plasma lactate levels might (a) not be an unfamiliar finding for quite a few people on the autism spectrum [2] and (b) might provide further evidence for the involvement of mitochondria in cases of autism among other things [3]. Mitochondria and autism, I might add, is still quite a complicated topic but a research area in the ascendancy.

Guevara-Campos et al reported on case reports for "three patients diagnosed with developmental delay, ID [intellectual disability] and ASD [autism spectrum disorder], and also with a possible mitochondrial disease accompanied by an ETC [electron transport chain] deficiency accompanied by hyperlactacidemia." There are various data provided following some clinical investigations including that based on muscle biopsy data. Just as important are some of the details on 'pharmacological treatment' of said issues and the observed impact on presented symptoms. Without cherry-picking too much, carnitine, a vitamin B complex, co-enzyme Q10 and folic acid combined seemed to have quite an effect on participants, particularly on "intellectual abilities". Some of these interventions have been trialled in other conditions where mitochondria or their important processes are suspected to show involvement (see here). I say this without providing endorsement or recommendation.

Appreciating that there is quite a bit more to do (experimentally) when it comes to "suspected mitochondrial involvement" specifically where autism is mentioned, and in particular, the need for quite a bit more controlled study on how such interventions might impact on symptoms in this group, I'm interested in the Guevara-Campos report. How many people on the autism spectrum their results hold true for is as yet unknown. With the growth in this area of research however, I'd be minded to suggest that we should really start directing a lot more resources to trying to answer that question if we are indeed going to start taking the plural autisms a little more seriously. Oh, and as per the sentiments of the paper by Zilberter and colleagues [4] there may yet be related factors which might be of "potential therapeutic significance."

Music: Roots Manuva - Witness. Brilliant.

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[1] Guevara-Campos J. et al. Autism and Intellectual Disability Associated with Mitochondrial Disease and Hyperlactacidemia. Int J Mol Sci. 2015 Feb 11;16(2):3870-3884.

[2] Oliveira G. et al. Mitochondrial dysfunction in autism spectrum disorders: a population-based study. Dev Med Child Neurol. 2005 Mar;47(3):185-9.

[3] Andersen LW. et al. Etiology and therapeutic approach to elevated lactate levels. Mayo Clin Proc. 2013 Oct;88(10):1127-40.

[4] Zilberter Y. et al. A unique array of neuroprotective effects of pyruvate in neuropathology. Front. Neurosci. 2015. Feb 17.

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Guevara-Campos J, González-Guevara L, & Cauli O (2015). Autism and Intellectual Disability Associated with Mitochondrial Disease and Hyperlactacidemia. International journal of molecular sciences, 16 (2), 3870-3884 PMID: 25679448

Carnitine and autism continued

The paper from everyone's favourite Saudi - Egyptian autism research tag-team that is Gehan Mostafa and Laila AL-Ayadhi [1] (open-access) on plasma polyunsaturated fatty acids and serum carnitine levels in a cohort of children diagnosed with autism / autism spectrum disorder (ASD) is served up for your reading delight today.

Regular readers of this blog might have heard me talk before about the pretty interesting research findings to come from this research partnership (see here and see here for example) covering all-manner of different sectors of the autism research environment.

This times around the name of the game was: "to investigate plasma levels of PUFAs [polyunsaturated fatty acids] and serum carnitine in relation to GI [gastrointestinal] manifestations in autistic children." The idea being that: "Carnitine and PUFAs are antiinflammatory molecules and their deficiency may result in GI inflammation and gut injury" following other work with autism in mind (see here).

A few pointers about the study might be in order:

• "This cross-sectional study was conducted on 100 children with autism." Participants ranged in age between 3-10 years and importantly were not taking additional fatty acids or anticonvulsants. A control group of 100 age- and sex-matched children asymptomatic for autism were also studied: "not related to the children with autism, and demonstrated no clinical findings suggestive of immunological, GI or neuropsychiatric disorders."
• Autism severity was assessed using the CARS and GI issues were examined "by an experienced pediatric gastroenterologist according to the Questionnaire on Pediatric Gastrointestinal Symptoms - Rome III Version used by previous studies that assessed gastrointestinal dysfunction in autism." Fasting blood samples were also provided and serum carnitine and plasma PUFAs examined.
• Results: both biological measures were lower as a group for the children with autism compared to controls. Indeed: "Low serum carnitine and plasma DHA [Docosahexaenoic acid], AA [Arachidonic acid], linolenic and linoleic acids, below the 5th percentile of the control values, were found in 66%, 62%, 60%, 43% and 38%, respectively of autistic children."
• Bearing in mind that PUFAs can exist in more than one form as per the old omega-3 / omega-6 issue (see here), authors also reported a group difference in the ratio of ω6/ω3 PUFAs: "ω6/w3 ratio (AA/DHA) was significantly higher in autistic patients" compared with controls. This is something also reported by the authors in other publications [2].
• GI symptoms were reported to be present in about half of the autism group. An important sentence is included about GI issues: "They were recurrent, severe and the patients were attending the clinic because of these agonizing symptoms." Further: "Autistic patients with GI manifestations had significantly lower serum carnitine and plasma DHA than patients without such manifestations."

I know I tend to say this about nearly every study I blog about but this is interesting work. Carnitine and autism is an area which I've got quite a bit of time for on this blog and the growing consistency in results suggesting lower levels of this stuff in quite a few cases of autism (see here). Some of the genetics of carnitine metabolism might also be 'linked' to at least some autism too (see here) with a particular focus on the idea of inborn errors of metabolism. Certainly, autism research should know a thing or two about them (see here).

The relationship between fatty acids and autism reflects a slightly less clear picture in terms of results. In a post going back to 2011, I talked about some of the peer-reviewed research on the use of supplementary fatty acids for autism (see here) and how certain comorbidity present in quite a bit of autism (see here) might be the bigger target. More recent research has kinda corroborated that idea (see here).

"How GI factors are related to autism is not yet clear" is another important quote from the authors. Yes, we are now in an era where there is general acceptance that certain functional GI issues are over-represented among those with autism (see here) but the hows and whys are still the source of significant speculation. I'd be minded to suggest that it's likely to be complicated and probably without a universal factor for everyone with autism and GI issues. There are some areas emerging that may yield further information such as examination (not hype) of those trillions of wee beasties which call our gut home (see here) and a 'possible' relationship with more pathological bowel states (see here). The Mostafa/AL-Ayadhi findings suggest another possible correlate.

I leave you with a quote from the authors: "these data should be treated with caution until further investigations are performed, with a larger subject population, to determine whether the occurrence of GI manifestations is a mere association or a consequence to reduced plasma PUFAs and serum carnitine levels in autistic patients." I couldn't agree more.

Music then. The Strokes with New York City Cops.

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[1] Mostafa GA. & AL-Ayadhi LY. Reduced levels of plasma polyunsaturated fatty acids and serum carnitine in autistic children: relation to gastrointestinal manifestations. Behavioral and Brain Functions 2015, 11:4.

[2] Mostafa GA. et al. A possible association between elevated serum levels of brain-specific auto-antibodies and reduced plasma levels of docosahexaenoic acid in autistic children. J Neuroimmunology. 2015. Jan 27.

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Mostafa, G., & AL-Ayadhi, L. (2015). Reduced levels of plasma polyunsaturated fatty acids and serum carnitine in autistic children: relation to gastrointestinal manifestations Behavioral and Brain Functions, 11 (1) DOI: 10.1186/s12993-014-0048-2

Very long chain acyl-CoA dehydrogenase (VLCAD) deficiency and autism?

I started this blogging year (2014) with a post talking about the need for further research consideration for a possible role of inborn errors of metabolism in relation to the presentation of at least some autism (see here). It is therefore timely that I (almost) end this year's blogging odyssey with reference to the paper by Amy Brown and colleagues [1] and their suggestion that although: "VLCAD deficiency does not have a significant impact on cognitive or motor skills" there may still be merit in looking further at very long chain acyl-CoA dehydrogenase (VLCAD) deficiency where: "Some children may be vulnerable to speech, social and behavioural issues."

Rule 42: All persons more than a mile high
must leave the court immediately

Although based on quite a small participant number (N=7), probably reflective of the fact that VLCAD deficiency is quite a rare disorder ("estimated at 1:30,000 in the US"), Brown and colleagues reported various findings based on the use of "a comprehensive neuropsychological assessment battery that assessed IQ, language, attention, memory, executive functioning, motor skills, behaviour, and social skills". Parents were also asked about their child's abilities in terms of things like social skills and behaviour.

Quite a few of the parameters investigated suggested that kids with VLCAD deficiency were average or above on their performance and presentation of skills. That being said: "Parents' questionnaires identified one child as having social skills deficits, and two as having behavioural problems such as hyperactivity. One child rated high on an autism spectrum subscale; another was formally diagnosed with autism spectrum disorder-both children were symptomatic at birth." I can't specifically provide you with answers as to how and why VLCAD deficiency might link to autism/autistic traits, but will provide you with a link to a previous paper [2] talking about long chain acyl-CoA dehydrogenase (LCAD) deficiency with a case report of autism in mind. LCAD and VLCAD have some interesting history. In that case, acyl-carnitines were discussed as potentially being relevant, which may very well tie into some other work in this area (see here). The fact also that carnitine is used as part of the treatment regime for some VLCAD deficiency might also overlap with other autism research (see here).

The two children talked about with autism in mind, one diagnosed with an autism spectrum disorder (ASD) and the other with some indication of autistic-like traits, hint at how some of the inborn errors of metabolism may yet provide some interesting insight into at least some autism. The paper by Burrage and colleagues [3] for example, updating on the field of branched-chain amino acid (BCAA) metabolism, offers further discussion on the possibility of a more generalised association as per the rise and rise of 'BCKDK autism'.

Bearing in mind our growing realisation of the plurality of autism - 'the autisms' - denoting not just the heterogeneity covered under the umbrella clinical description but also that more than one road might lead to the presentation of autistic traits, examining the inborn errors of metabolism with autism in mind is an area of great scientific potential. It benefits from some testable genetic/biological starting points with the detection of those inborn errors of metabolism which may provide some important insights into how [some] autism might come about. Given that some of those errors of metabolism can also be 'corrected' in various ways, it might also provide some pretty interesting data on how behavioural presentation might also be affected. Remember PKU? I'd like to see a lot more in this area.

A quick heads-up... tomorrow (30th December 2014) I'm gonna publish my annual round-up of some of the blogging highlights here on Questioning Answers in 2014. You're all invited to drop in and take a gander...

And then to some music: Elvis and Suspicious Mind.

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[1] Brown A. et al. Neurodevelopmental profiles of children with very long chain acyl-CoA dehydrogenase deficiency diagnosed by newborn screening. Mol Genet Metab. 2014 Oct 12. pii: S1096-7192(14)00314-X.

[2] Clark-Taylor T. & Clark-Taylor BE. Is autism a disorder of fatty acid metabolism? Possible dysfunction of mitochondrial beta-oxidation by long chain acyl-CoA dehydrogenase. Med Hypotheses. 2004;62(6):970-5.

[3] Burrage LC. et al. Branched-chain amino acid metabolism: from rare Mendelian diseases to more common disorders. Hum Mol Genet. 2014 Sep 15;23(R1):R1-8.

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Brown A, Crowe L, Andresen BS, Anderson V, & Boneh A (2014). Neurodevelopmental profiles of children with very long chain acyl-CoA dehydrogenase deficiency diagnosed by newborn screening. Molecular genetics and metabolism PMID: 25456746

Ketogenic diet and the valproate mouse model of autism

A brief entry today and yet another blog post that starts with a quote (sorry)... "The offspring exposed to VPA [valproic acid] prenatally demonstrated a significant decrease in the number of play initiations/attacks and this was reversed with the KD [ketogenic diet]".

Gloucester Old Spot @ Wikipedia 

That finding reported in the paper by Ahn and colleagues [1] continues my interest in all-things related to prenatal VPA exposure and the reported effects on some offspring (see here). The added bonus of including some discussion about how the use of a ketogenic diet might reverse some of the effects of VPA exposure (in rats at least) is also worthwhile mentioning.

A couple of pointers perhaps...

• Rats, Sprague-Dawley mother rats, were given VPA or saline (as a control) during pregnancy and their pups (VPA-exposed vs. controls) were subjected to measures looking at "juvenile play behavior" and eventually "mitochondrial bioenergetic analysis" as a function of the use of a ketogenic or standard diet.
• Results: "Prenatal VPA exposure also disrupted the pattern of play responses". Not a great surprise there given everything else that has been linked to VPA exposure in-utero. But.. use of the ketogenic diet "was able to modify complex social behaviors and mitochondrial respiration". As noted previously, the reduction in play initiations made by the VPA exposed mice was to some degree rescued following use of the ketogenic diet.

Yes, I know that this was a study of rats, and whilst useful, rats are rats not humans. But I am nevertheless intrigued by the suggestion that something like a ketogenic diet - more typically indicated for some types of treatment resistant epilepsy - might to some degree, affect the behaviour and physiology of animals exposed to a traditional anticonvulsant like valproate during the nine months that made them. Does anyone else find that a little ironic? Also throw in mention of the words 'autism spectrum disorder' alongside that animal VPA exposure model alongside the ketogenic diet (see here) and I'm sure there's some more research to be done in this area.

Mode of action? I dunno. I will draw your attention to some interesting work on carnitine homoeostasis as a function of valproate administration [2] which might be relevant. Carnitine plays a role in mitochondrial function [3] and there is some suggestion that a ketogenic diet might help maintain carnitine levels in the presence of VPA [4]. Whether this applies to brain structures or neurochemistry potentially already affected by prenatal exposure to VPA is a question not yet asked or answered. Bearing in mind the gastrointestinal (GI) effects also noted in VPA exposure models (see here) I might also be inclined to 'look to the bowels' in terms of any potential effects from the ketogenic diet in that organ too.

Music to close and I was taken aback by the performance from Pumeza at the opening to the 2014 Commonwealth Games and her version of Freedom Come All Ye...

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[1] Ahn Y. et al. The Ketogenic Diet Modifies Social and Metabolic Alterations Identified in the Prenatal Valproic Acid Model of Autism Spectrum Disorder. Dev Neurosci. 2014 Jul 8.

[2] Morand R. et al. Effect of short- and long-term treatment with valproate on carnitine homeostasis in humans. Ther Drug Monit. 2012 Aug;34(4):406-14.

[3] Zammit VA. et al. Carnitine, mitochondrial function and therapy. Adv Drug Deliv Rev. 2009 Nov 30;61(14):1353-62.

[4] Coppola G. et al. Plasma free carnitine in epilepsy children, adolescents and young adults treated with old and new antiepileptic drugs with or without ketogenic diet. Brain Dev. 2006 Jul;28(6):358-65.

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Ahn Y, Narous M, Tobias R, Rho JM, & Mychasiuk R (2014). The Ketogenic Diet Modifies Social and Metabolic Alterations Identified in the Prenatal Valproic Acid Model of Autism Spectrum Disorder. Developmental neuroscience PMID: 25011527

BCKDK mutations and autism continued?

February 19th 2014 (yesterday). Questioning Answers celebrates 3 years of life as a blog. Happy birthday. To quote from one website: "Congratulations, you have survived the "terrible two's!"'. Maybe I should start claiming some free early education for it too?

You are 3 @ Paul Whiteley

Anyhow...

Cast your mind back to September 2012 and the publication of a paper by Gaia Novarino and colleagues* which I posted about (see here) discussing some interesting observations with respect to a potentially treatable form of autism.

The main point of the Novarino research was how genetic mutations upset a delicate biochemical balance. Various 'inactivating' genetic mutations in respect of the BCKDK (Branched Chain Ketoacid Dehydrogenase Kinase) gene have knock-on implications for the production or rather non-production of BCKDK protein. Lower levels of this protein/enzyme left another important group of enzymes, the branched-chain keto-acid dehydrogenase complex (BCKD) to run unchecked which has implications for the metabolism and over-zealous degradation of branched-chain amino acids. The resulting lower levels of these branched-chain amino acids (BCAAs) seemed to correlate with presence of epilepsy and behavioural issues not a million miles away from those which characterise autism. Supplementation with BCAAs seemed to have a positive restorative effect on said behavioural issues. And rest.

Both then and now I consider these findings to be extremely interesting. As indicated on my post at the time, they hint at both some interesting roles for various amino acids in cases of autism (see here) and provided some exquisite evidence for the plurality of autism: the autisms (see here). That they also follow a pattern of inborn errors of metabolism being potentially linked to autism was also an important point.

Enter then another paper on BCKDK mutations by García-Cazorla and colleagues** found in two children presenting with "developmental delay, microcephaly and neurobehavioral abnormalities" also demonstrating the potential power of the BCAAs in affecting symptom presentation.

A few details about the latest study:

• It was a small study looking at two children who presented with "two novel exonic BCKDK mutations, c.520C>G/p.R174G and c.1166T>C/p.L389P, [that] were identified at the homozygous state". Homozygous refers to the issue of genetic zygosity which, as the results go on to report, very much impacted on the function of BCKDK: "Mutation p.L389P showed total loss of kinase activity".
• Authors also confirmed how detrimental the mutations were based on analysis of "patient-derived fibroblasts" and how undetectable or barely detectable levels of BCKDK protein (the product of the BCKDK gene) resulted "in increased BCKD activity and the very rapid BCAA catabolism".
• They conclude that for one of the children who presented with undetectable levels of the BCKDK protein, use of a BCAA supplement "normalized plasma BCAA levels and improved growth, developmental and behavioral variables".

Obviously one has to be a little careful in extrapolating these results to the wider population of autism. This is a report of specific identified cases where genetic mutations of the BCKDK gene were found and the resultant biological effect of mutation confirmed. They don't imply that supplementing willy-nilly with BCAAs will impact on all cases of autism and neither am I suggesting they will. Autisms not autism...

That being said, there is another message to come from this paper on the value of keeping an open mind when it comes to look at the potential underlying genetics and biochemistry of autism. Not so long ago for example, I talked about the paper by Spilioti and colleagues (see here) on the presence of inborn errors of metabolism in their participant group diagnosed with autism. The message there was if you don't look, don't expect to find anything. Indeed, if you need some further evidence for this message I would also direct you to the work by Celestino-Soper and colleagues (see here) on another compound, carnitine, and their analysis of the gene Trimethyllysine Hydroxylase, Epsilon (TMLHE) in relation to carnitine and autism. Again, you've got to look to find or not find. And the papers keep coming***...

I'm going to keep my eye out for any further information on BCKDK mutations...

To close, no music link today but rather following some rather interesting conversations with my brood about winners and losers across the previous 6 films of the Star Wars saga, I vote for Darth Sidious as being the real winner... "unlimited power" and all that.

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* Novarino G. et al. Mutations in BCKD-kinase Lead to a Potentially Treatable Form of Autism with Epilepsy. Science. 2012; 338: 394-397.

** García-Cazorla A. et al. Two Novel Mutations in the BCKDK Gene (Branched-Chain Keto-Acid Dehydrogenase Kinase) are Responsible of a Neurobehavioral Deficit in two Pediatric Unrelated Patients. Hum Mutat. 2014 Jan 21. doi: 10.1002/humu.22513.

*** Helsmoortel C. et al. A SWI/SNF-related autism syndrome caused by de novo mutations in ADNP. Nature Genetics. 2014. 16 Feb.

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García-Cazorla A, Oyarzabal A, Fort J, Robles C, Castejón E, Ruiz-Sala P, Bodoy S, Merinero B, Lopez-Sala A, Dopazo J, Nunes V, Ugarte M, Artuch R, Palacín M, Rodríguez-Pombo P, & Working Group (2014). Two Novel Mutations in the BCKDK Gene (Branched-Chain Keto-Acid Dehydrogenase Kinase) are Responsible of a Neurobehavioral Deficit in two Pediatric Unrelated Patients. Human mutation PMID: 24449431

Acyl-carnitine profiles and autism

No doubt alongside quite a few others, I was interested to read the latest paper from Richard Frye and colleagues* (open-access) discussing the potential links between an animal model of autism spectrum disorder (ASD) and real-world ASD in a particular cohort of participants.

This is not the first time that I've talked about (a) the work of Dr Frye - as per my [don't panic] post on folate receptor autoantibodies in cases of autism (see here) and (b) some of the difficulties attached to translating rat/mouse model findings in autism research into 'real-world' findings (see here). Indeed this last point might also tie into my recent musings on the use of LPS in autism research (see here).

Daddy o' microbiology @ Wikipedia  

My interest in the latest Frye paper was further piqued upon realising that Derrick MacFabe was also part of the authorship team given his insightful work on how administration of propionic acid - a short chain fatty acid - to rodents might provide some interesting perspectives overlapping with various findings noted in cases of autism (see my previous post here).

The Frye paper is open-access and available to all but here are a few points worth mentioning:

• Based on the animal model work looking at propionic acid (PPA) infusions and some of the collected effects (presentation of autism-type behaviours**, mitochondrial issues***, abnormal measures of glutathione, etc.), the aim of the current study was to map out whether some of the observations in the PPA rodent model were also present in real-life autism.
• Based on a clinic-based sample of children with autism (N=326), acyl-carnitine panels were conducted on about two-thirds of participants (n=213).
• Acyl-carnitines, as their name suggests, are related to carnitine which has previously been tied back to autism (see here) and represent a number of different compounds - complexes of carnitine and various fatty acids - involved in the transport of fatty acids into the mitochondrial matrix.
• The previous PPA rodent model indicated some disturbance in the amount of various acyl-carnitines, particularly with regards to short- and long-chain fatty acids (not so much in the medium-chain fatty acids) reflective of some mitochondrial dysfunction.
• So the authors looked at short-, medium- and long-chain acyl-carnitines in their autism cohort to see if there were any overlaps compared with the PPA treated animals.
• Results: 35% of the autism cohort showed "an increase in three or more acyl-carnitines when initially measured". When further testing were conducted on some of these participants showing elevation, this figure was revised to 17% of the cohort who "demonstrated consistent elevations in short-chain and long-chain, but not medium-chain, acyl-carnitines". You'll note the similarity with PPA animal model in terms of the short- and long-chain acyl-carnitines.
• Furthermore, four participants were also examined with regards to glutathione and oxidative stress markers. As per the quite consistent literature on things like total- and free-reduced glutathione (see this post), compared with controls, there were some familiar trends emerging.

And relax.

There is a lot to take in from this paper both in the protocol and testing undertaken and the possible interpretation of findings. A quote best sums up the results: "This study has demonstrated that ~17% of children with ASD manifest biomarkers of abnormal mitochondrial fatty-acid metabolism that parallel similar biomarkers in the PPA rodent model of ASD".

As perhaps expected, there has been some press attention following the publication of this paper. The headline: Researchers discover link between certain types of autism and gut bacteria has been a common one, reflective of the fact that when it comes to the production of PPA outside of injecting the stuff directly into the rodent brain, the gut and in particular, certain types of gut bacteria, have been suggested as a route to PPA production in cases of autism. Indeed, I'm taken back to the findings by Wang and colleagues from Oz (see this post) on levels of fecal short chain fatty acids (SCFAs) in their cohort of children with autism which included propionic acid (propionate). I'm also reliably informed that there is 'more to come' from the Australian research group in the coming months.

There are a few final aspects to the Frye study which I should have mentioned earlier. This includes the notion that "it is very likely that MD [mitochondrial dysfunction] is acquired" given that in the most part, both nuclear and mitochondrial DNA examinations in their cohort were negative for anything that might genetically account for the results. I'll be coming to DNA and mitochondrial issues in a subsequent 'training' post scheduled quite soon but one can perhaps see how this might strengthen any argument of disruptions to gut bacteria/microflora facilitating the production of elevated PPA onwards to disrupting acyl-carnitine chemistry.

Additionally there is the implication based on the PPA rodent model that affecting PPA production could be a potential therapeutic route for some cases of autism at some point. I don't really want to go too far into the hows and whys of this suggestion at the current time given the current lack of evidence for any effect. Aside that is, from drawing your attention to a related field of inquiry into how disrupting PPA-producing gut bacteria might have some important implications for those suffering from conditions like propionic acidaemia****.

I'm not going to get too carried away with the Frye results as they stand. As per my previous post on PPA, there is still a bit of a stretch from injecting rodent brains with PPA and recording outcomes to suggesting that gut bacteria will be able to produce enough PPA so as to exert a similar effect in cases of autism. At least one group is asking the same question***** (open-access). That being said, I am still interested in the details of this study and how the authors have at least tried to model animal findings into real-life autism. Indeed how we really should be putting a lot more research effort into looking at mitochondria and autism as we potentially also should gut bacteria and autism.

A song to finish... something mellow yet catchy.... Lovefool by the Cardigans.

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* Frye RE. et al. Unique acyl-carnitine profiles are potential biomarkers for acquired mitochondrial disease in autism spectrum disorder. Translational Psychiatry. 2013; 3: e220.

** MacFabe DF. et al. Effects of the enteric bacterial metabolic product propionic acid on object-directed behavior, social behavior, cognition, and neuroinflammation in adolescent rats: Relevance to autism spectrum disorder. Behav Brain Res. 2011; 217: 47-54.

*** Thomas RH. et al. Altered brain phospholipid and acylcarnitine profiles in propionic acid infused rodents: further development of a potential model of autism spectrum disorders. J Neurochem. 2010; 113: 515-529.

**** Mellon AF. et al. Effect of oral antibiotics on intestinal production of propionic acid. Arch Dis Child 2000; 82: 169-172.

***** El-Ansary AK. et al. Comparative study on the protective effect of carnosine and carnitine against pro-inflammatory/pro-oxidant effects of clindamycin and propionic acid administrations to hamsters. African Journal of Microbiology Research. 2013; 7: 103-114.

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Frye RE, Melnyk S, & Macfabe DF (2013). Unique acyl-carnitine profiles are potential biomarkers for acquired mitochondrial disease in autism spectrum disorder. Translational psychiatry, 3 PMID: 23340503