Autism and the measurement of urinary amino acids

Today's post concerns the findings reported by Aiping Liu and colleagues [1] who, following the analysis of urine samples from a group of children diagnosed with an autism spectrum disorder (ASD) and a not-autism control group, concluded that there may be something to see with regards to the urinary excretion of amino acids.

First things first, amino acids are the building blocks of proteins. Long chains of amino acids form different proteins (and peptides) that serve multiple biological functions. But making up proteins is but one of the roles of amino acids, as a variety of other functions are also included in their repertoire; notably also being the raw material for the formation of some neurotransmitters and related compounds (see here for example).

Liu et al approached their analysis of amino acids in relation to autism from the point of view of their measurement being "potential novel metabolic biomarkers for ASD." This follows something of a trend in autism research circles whereby patterns of certain amino acids and their associated chemistry in certain biofluids might have such 'potential' for some types of autism (see here and see here and see here for some other examples) albeit with certain caveats. Researchers utilised some quite well known methods when it came to their analysis - "liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based analysis" - and set to work using a tried-and-tested method (see here): "a two-step discovery–validation approach."

Analysing urine samples from nearly 60 children with autism and over 80 not-autism controls ("28 ASD and 41 TD  [typically developing] children for the discovery stage and from an additional cohort of 29 ASD and 41 TD children for the validation stage"), researchers reported detecting and identifying "63 UAA [urinary amino acid] indicators." Twenty-one of these amino acids and/or amino acid metabolites were observed to be "present at significantly different levels in the urine of ASD children compared with TD children" in both participant sets. These compounds were fairly evenly either higher or lower in the kids with autism group (10 higher and 11 lower). I was particularly interested to see that creatinine was observed to be in the significantly higher category associated with the autism group given some other results that were counter to this finding (see here and see here) including some of my own published data [2] from a few years back. Authors also mention how they "identified a panel of 7 UAA indicators that [most effectively] discriminated between the samples from ASD and TD children (lysine, 2-aminoisobutyric acid, 5-hydroxytryptamine, proline, aspartate, arginine/ornithine, and 4-hydroxyproline)."

From those compounds, a few themes emerged with regards to the biochemistry that *might* show some involvement with autism. So: "Abnormalities in the Methionine Cycle in Children With ASD", "Evidence of High Oxidative Stress Levels in Children With ASD" and "Abnormalities in 5HT Metabolism in Children With ASD" are some of the systems potentially implicated by Liu et al. Needless to say that such biological systems are by no means strangers to autism research (see here and see here for examples) albeit not necessarily always in the same direction as the Liu findings.

Caveats? Well yes, a few, such as a reliance solely on single spot urine samples rather than multiple samples from the same person, no other measures of amino acid content in blood for example, and the focus on participants diagnosed with autism excluding things like "attention-deficit hyperactivity disorder, obsessive compulsive disorder" where 'real-life autism' rarely exists in some sort of diagnostic vacuum (see here). But, the findings are interesting and once again highlight how metabolomics is something particularly valuable to autism research (see here) and complementary to genetic studies for example, when trying to decipher the very heterogeneous autisms (plural). Issues with certain amino acids when identified in the context of autism *might* also point to a wider issue (see here) that could also indicate intervention too...

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[1] Liu A. et al. Altered urinary amino acids in children with autism spectrum disorders. Front. Cell. Neurosci. 2019. Jan 10.

[2] Whiteley P. et al. Spot urinary creatinine excretion in pervasive developmental disorders. Pediatr Int. 2006 Jun;48(3):292-7.

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"a 256-peptide immunosignature with the ability to separate ME/CFS cases from controls"

The findings reported by Oliver Günther and colleagues [1] (open-access available here) really interested me. They interested me because they talked about the "hit and run" hypothesis being pertinent to myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS), where "a pathogen or other immunological insult experienced by a subject may be gone, but leaves behind physiological disequilibrium." They interested me because researchers turned to "an immunosignature assay (ISA) that employs a microarray of thousands of random-sequence peptides to interrogate antibodies in a broad and unbiased fashion" to try and pick out the biological effects of the 'hit and run' hypothesis in relation to ME/CFS. And they interested me because researchers reported that they were able to identify "a 256-peptide signature that separates ME/CFS samples from healthy controls, suggesting that the hit-and-run hypothesis of immune dysfunction merits further investigation." Lots of interesting things (honest!).

The Günther paper is open-access so doesn't really need too grand an explanation from me. The long-and-short of it was that following the adoption of a discovery and validation methodology (an increasingly favoured option in ME/CFS research circles), authors came up with a sort of 'biological fingerprint' "optimally separating ME/CFS cases and controls" based on the examination of serum samples. They also noted that in amongst their 256-peptide signature, one particular peptide - "LRVVWLSGVASG" - was also mentioned in another independent study with similar aims [2] perhaps therefore requiring further research focus. For those who might not be totally au fait with peptide designation, that string of letters is not meant to be pronounced, but rather each letter corresponds to an amino acid making up that particular peptide.

Whilst this is great work and indeed, represents some really quite detailed analysis, the authors caution that the science is not quite there yet when it comes to a 'biological test' for ME/CFS. So: "the heterogenous nature of ME/CFS clinical presentation and the variance natural present amongst control samples means that group labels in the Discovery and Validation Sets are not based on any gold standard." Diagnosis of ME/CFS still remains a point of real contention in various circles (see here) given the variety of diagnostic criteria available. Indeed, some commentators have suggested that the combination of 'ME/CFS' as a unified diagnostic label simply cannot ever exist (see here). The authors further note that: "Even the best research case definitions are often subjective and—in the absence of clear biomarkers—any group of ME/CFS cases likely comprise a heterogeneous set of pathologies."

I'm also minded to suggest that as per the lessons being learned in connection to autism biomarker research for example (see here), one needs to perhaps think about getting different research groups together who are looking at ME/CFS from different angles (see here). Y'know, sort of combining various different biomarker studies looking at various different biological 'angles' and sorta meta-analysing all the collected data to see if a larger, grander, range of variables might provide a more accurate biomarker picture of the condition(s)...

Still, the Günther study represents some good science and good value-for-research-money. It stresses how, by utilising the pretty sophisticated analytical equipment available these days, one can start creeping ever closer to some of the possible biochemistry that underpins ME/CFS (or at least some ME/CFS) and perhaps then also start some conversations centred on what can be done to alleviate symptoms and cure such a devastating illness.

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[1] Günther OP. et al. Immunosignature Analysis of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS). Mol Neurobiol. 2018 Oct 8.

[2] Singh S. et al. Humoral Immunity Profiling of Subjects with Myalgic Encephalomyelitis Using a Random Peptide Microarray Differentiates Cases from Controls with High Specificity and Sensitivity. Mol Neurobiol. 2018 Jan;55(1):633-641.

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