Urinary Untargeted Metabolic Profile Differentiates Children with Autism from Their Unaffected Siblings

Abstract

Autism Spectrum Disorder (ASD) encompasses a clinical spectrum of neurodevelopmental conditions that display significant heterogeneity in etiology, symptomatology, and severity. We previously compared 30 young children with idiopathic ASD and 30 unrelated typically-developing controls, detecting an imbalance in several compounds belonging mainly to the metabolism of purines, tryptophan and other amino acids, as well as compounds derived from the intestinal flora, and reduced levels of vitamins B6, B12 and folic acid. The present study describes significant urinary metabolomic differences within 14 pairs, including one child with idiopathic ASD and his/her typically-developing sibling, tightly matched by sex and age to minimize confounding factors, allowing a more reliable identification of the metabolic fingerprint related to ASD. By using a highly sensitive, accurate and unbiased approach, suitable for ensuring broad metabolite detection coverage on human urine, and by applying multivariate statistical analysis, we largely replicate our previous results, demonstrating a significant perturbation of the purine and tryptophan pathways, and further highlight abnormalities in the “phenylalanine, tyrosine and tryptophan” pathway, essentially involving increased phenylalanine and decreased tyrosine levels, as well as enhanced concentrations of bacterial degradation products, including phenylpyruvic acid, phenylacetic acid and 4-ethylphenyl-sulfate. The outcome of these within-family contrasts consolidates and extends our previous results obtained from unrelated individuals, adding further evidence that these metabolic imbalances may be linked to ASD rather than to environmental differences between cases and controls. It further underscores the excess of some gut microbiota-derived compounds in ASD, which could have diagnostic value in a network model differentiating the metabolome of autistic and unaffected siblings. Finally, it points toward the existence of a “metabolic autism spectrum” distributed as an endophenotype, with unaffected siblings possibly displaying a metabolic profile intermediate between their autistic siblings and unrelated typically-developing controls.

Urine metabolomic profiles of autism and autistic traits – a twin study

“…Despite the fact that we only found nominally associated metabolites, our findings align with previously reported urine-based metabolomic studies for autism diagnosis. For instance, we observed elevated levels of phenylpyruvate (Noto et al, 2014;Timperio et al, 2022) and taurine (Ma et al, 2021;Mavel et al, 2013;Nadal-Desbarats et al, 2014;Yap et al, 2010), as noted earlier in several studies. We also show decreased levels of carnitine, similarly to what was reported in other biological samples (Fahmy et al, 2013;Filipek et al, 2004).…”

“…This is an important aspect to account for, when investigating suitable biomarkers in lieu of the high prevalence of co-occurring NDCs in autism (Khachadourian et al, 2023). The size of our study cohort (N=96, with n=38 diagnosed with autism) is larger when compared to several recently published studies using urine-based mass spectrometry, such as 60 available samples from an otherwise larger cohort (Liu et al, 2022), 57 participants (Mussap et al, 2020) and 14 autism discordant sibling pairs (Timperio et al, 2022). The twin cohort also gives us the possibility to analyse the effects of genetic and environmental factors on these potential biomarkers.…”

“…Early efforts to understand the urine metabolome in autism began several decades ago, with initial biochemical studies searching for associations (Himwich et al, 1972;Young et al, 1978). The investigations have shifted to more untargeted and high-precision analytical approaches using NMR (Ma et al, 2021;Mavel et al, 2013) and MS (Mussap et al, 2020;Noto et al, 2014;Timperio et al, 2022).…”

Maternal Inflammation with Elevated Kynurenine Metabolites Is Related to the Risk of Abnormal Brain Development and Behavioral Changes in Autism Spectrum Disorder

“…These are all considered risk factors for ASD [ 204 , 205 , 206 ]. Even though single compound changes of KP are still controversial in each clinical study ( Table 1 ) [ 1 , 2 , 8 , 207 , 208 , 209 , 210 , 211 , 212 , 213 , 214 , 215 , 216 , 217 , 218 , 219 ], it is clear that an abnormal balance of KP metabolites may be associated with adverse fetal neurodevelopment and the pathogenesis of ASD.…”

Physical Activity, Gut Microbiota, and Genetic Background for Children and Adolescents with Autism Spectrum Disorder

“…In another urinary metabolomic study, the authors identified significant dysregulation to the purine, tyrosine, phenylalanine and tryptophan pathways, characterized by increased phenylalanine and decreased tyrosine levels. As a result of bacterial degradation, the concentrations of phenylacetic acid, phenylpyruvic acid, and 4-ethylphenyl-sulfate were also elevated [ 168 ].…”