Amino Acid Dysregulation Metabotypes: Potential Biomarkers for Diagnosis and Individualized Treatment for Subtypes of Autism Spectrum Disorder

Abstract

Background: Autism spectrum disorder (ASD) is behaviorally and biologically heterogeneous and likely represents a series of conditions arising from different underlying genetic, metabolic, and environmental factors. There are currently no reliable diagnostic biomarkers for ASD. Based on evidence that dysregulation of branched-chain amino acids (BCAAs) may contribute to the behavioral characteristics of ASD, we tested whether dysregulation of amino acids (AAs) was a pervasive phenomenon in individuals with ASD. This is the first article to report results from the Children's Autism Metabolome Project (CAMP), a large-scale effort to define autism biomarkers based on metabolomic analyses of blood samples from young children.

Methods: Dysregulation of AA metabolism was identified by comparing plasma metabolites from 516 children with ASD with those from 164 age-matched typically developing children recruited into the CAMP. ASD subjects were stratified into subpopulations based on shared metabolic phenotypes associated with BCAA dysregulation.

Results: We identified groups of AAs with positive correlations that were, as a group, negatively correlated with BCAA levels in ASD. Imbalances between these two groups of AAs identified three ASD-associated amino acid dysregulation metabotypes. The combination of glutamine, glycine, and ornithine amino acid dysregulation metabotypes identified a dysregulation in AA/BCAA metabolism that is present in 16.7% of the CAMP subjects with ASD and is detectable with a specificity of 96.3% and a positive predictive value of 93.5% within the ASD subject cohort.

Conclusions: Identification and utilization of metabotypes of ASD can lead to actionable metabolic tests that support early diagnosis and stratification for targeted therapeutic interventions.

Trial registration: ClinicalTrials.gov NCT02548442.

Keywords: Amino acids; Autism; Biomarker; Diagnosis; Metabolomics; Metabotype.

Copyright © 2018 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Figures

Figure 1.

Heat map with hierarchical clustering dendrograms from pairwise Pearson correlations of metabolite abundances for the training set ASD subjects. Red filled boxes associated with the dendrograms identify statistically significant clusters following bootstrap resampling. The names of these clusters appear within the red boxes. The green open boxes highlight the BCAA cluster in the columns and the glycine cluster in the rows. The intersection of the two green boxes, marked by a yellow open rectangle, identifies the block of negative correlations shared by the glycine and BCAA clusters. Abbreviations: BCAA, branched chain amino acids; BAIBA, β-Aminoisobutyric Acid; GABA, γ-Aminobutyric acid, bAla, β-alanine; Hci, Homocitrulline; Hse, Homoserine; ETA, Ethanolamine; Sar, Sarcosine; Tau, Taruine; Hyp, 4-Hydroxyproline; Cit, Citrulline

Figure 2.

Levels of amino acid ratios and of individual amino acids; diagnostic threshold set in the training set (red lines). Venn diagrams of the metabotype-positive subjects identified by each of the AA:BCAA diagnostics for AADMglutamine (A-C), AADMglycine (D-F), and AADMornithine (G-I) in the training and test sets. A, D, and G) Scatter plots of the AA:BCAA ratios used to create an AADM diagnostic test. Red points represent AADM positive subjects and black points represent AADM negative subjects. The red horizontal line is the diagnostic threshold set in the training set. B, E, and H) Scatter plots of individual amino acids used in the creation of the ratios. Red dots indicate AADM positive subjects and black points represent those that are AADM negative. C, F, and I) Venn diagram of metabotype-positive subjects identified by the three ratios used to create each AA:BCAA diagnostic. Each circle represents the subjects identified by the diagnostic threshold for a given ratio. The intersection of the Venn diagram indicates the subjects called AADM positive (red dots in scatter plots). Performance metrics above the Venn diagram represent entire study population (training and test sets).Abbreviations: AA, amino acid; BCAA, branched chain amino acid; ASD, autism spectrum disorder; TYP, typically developing; AADM, amino acid dysregulation metabotype.

Figure 3.

A) Venn diagram of the 92 AADMtotal subjects identified by each of the AADMs. At least 50% of the subjects identified by one AADM were identified by the other 2 AADMs. The AADMtotal population is comprised of 86 ASD and 6 TYP subjects. The overall prevalence of metabolic dysregulation in the CAMP ASD population is 16.7% (86 AADMtotal ASD / 516 CAMP ASD), specificity 96.3 % (158 AADM-negative TYP / 164 CAMP TYP), PPV 93.5% (86 AADMtotal ASD / 92 AADMtotal). B) PCA analysis of the metabolite ratios used in the metabolic signature of the reproducible AADMs creating the AADMtotal estimates in the CAMP study population. Black circle is the 95% confidence interval from the Hoetellings T2. Red letters are AADMtotal positive (N=92), Black letters are AADMtotal negative (N=588). A=ASD and T=TYP. Abbreviations: BCAA, branched chain amino acid; Orn, Ornithine; ASD, autism spectrum disorder; TYP, typically developing; AADM, amino acid dysregulation metabotype; CAMP, Children’s Autism Metabolome Project.