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We used next-generation metabolic screening to identify new biomarkers for improved diagnosis and pathophysiological understanding of glucose transporter type 1 deficiency syndrome (GLUT1DS), comparing metabolic cerebrospinal fluid (CSF) profiles from 12 patients to those of 116 controls. This confirmed decreased CSF glucose and lactate levels in patients with GLUT1DS and increased glutamine at group level. We identified three novel biomarkers significantly decreased in patients, namely gluconic + galactonic acid, xylose-α1-3- glucose, and xylose-α1-3-xylose-α1-3- glucose, of which the latter two have not previously been identified in body fluids. CSF concentrations of gluconic + galactonic acid may be reduced as these metabolites could serve as alternative substrates for the pentose phosphate pathway. Xylose-α1-3-glucose and xylose-α1-3- xylose-α1-3-glucose may originate from glycosylated proteins ; their decreased levels are hypothetically the consequence of insufficient glucose, one of two substrates for O- glucosylation. Since many proteins are O- glucosylated, this deficiency may affect cellular processes and thus contribute to GLUT1DS pathophysiology. The novel CSF biomarkers have the potential to improve the biochemical diagnosis of GLUT1DS. Our findings imply that brain glucose deficiency in GLUT1DS may cause disruptions at the cellular level that go beyond energy metabolism, underlining the importance of developing treatment strategies that directly target cerebral glucose uptake.

The identification of disease biomarkers plays a crucial role in developing diagnostic strategies for inborn errors of metabolism and understanding their pathophysiology. A primary metabolite that accumulates in the inborn error phenylketonuria is phenylalanine, however its levels do not always directly correlate with clinical outcomes. Here we combine infrared ion spectroscopy and NMR spectroscopy to identify the Phe-glucose Amadori rearrangement product as a biomarker for phenylketonuria. Additionally, we find analogous amino acid-glucose metabolites formed in the body fluids of patients accumulating methionine, lysine, proline and citrulline. Amadori rearrangement products are well-known intermediates in the formation of advanced glycation end-products and have been associated with the pathophysiology of diabetes mellitus and ageing, but are now shown to also form under conditions of aminoacidemia. They represent a general class of metabolites for inborn errors of amino acid metabolism that show potential as biomarkers and may provide further insight in disease pathophysiology., Rianne van Outersterp et al. combine mass spectrometry, NMR, and infrared ion spectroscopy to identify amino acid-hexose conjugates in the blood plasma from patients with metabolic disorders such as phenylketonuria (PKU). These conjugates, or Amadori rearrangement products, are generally not detectable in blood samples from unaffected individuals, and may therefore represent disease biomarkers.

Timely diagnosis is essential for patients with neurometabolic disorders to enable targeted treatment. Next‐Generation Metabolic Screening (NGMS) allows for simultaneous screening of multiple diseases and yields a holistic view of disturbed metabolic pathways. We applied this technique to define a cerebrospinal fluid (CSF) reference metabolome and validated our approach with patients with known neurometabolic disorders. Samples were measured using ultra‐high‐performance liquid chromatography‐quadrupole time‐of‐flight mass spectrometry followed by (un)targeted analysis. For the reference metabolome, CSF samples from patients with normal general chemistry results and no neurometabolic diagnosis were selected and grouped based on sex and age (0‐2/2‐5/5‐10/10‐15 years). We checked the levels of known biomarkers in CSF from seven patients with five different neurometabolic disorders to confirm the suitability of our method for diagnosis. Untargeted analysis of 87 control CSF samples yielded 8036 features for semiquantitative analysis. No sex differences were found, but 1782 features (22%) were different between age groups (q

The implementation of whole-exome sequencing in clinical diagnostics has generated a need for functional evaluation of genetic variants. In the field of inborn errors of metabolism (IEM), a diverse spectrum of targeted biochemical assays is employed to analyze a limited amount of metabolites. We now present a single-platform, high-resolution liquid chromatography quadrupole time of flight (LC-QTOF) method that can be applied for holistic metabolic profiling in plasma of individual IEM-suspected patients. This method, which we termed “next-generation metabolic screening” (NGMS), can detect >10,000 features in each sample. In the NGMS workflow, features identified in patient and control samples are aligned using the “various forms of chromatography mass spectrometry (XCMS)” software package. Subsequently, all features are annotated using the Human Metabolome Database, and statistical testing is performed to identify significantly perturbed metabolite concentrations in a patient sample compared with controls. We propose three main modalities to analyze complex, untargeted metabolomics data. First, a targeted evaluation can be done based on identified genetic variants of uncertain significance in metabolic pathways. Second, we developed a panel of IEM-related metabolites to filter untargeted metabolomics data. Based on this IEM-panel approach, we provided the correct diagnosis for 42 of 46 IEMs. As a last modality, metabolomics data can be analyzed in an untargeted setting, which we term “open the metabolome” analysis. This approach identifies potential novel biomarkers in known IEMs and leads to identification of biomarkers for as yet unknown IEMs. We are convinced that NGMS is the way forward in laboratory diagnostics of IEMs. Electronic supplementary material The online version of this article (10.1007/s10545-017-0131-6) contains supplementary material, which is available to authorized users.