Publications by authors named "Leo A J Kluijtmans"

69 Publications

Amadori rearrangement products as potential biomarkers for inborn errors of amino-acid metabolism.

Commun Biol 2021 Mar 19;4(1):367. Epub 2021 Mar 19.

Radboud University, Institute for Molecules and Materials, FELIX Laboratory, Toernooiveld 7, Nijmegen, the Netherlands.

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.
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http://dx.doi.org/10.1038/s42003-021-01909-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7979741PMC
March 2021

Monitoring phenylalanine concentrations in the follow-up of phenylketonuria patients: An inventory of pre-analytical and analytical variation.

JIMD Rep 2021 Mar 22;58(1):70-79. Epub 2020 Nov 22.

Translational Metabolic Laboratory, Department of Laboratory Medicine Radboud University Medical Centre Nijmegen The Netherlands.

Background: Reliable measurement of phenylalanine (Phe) is a prerequisite for adequate follow-up of phenylketonuria (PKU) patients. However, previous studies have raised concerns on the intercomparability of plasma and dried blood spot (DBS) Phe results. In this study, we made an inventory of differences in (pre-)analytical methodology used for Phe determination across Dutch laboratories, and compared DBS and plasma results.

Methods: Through an online questionnaire, we assessed (pre-)analytical Phe measurement procedures of seven Dutch metabolic laboratories. To investigate the difference between plasma and DBS Phe, participating laboratories received simultaneously collected plasma-DBS sets from 23 PKU patients. In parallel, 40 sample sets of DBS spotted from either venous blood or capillary fingerprick were analyzed.

Results: Our data show that there is no consistency on standard operating procedures for Phe measurement. The association of DBS to plasma Phe concentration exhibits substantial inter-laboratory variation, ranging from a mean difference of -15.5% to +30.6% between plasma and DBS Phe concentrations. In addition, we found a mean difference of +5.8% in Phe concentration between capillary DBS and DBS prepared from venous blood.

Conclusions: The results of our study point to substantial (pre-)analytical variation in Phe measurements, implicating that bloodspot Phe results should be interpreted with caution, especially when no correction factor is applied. To minimize variation, we advocate pre-analytical standardization and analytical harmonization of Phe measurements, including consensus on application of a correction factor to adjust DBS Phe to plasma concentrations.
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http://dx.doi.org/10.1002/jmd2.12186DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7932865PMC
March 2021

Response to Biesecker et al.

Genet Med 2021 Apr 8;23(4):793-794. Epub 2021 Jan 8.

PerkinElmer Genomics, Duluth, GA, USA.

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http://dx.doi.org/10.1038/s41436-020-01055-zDOI Listing
April 2021

The role of clinical response to treatment in determining pathogenicity of genomic variants.

Genet Med 2021 Mar 22;23(3):581-585. Epub 2020 Oct 22.

PerkinElmer Genomics, Duluth, GA, USA.

Purpose: The 2015 American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines for the interpretation of sequence variants provide a framework to standardize terminology in the classification of variants uncovered through genetic testing. We aimed to assess the validity of utilizing clinical response to therapies specifically targeted to a suspected disease in clarifying variant pathogenicity.

Methods: Five families with disparate clinical presentations and different genetic diseases evaluated and treated in multiple diagnostic settings are summarized.

Results: Extended evaluations indicated possible genetic diagnoses and assigned candidate causal variants, but the cumulative clinical, biochemical, and molecular information in each instance was not completely consistent with the identified disease. Initiation of treatment specific to the suspected diagnoses in the affected individuals led to clinical improvement in all five families.

Conclusion: We propose that the effect of therapies that are specific and targeted to treatable genetic diseases embodies an in vivo physiological response and could be considered as additional criteria within the 2015 ACMG/AMP guidelines in determining genomic variant pathogenicity.
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http://dx.doi.org/10.1038/s41436-020-00996-9DOI Listing
March 2021

A newborn screening approach to diagnose 3-hydroxy-3-methylglutaryl-CoA lyase deficiency.

JIMD Rep 2020 Jul 14;54(1):79-86. Epub 2020 Apr 14.

Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacký University Olomouc Olomouc Czech Republic.

3-Hydroxy-3-methylglutaryl-coenzyme A lyase deficiency (HMGCLD) is a rare autosomal recessively inherited metabolic disorder. Patients suffer from avoidable neurologically devastating metabolic decompensations and thus would benefit from newborn screening (NBS). The diagnosis is currently made by measuring dry blood spot acylcarnitines (C5OH and C6DC) followed by urinary organic acid profiling for the differential diagnosis from several other disorders. Using untargeted metabolomics (reversed-phase UHPLC coupled to an Orbitrap Elite hybrid mass spectrometer) of plasma samples from 5 HMGCLD patients and 19 age-matched controls, we found 3-methylglutaconic acid and 3-hydroxy-3-methylglutaric acid, together with 3-hydroxyisovalerylcarnitine as the most discriminating metabolites between the groups. In order to evaluate the NBS potential of these metabolites we quantified the most discriminating metabolites from untargeted metabolomics in 23 blood spots from 4 HMGCLD patients and 55 controls by UHPLC tandem mass spectrometry. The results provide a tool for expanded NBS of HMGCLD using tandem mass spectrometry. Selected reaction monitoring transition 262/85 could be used in a first-tier NBS analysis to screen for elevated 3-hydroxyisovalerylcarnitine. In a positive case, a second-tier analysis of 3-hydroxy-3-methylglutaric acid and 3-methylglutaconic acid in a dry blood spot using UHPLC tandem mass spectrometry instruments confirms the diagnosis. In conclusion, we describe the identification of new diagnostic biomarkers for HMGCLD and their application in NBS in dry blood spots. By using second-tier testing, all patients with HMGCLD were unequivocally and correctly diagnosed.
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http://dx.doi.org/10.1002/jmd2.12118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7358667PMC
July 2020

Evaluation of 11 years of newborn screening for maple syrup urine disease in the Netherlands and a systematic review of the literature: Strategies for optimization.

JIMD Rep 2020 Jul 13;54(1):68-78. Epub 2020 May 13.

Department of Pediatrics, Division of Metabolic Disorders Amsterdam UMC, University of Amsterdam Amsterdam The Netherlands.

Maple syrup urine disease (MSUD) leads to severe neurological deterioration unless diagnosed early and treated immediately. We have evaluated the effectiveness of 11 years of MSUD newborn screening (NBS) in the Netherlands (screening >72 hours, referral if both total leucine (Xle) and valine ≥400 μmol/L blood) and have explored possibilities for improvement by combining our data with a systematic literature review and data from Collaborative Laboratory Integrated Reports (CLIR). Dutch MSUD NBS characteristics and accuracy were determined. The hypothetical referral numbers in the Dutch population of additional screening markers suggested by CLIR were calculated. In a systematic review, articles reporting NBS leucine concentrations of confirmed patients were included. Our data showed that NBS of 1 963 465 newborns identified 4 MSUD patients and led to 118 false-positive referrals (PPV 3.28%; incidence 1:491 000 newborns). In literature, leucine is the preferred NBS parameter. Total leucine (Xle) concentrations (mass-spectrometry) of 53 detected and 8 false-negative patients (sampling age within 25 hours in 3 patients) reported in literature ranged from 288 to 3376 (median 900) and 42 to 325 (median 209) μmol/L blood respectively. CLIR showed increasing Xle concentrations with sampling age and early NBS sampling and milder variant MSUD phenotypes with (nearly) normal biochemical profiles are causes of false-negative NBS results. We evaluated the effect of additional screening markers and established the Xle/phenylalanine ratio as a promising additional marker ratio for increasing the PPV, while maintaining high sensitivity in the Dutch MSUD NBS.
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http://dx.doi.org/10.1002/jmd2.12124DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7358668PMC
July 2020

metPropagate: network-guided propagation of metabolomic information for prioritization of metabolic disease genes.

NPJ Genom Med 2020 2;5:25. Epub 2020 Jul 2.

BC Children's Hospital Research Institute, Centre for Molecular Medicine and Therapeutics, University of British Columbia, Vancouver, Canada.

Many inborn errors of metabolism (IEMs) are amenable to treatment, therefore early diagnosis is imperative. Whole-exome sequencing (WES) variant prioritization coupled with phenotype-guided clinical and bioinformatics expertise is typically used to identify disease-causing variants; however, it can be challenging to identify the causal candidate gene when a large number of rare and potentially pathogenic variants are detected. Here, we present a network-based approach, metPropagate, that uses untargeted metabolomics (UM) data from a single patient and a group of controls to prioritize candidate genes in patients with suspected IEMs. We validate metPropagate on 107 patients with IEMs diagnosed in Miller et al. (2015) and 11 patients with both CNS and metabolic abnormalities. The metPropagate method ranks candidate genes by label propagation, a graph-smoothing algorithm that considers each gene's metabolic perturbation in addition to the network of interactions between neighbors. metPropagate was able to prioritize at least one causative gene in the top 20 percentile of candidate genes for 92% of patients with known IEMs. Applied to patients with suspected neurometabolic disease, metPropagate placed at least one causative gene in the top 20 percentile in 9/11 patients, and ranked the causative gene more highly than Exomiser's phenotype-based ranking in 6/11 patients. Interestingly, ranking by a weighted combination of metPropagate and Exomiser scores resulted in improved prioritization. The results of this study indicate that network-based analysis of UM data can provide an additional mode of evidence to prioritize causal genes in patients with suspected IEMs.
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http://dx.doi.org/10.1038/s41525-020-0132-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7331614PMC
July 2020

Correction to: A comparison of high‑throughput plasma NMR protocols for comparative untargeted metabolomics.

Metabolomics 2020 May 20;16(6):67. Epub 2020 May 20.

Translational Metabolic Laboratory, Department of Laboratory Medicine, Radboudumc, Geert Grooteplein Zuid 10, 6525 GA, Nijmegen, The Netherlands.

Following publication of the original article, the authors would like to correct a sentence in the paragraph "H-NMR spectra were recorded at 298 K…" under the heading "NMR experiments".
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http://dx.doi.org/10.1007/s11306-020-01688-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7645580PMC
May 2020

A comparison of high-throughput plasma NMR protocols for comparative untargeted metabolomics.

Metabolomics 2020 05 1;16(5):64. Epub 2020 May 1.

Translational Metabolic Laboratory, Department of Laboratory Medicine, Radboudumc, Geert Grooteplein Zuid 10, 6525 GA, Nijmegen, The Netherlands.

Introduction: When analyzing the human plasma metabolome with Nuclear Magnetic Resonance (NMR) spectroscopy, the Carr-Purcell-Meiboom-Gill (CPMG) experiment is commonly employed for large studies. However, this process can lead to compromised statistical analyses due to residual macromolecule signals. In addition, the utilization of Trimethylsilylpropanoic acid (TSP) as an internal standard often leads to quantification issues, and binning, as a spectral summarization step, can result in features not clearly assignable to metabolites.

Objectives: Our aim was to establish a new complete protocol for large plasma cohorts collected with the purpose of describing the comparative metabolic profile of groups of samples.

Methods: We compared the conventional CPMG approach to a novel procedure that involves diffusion NMR, using the Longitudinal Eddy-Current Delay (LED) experiment, maleic acid (MA) as the quantification reference and peak picking for spectral reduction. This comparison was carried out using the ultrafiltration method as a gold standard in a simple sample classification experiment, with Partial Least Squares-Discriminant Analysis (PLS-DA) and the resulting metabolic signatures for multivariate data analysis. In addition, the quantification capabilities of the method were evaluated.

Results: We found that the LED method applied was able to detect more metabolites than CPMG and suppress macromolecule signals more efficiently. The complete protocol was able to yield PLS-DA models with enhanced classification accuracy as well as a more reliable set of important features than the conventional CPMG approach. Assessment of the quantitative capabilities of the method resulted in good linearity, recovery and agreement with an established amino acid assay for the majority of the metabolites tested. Regarding repeatability, ~ 85% of all peaks had an adequately low coefficient of variation (< 30%) in replicate samples.

Conclusion: Overall, our comparison yielded a high-throughput untargeted plasma NMR protocol for optimized data acquisition and processing that is expected to be a valuable contribution in the field of metabolic biomarker discovery.
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http://dx.doi.org/10.1007/s11306-020-01686-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7196944PMC
May 2020

Infrared ion spectroscopy: New opportunities for small-molecule identification in mass spectrometry - A tutorial perspective.

Anal Chim Acta 2020 Jan 24;1093:1-15. Epub 2019 Oct 24.

Radboud University, Institute for Molecules and Materials, FELIX Laboratory, Toernooiveld 7, 6525 ED, Nijmegen, the Netherlands; van't Hoff Institute for Molecular Sciences, University of Amsterdam, 1098XH, Amsterdam, Science Park 908, the Netherlands. Electronic address:

Combining the individual analytical strengths of mass spectrometry and infrared spectroscopy, infrared ion spectroscopy is increasingly recognized as a powerful tool for small-molecule identification in a wide range of analytical applications. Mass spectrometry is itself a leading analytical technique for small-molecule identification on the merit of its outstanding sensitivity, selectivity and versatility. The foremost shortcoming of the technique, however, is its limited ability to directly probe molecular structure, especially when contrasted against spectroscopic techniques. In infrared ion spectroscopy, infrared vibrational spectra are recorded for mass-isolated ions and provide a signature that can be matched to reference spectra, either measured from standards or predicted using quantum-chemical calculations. Here we present an overview of the potential for this technique to develop into a versatile analytical method for identifying molecular structures in mass spectrometry-based analytical workflows. In this tutorial perspective, we introduce the reader to the technique of infrared ion spectroscopy and highlight a selection of recent experimental advances and applications in current analytical challenges, in particular in the field of untargeted metabolomics. We report on the coupling of infrared ion spectroscopy with liquid chromatography and present experiments that serve as proof-of-principle examples of strategies to address outstanding challenges.
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http://dx.doi.org/10.1016/j.aca.2019.10.043DOI Listing
January 2020

Bi-allelic GOT2 Mutations Cause a Treatable Malate-Aspartate Shuttle-Related Encephalopathy.

Am J Hum Genet 2019 09 15;105(3):534-548. Epub 2019 Aug 15.

On behalf of "United for Metabolic Diseases," 1105AZ Amsterdam, the Netherlands; Department of Laboratory Medicine, Translational Metabolic Laboratory, Radboud University Medical Centre, 6525 GA Nijmegen, the Netherlands. Electronic address:

Early-infantile encephalopathies with epilepsy are devastating conditions mandating an accurate diagnosis to guide proper management. Whole-exome sequencing was used to investigate the disease etiology in four children from independent families with intellectual disability and epilepsy, revealing bi-allelic GOT2 mutations. In-depth metabolic studies in individual 1 showed low plasma serine, hypercitrullinemia, hyperlactatemia, and hyperammonemia. The epilepsy was serine and pyridoxine responsive. Functional consequences of observed mutations were tested by measuring enzyme activity and by cell and animal models. Zebrafish and mouse models were used to validate brain developmental and functional defects and to test therapeutic strategies. GOT2 encodes the mitochondrial glutamate oxaloacetate transaminase. GOT2 enzyme activity was deficient in fibroblasts with bi-allelic mutations. GOT2, a member of the malate-aspartate shuttle, plays an essential role in the intracellular NAD(H) redox balance. De novo serine biosynthesis was impaired in fibroblasts with GOT2 mutations and GOT2-knockout HEK293 cells. Correcting the highly oxidized cytosolic NAD-redox state by pyruvate supplementation restored serine biosynthesis in GOT2-deficient cells. Knockdown of got2a in zebrafish resulted in a brain developmental defect associated with seizure-like electroencephalography spikes, which could be rescued by supplying pyridoxine in embryo water. Both pyridoxine and serine synergistically rescued embryonic developmental defects in zebrafish got2a morphants. The two treated individuals reacted favorably to their treatment. Our data provide a mechanistic basis for the biochemical abnormalities in GOT2 deficiency that may also hold for other MAS defects.
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http://dx.doi.org/10.1016/j.ajhg.2019.07.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6732527PMC
September 2019

Long-term treatment effect in cerebrotendinous xanthomatosis depends on age at treatment start.

Neurology 2019 01 7;92(2):e83-e95. Epub 2018 Dec 7.

From the Department of Neurology (B.M.L.S.), Catharina Hospital, Eindhoven; Department of Neurology (B.M.L.S., A.V.), Canisius Wilhelmina Hospital, Nijmegen; Department of Pediatrics (H.H.H.), Center for Lysosomal and Metabolic Diseases, Erasmus Medical Center-University Hospital, Rotterdam; Department of Neurology (B.P.C.v.d.W.), Donders Institute for Brain, Cognition and Behaviour (B.P.C.v.d.W., R.A.W.), and Department of Laboratory Medicine (L.A.J.K., R.A.W.), Translational Metabolic Laboratory, Radboud University Medical Center, Nijmegen; Department of Genetics (E.H.B.), University Medical Center Utrecht; Department of Internal Medicine (C.E.M.H.), Division of Endocrinology and Metabolism, Academic Medical Center, Amsterdam; Department of Internal Medicine (H.R.H.), Máxima Medical Center Eindhoven; Department of Internal Medicine (H.R.H.), Maastricht University Medical Center; and CAPHRI School for Public Health and Primary Care, Ageing and Long-Term Care (H.R.H.), Maastricht University, the Netherlands.

Objective: To evaluate the effect of chenodeoxycholic acid treatment on disease progression in cerebrotendinous xanthomatosis (CTX).

Methods: In this retrospective cohort study, we report the clinical long-term follow-up characteristics of 56 Dutch patients with CTX. Age at diagnosis was correlated with clinical characteristics and with the course of modified Rankin Scale (mRS) and Expanded Disability Status Scale (EDSS) scores at follow-up.

Results: Median follow-up time was 8 years (6 months-31.5 years). Patients diagnosed and treated before the age of 24 years had a significantly better outcome at follow-up. When considering only patients with a good treatment adherence (n = 43), neurologic symptoms, if present, disappeared in all patients who were diagnosed before the age of 24 and treated since. Furthermore, treatment prevented the development of new neurologic symptoms during follow-up. In contrast, 61% of the patients diagnosed and treated after the age of 24 showed deterioration of the neurologic symptoms, with parkinsonism as a treatment-resistant feature. There was an improvement or stabilization in favor of patients diagnosed and treated before the age of 24 compared to those treated after the age of 24: 100% vs 58% for mRS scores and 100% vs 50% for EDSS scores, respectively.

Conclusions: Treatment start at an early age can reverse and even prevent the development of neurologic symptoms in CTX. This study emphasizes the importance of early diagnosis in CTX and provides a rationale to include CTX in newborn screening programs.
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http://dx.doi.org/10.1212/WNL.0000000000006731DOI Listing
January 2019

Integration of genomics and metabolomics for prioritization of rare disease variants: a 2018 literature review.

J Inherit Metab Dis 2018 05 2;41(3):435-445. Epub 2018 May 2.

BC Children's Hospital Research Institute, Centre for Molecular Medicine and Therapeutics, University of British Columbia, Vancouver, BC, Canada.

Many inborn errors of metabolism (IEMs) are amenable to treatment; therefore, early diagnosis and treatment is imperative. Despite recent advances, the genetic basis of many metabolic phenotypes remains unknown. For discovery purposes, whole exome sequencing (WES) variant prioritization coupled with clinical and bioinformatics expertise is the primary method used to identify novel disease-causing variants; however, causation is often difficult to establish due to the number of plausible variants. Integrated analysis of untargeted metabolomics (UM) and WES or whole genome sequencing (WGS) data is a promising systematic approach for identifying disease-causing variants. In this review, we provide a literature-based overview of UM methods utilizing liquid chromatography mass spectrometry (LC-MS), and assess approaches to integrating WES/WGS and LC-MS UM data for the discovery and prioritization of variants causing IEMs. To embed this integrated -omics approach in the clinic, expansion of gene-metabolite annotations and metabolomic feature-to-metabolite mapping methods are needed.
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http://dx.doi.org/10.1007/s10545-018-0139-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5959954PMC
May 2018

Unraveling the unknown areas of the human metabolome: the role of infrared ion spectroscopy.

J Inherit Metab Dis 2018 05 19;41(3):367-377. Epub 2018 Mar 19.

Radboud University, Institute for Molecules and Materials, FELIX Laboratory, Toernooiveld 7c, 6525ED, Nijmegen, The Netherlands.

The identification of molecular biomarkers is critical for diagnosing and treating patients and for establishing a fundamental understanding of the pathophysiology and underlying biochemistry of inborn errors of metabolism. Currently, liquid chromatography/high-resolution mass spectrometry and nuclear magnetic resonance spectroscopy are the principle methods used for biomarker research and for structural elucidation of small molecules in patient body fluids. While both are powerful techniques, several limitations exist that often make the identification of unknown compounds challenging. Here, we describe how infrared ion spectroscopy has the potential to be a valuable orthogonal technique that provides highly-specific molecular structure information while maintaining ultra-high sensitivity. Here, we characterize and distinguish two well-known biomarkers of inborn errors of metabolism, glutaric acid for glutaric aciduria and ethylmalonic acid for short-chain acyl-CoA dehydrogenase deficiency, using infrared ion spectroscopy. In contrast to tandem mass spectra, in which ion fragments can hardly be predicted, we show that the prediction of an IR spectrum allows reference-free identification in the case that standard compounds are either commercially or synthetically unavailable. Finally, we illustrate how functional group information can be obtained from an IR spectrum for an unknown and how this is valuable information to, for example, narrow down a list of candidate structures resulting from a database query. Early diagnosis in inborn errors of metabolism is crucial for enabling treatment and depends on the identification of biomarkers specific for the disorder. Infrared ion spectroscopy has the potential to play a pivotal role in the identification of challenging biomarkers.
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http://dx.doi.org/10.1007/s10545-018-0161-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5959965PMC
May 2018

Next-generation metabolic screening: targeted and untargeted metabolomics for the diagnosis of inborn errors of metabolism in individual patients.

J Inherit Metab Dis 2018 05 16;41(3):337-353. Epub 2018 Feb 16.

Department of Laboratory Medicine, Translational Metabolic Laboratory (TML), Radboud University Medical Center, Geert Groote Plein Zuid 10, 6525, GA, Nijmegen, The Netherlands.

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.
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http://dx.doi.org/10.1007/s10545-017-0131-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5959972PMC
May 2018

Fast and accurate quantitative organic acid analysis with LC-QTOF/MS facilitates screening of patients for inborn errors of metabolism.

J Inherit Metab Dis 2018 05 12;41(3):415-424. Epub 2018 Feb 12.

Department of Clinical Genetics, Maastricht University Medical Centre, Maastricht, The Netherlands.

Since organic acid analysis in urine with gaschromatography-mass spectrometry (GC-MS) is a time-consuming technique, we developed a new liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF/MS) method to replace the classical analysis for diagnosis of inborn errors of metabolism (IEM). Sample preparation is simple and experimental time short. Targeted mass extraction and automatic calculation of z-scores generated profiles characteristic for the IEMs in our panel consisting of 71 biomarkers for defects in amino acids, neurotransmitters, fatty acids, purine, and pyrimidine metabolism as well as other disorders. In addition, four medication-related metabolites were included in the panel. The method was validated to meet Dutch NEN-EN-ISO 15189 standards. Cross validation of 24 organic acids from 28 urine samples of the ERNDIM scheme showed superiority of the UPLC-QTOF/MS method over the GC-MS method. We applied our method to 99 patient urine samples with 32 different IEMs, and 88 control samples. All IEMs were unambiguously established/diagnosed using this new QTOF method by evaluation of the panel of 71 biomarkers. In conclusion, we present a LC-QTOF/MS method for fast and accurate quantitative organic acid analysis which facilitates screening of patients for IEMs. Extension of the panel of metabolites is easy which makes this application a promising technique in metabolic diagnostics/laboratories.
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http://dx.doi.org/10.1007/s10545-017-0129-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5959959PMC
May 2018

The genotypic and phenotypic spectrum of MTO1 deficiency.

Mol Genet Metab 2018 01 15;123(1):28-42. Epub 2017 Nov 15.

Department of Pediatrics, University of British Columbia, Vancouver, BC, Canada; Centre for Molecular Medicine and Therapeutics, Vancouver, BC, Canada; BC Children's Hospital Research Institute, University of British Columbia, Vancouver, Canada; Departments of Pediatrics and Clinical Genetics, Academic Medical Center, Amsterdam, The Netherlands. Electronic address:

Background: Mitochondrial diseases, a group of multi-systemic disorders often characterized by tissue-specific phenotypes, are usually progressive and fatal disorders resulting from defects in oxidative phosphorylation. MTO1 (Mitochondrial tRNA Translation Optimization 1), an evolutionarily conserved protein expressed in high-energy demand tissues has been linked to human early-onset combined oxidative phosphorylation deficiency associated with hypertrophic cardiomyopathy, often referred to as combined oxidative phosphorylation deficiency-10 (COXPD10).

Material And Methods: Thirty five cases of MTO1 deficiency were identified and reviewed through international collaboration. The cases of two female siblings, who presented at 1 and 2years of life with seizures, global developmental delay, hypotonia, elevated lactate and complex I and IV deficiency on muscle biopsy but without cardiomyopathy, are presented in detail.

Results: For the description of phenotypic features, the denominator varies as the literature was insufficient to allow for complete ascertainment of all data for the 35 cases. An extensive review of all known MTO1 deficiency cases revealed the most common features at presentation to be lactic acidosis (LA) (21/34; 62% cases) and hypertrophic cardiomyopathy (15/34; 44% cases). Eventually lactic acidosis and hypertrophic cardiomyopathy are described in 35/35 (100%) and 27/34 (79%) of patients with MTO1 deficiency, respectively; with global developmental delay/intellectual disability present in 28/29 (97%), feeding difficulties in 17/35 (49%), failure to thrive in 12/35 (34%), seizures in 12/35 (34%), optic atrophy in 11/21 (52%) and ataxia in 7/34 (21%). There are 19 different pathogenic MTO1 variants identified in these 35 cases: one splice-site, 3 frameshift and 15 missense variants. None have bi-allelic variants that completely inactivate MTO1; however, patients where one variant is truncating (i.e. frameshift) while the second one is a missense appear to have a more severe, even fatal, phenotype. These data suggest that complete loss of MTO1 is not viable. A ketogenic diet may have exerted a favourable effect on seizures in 2/5 patients.

Conclusion: MTO1 deficiency is lethal in some but not all cases, and a genotype-phenotype relation is suggested. Aside from lactic acidosis and cardiomyopathy, developmental delay and other phenotypic features affecting multiple organ systems are often present in these patients, suggesting a broader spectrum than hitherto reported. The diagnosis should be suspected on clinical features and the presence of markers of mitochondrial dysfunction in body fluids, especially low residual complex I, III and IV activity in muscle. Molecular confirmation is required and targeted genomic testing may be the most efficient approach. Although subjective clinical improvement was observed in a small number of patients on therapies such as ketogenic diet and dichloroacetate, no evidence-based effective therapy exists.
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http://dx.doi.org/10.1016/j.ymgme.2017.11.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5780301PMC
January 2018

Structural elucidation of novel biomarkers of known metabolic disorders based on multistage fragmentation mass spectra.

J Inherit Metab Dis 2018 05 14;41(3):407-414. Epub 2017 Nov 14.

Translational Metabolic Laboratory - 830 TML, Department of Laboratory Medicine, Radboud University Medical Centre, Geert Grooteplein 10, 6525, GA, Nijmegen, the Netherlands.

Specific diagnostic markers are the key to effective diagnosis and treatment of inborn errors of metabolism (IEM). Untargeted metabolomics allows for the identification of potential novel diagnostic biomarkers. Current separation techniques coupled to high-resolution mass spectrometry provide a powerful tool for structural elucidation of unknown compounds in complex biological matrices. This is a proof-of-concept study testing this methodology to determine the molecular structure of as yet uncharacterized m/z signals that were significantly increased in plasma samples from patients with phenylketonuria and 3-hydroxy-3-methylglutaryl-CoA lyase deficiency. A hybrid linear ion trap-orbitrap high resolution mass spectrometer, capable of multistage fragmentation, was used to acquire accurate masses and product ion spectra of the uncharacterized m/z signals. In order to determine the molecular structures, spectral databases were searched and fragmentation prediction software was used. This approach enabled structural elucidation of novel compounds potentially useful as biomarkers in diagnostics and follow-up of IEM patients. Two new conjugates, glutamyl-glutamyl-phenylalanine and phenylalanine-hexose, were identified in plasma of phenylketonuria patients. These novel markers showed high inter-patient variation and did not correlate to phenylalanine levels, illustrating their potential added value for follow-up. As novel biomarkers for 3-hydroxy-3-methylglutaryl-CoA lyase deficiency, three positional isomers of 3-methylglutaconyl carnitine could be detected in patient plasma. Our results highlight the applicability of current accurate mass multistage fragmentation techniques for structural elucidation of unknown metabolites in human biofluids, offering an unprecedented opportunity to gain further biochemical insights in known inborn errors of metabolism by enabling high confidence identification of novel biomarkers.
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http://dx.doi.org/10.1007/s10545-017-0109-4DOI Listing
May 2018

Autism spectrum disorder: an early and frequent feature in cerebrotendinous xanthomatosis.

J Inherit Metab Dis 2018 07 11;41(4):641-646. Epub 2017 Sep 11.

Department of Neurology, Canisius Wilhelmina Hospital, PO Box 9015, 6500GS, Nijmegen, The Netherlands.

Background: Cerebrotendinous xanthomatosis (CTX) is an autosomal recessively inherited inborn error of metabolism (IEM) due to mutations in the CYP27A1 gene. The clinical picture ranges from being nearly asymptomatic in early childhood, up to severe disability at adult age. Infantile-onset diarrhea and juvenile-onset cataract are the earliest symptoms in childhood. In the current study, we evaluated the presence of autism spectrum disorder (ASD) in a large cohort of CTX patients.

Methods: We performed a retrospective patient file study in 77 genetically confirmed Dutch CTX patients to determine the frequency of ASD. In addition, we compared plasma cholestanol levels in CTX patients with and without a diagnosis of ASD and tried to establish a relation between CYP27A1 genotype and ASD.

Results: In our CTX cohort, 10 patients (13%; nine pediatric and one adult) with ASD were identified. At the time of diagnosis of ASD, most patients only exhibited symptoms of diarrhea and/or intellectual disability without signs of cataract or neurological symptoms. No correlation was found between the presence of ASD and the level of cholestanol or CYP27A1 genotype. The behavioral problems stabilized or improved after treatment initiation with chenodeoxycholic acid (CDCA) in all pediatric patients.

Conclusions: We conclude that ASD is an early and probably underestimated frequent feature in CTX. Metabolic screening for CTX should be performed in patients with ASD when accompanied by diarrhea, intellectual disability, juvenile cataract, and/or neurological involvement. Early recognition allows for earlier initiation of specific treatment and will improve clinical outcome. Our results add CTX to the list of treatable IEMs associated with ASD.
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http://dx.doi.org/10.1007/s10545-017-0086-7DOI Listing
July 2018

Molecular identification in metabolomics using infrared ion spectroscopy.

Sci Rep 2017 06 13;7(1):3363. Epub 2017 Jun 13.

Radboud University, Institute for Molecules and Materials, FELIX Laboratory, Toernooiveld 7c, 6525ED, Nijmegen, The Netherlands.

Small molecule identification is a continually expanding field of research and represents the core challenge in various areas of (bio)analytical science, including metabolomics. Here, we unequivocally differentiate enantiomeric N-acetylhexosamines in body fluids using infrared ion spectroscopy, providing orthogonal identification of molecular structure unavailable by standard liquid chromatography/high-resolution tandem mass spectrometry. These results illustrate the potential of infrared ion spectroscopy for the identification of small molecules from complex mixtures.
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http://dx.doi.org/10.1038/s41598-017-03387-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5469762PMC
June 2017

Mild orotic aciduria in UMPS heterozygotes: a metabolic finding without clinical consequences.

J Inherit Metab Dis 2017 05 15;40(3):423-431. Epub 2017 Feb 15.

Department of Genetics, Kaiser Permanente, Los Angeles, CA, USA.

Background: Elevated urinary excretion of orotic acid is associated with treatable disorders of the urea cycle and pyrimidine metabolism. Establishing the correct and timely diagnosis in a patient with orotic aciduria is key to effective treatment. Uridine monophosphate synthase is involved in de novo pyrimidine synthesis. Uridine monophosphate synthase deficiency (or hereditary orotic aciduria), due to biallelic mutations in UMPS, is a rare condition presenting with megaloblastic anemia in the first months of life. If not treated with the pyrimidine precursor uridine, neutropenia, failure to thrive, growth retardation, developmental delay, and intellectual disability may ensue.

Methods And Results: We identified mild and isolated orotic aciduria in 11 unrelated individuals with diverse clinical signs and symptoms, the most common denominator being intellectual disability/developmental delay. Of note, none had blood count abnormalities, relevant hyperammonemia or altered plasma amino acid profile. All individuals were found to have heterozygous alterations in UMPS. Four of these variants were predicted to be null alleles with complete loss of function. The remaining variants were missense changes and predicted to be damaging to the normal encoded protein. Interestingly, family screening revealed heterozygous UMPS variants in combination with mild orotic aciduria in 19 clinically asymptomatic family members.

Conclusions: We therefore conclude that heterozygous UMPS-mutations can lead to mild and isolated orotic aciduria without clinical consequence. Partial UMPS-deficiency should be included in the differential diagnosis of mild orotic aciduria. The discovery of heterozygotes manifesting clinical symptoms such as hypotonia and developmental delay are likely due to ascertainment bias.
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http://dx.doi.org/10.1007/s10545-017-0015-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5393157PMC
May 2017

Reliable Diagnosis of Carnitine Palmitoyltransferase Type IA Deficiency by Analysis of Plasma Acylcarnitine Profiles.

JIMD Rep 2017 14;32:33-39. Epub 2016 Jun 14.

Department of Laboratory Medicine, Laboratory of Metabolic Diseases, University of Groningen, University Medical Center Groningen, Room Y2.125, HPA EA60, 30.001, NL-9700 RB, Groningen, The Netherlands.

Background: Carnitine palmitoyltransferase IA (CPT-IA) deficiency is an inherited disorder of the carnitine cycle (MIM #255120). Patients affected by this deficiency might be missed easily because of lack of specific and sensitive biochemical markers. In this study, sensitivity and specificity of plasma free carnitine (C0) and long-chain acylcarnitines (lc-ac: C16:0-, C16:1-, C18:0-, C18:1- and C18:2-ac) was evaluated, including the sum of lc-ac (∑lc-ac) and the molar ratios C0/(C16:0-ac+C18:0-ac) and C0/∑lc-ac.

Methods: Nine plasma acylcarnitine profiles of 4 CPT-IA deficient patients were compared with profiles of 2,190 subjects suspected of or diagnosed with an inherited disorder of metabolism. Age-dependent reference values were calculated based on the patient population without a definite diagnosis of an inborn error of metabolism (n = 1,600). Sensitivity, specificity, and Receiver Operating Characteristic (ROC) curves were calculated based on samples of the whole patient population.

Results: Concentrations of C0 in plasma were normal in all CPT-IA deficient patient samples. ROC analyses showed highest diagnostic values for C18:0-ac, C18:1-ac, and ∑lc-ac (AUC 1.000) and lowest for C0 (AUC 0.738). Combining two markers, i.e., a plasma C18:1-ac concentration <0.05 μmol/L and a molar ratio of C0/(C16:0-ac+C18:0-ac) >587, specificity to diagnose CPT-IA deficiency increased to 99.3% compared with either C18:1-ac (97.4%) or C0/(C16:0-ac+C18:0-ac) (96.9%) alone, all at a sensitivity of 100%.

Conclusions: Combination of a low concentration of C18:1-ac with a high molar ratio of C0/(C16:0-ac+C18:0-ac) ratio in plasma has high diagnostic value for CPT-IA deficiency. Patients with a clinical suspicion of CPT-IA deficiency can be diagnosed with this test combination.
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http://dx.doi.org/10.1007/8904_2016_564DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5362552PMC
June 2016

Hydrogen cyanide emission in the lung by Staphylococcus aureus.

Eur Respir J 2016 08 26;48(2):577-9. Epub 2016 May 26.

Dept of Paediatrics, Amalia Children's Hospital, Radboud University Medical Center, Nijmegen, The Netherlands Dept of Paediatrics, Canisius Wilhelmina Hospital, Nijmegen, The Netherlands Both authors contributed equally.

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http://dx.doi.org/10.1183/13993003.02093-2015DOI Listing
August 2016

NANS-mediated synthesis of sialic acid is required for brain and skeletal development.

Nat Genet 2016 07 23;48(7):777-84. Epub 2016 May 23.

Centre for Molecular Diseases, Lausanne University Hospital (CHUV), University of Lausanne, Lausanne, Switzerland.

We identified biallelic mutations in NANS, the gene encoding the synthase for N-acetylneuraminic acid (NeuNAc; sialic acid), in nine individuals with infantile-onset severe developmental delay and skeletal dysplasia. Patient body fluids showed an elevation in N-acetyl-D-mannosamine levels, and patient-derived fibroblasts had reduced NANS activity and were unable to incorporate sialic acid precursors into sialylated glycoproteins. Knockdown of nansa in zebrafish embryos resulted in abnormal skeletal development, and exogenously added sialic acid partially rescued the skeletal phenotype. Thus, NANS-mediated synthesis of sialic acid is required for early brain development and skeletal growth. Normal sialylation of plasma proteins was observed in spite of NANS deficiency. Exploration of endogenous synthesis, nutritional absorption, and rescue pathways for sialic acid in different tissues and developmental phases is warranted to design therapeutic strategies to counteract NANS deficiency and to shed light on sialic acid metabolism and its implications for human nutrition.
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http://dx.doi.org/10.1038/ng.3578DOI Listing
July 2016

Hyperammonemia due to Adult-Onset N-Acetylglutamate Synthase Deficiency.

JIMD Rep 2017 5;31:95-99. Epub 2016 May 5.

Department of Internal Medicine, Radboud University Medical Center, 9101, 6500 HB, Nijmegen, The Netherlands.

A 59-year-old woman, with a medical history of intellectual disability after perinatal asphyxia, was admitted because of coma due to hyperammonemia after she was treated for a fracture of the pelvis. The ammonia level was 280 μM. Acquired disorders as explanation for the hyperammonemia were excluded. Metabolic investigations showed an elevated glutamine and alanine and low citrulline, suspect for a urea cycle defect (UCD). Orotic acid could not be demonstrated in urine. DNA investigations were negative for mutations or deletions in the OTC and CPS1 gene, but revealed a homozygous c.603G>C mutation in exon 2 of the N-acetylglutamate synthase (NAGS) gene (NM_153006.2:c.603G>C), which mandates p.Lys201Asn. This is a novel mutation in the NAGS gene.After the diagnosis of NAGS deficiency was made carbamylglutamate was started in a low dose. In combination with mild protein restriction the ammonia level decreased to 26 μM.This is one of the first patients in literature in whom the diagnosis of a UCD is made at such an advanced age. It is important for the adult physician to consider a metabolic disorder at every age.
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http://dx.doi.org/10.1007/8904_2016_565DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5272844PMC
May 2016

Microbial Metabolism Shifts Towards an Adverse Profile with Supplementary Iron in the TIM-2 In vitro Model of the Human Colon.

Front Microbiol 2015 6;6:1481. Epub 2016 Jan 6.

Department of Laboratory Medicine - Translational Metabolic Laboratory, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center Nijmegen, Netherlands.

Oral iron administration in African children can increase the risk for infections. However, it remains unclear to what extent supplementary iron affects the intestinal microbiome. We here explored the impact of iron preparations on microbial growth and metabolism in the well-controlled TNO's in vitro model of the large intestine (TIM-2). The model was inoculated with a human microbiota, without supplementary iron, or with 50 or 250 μmol/L ferrous sulfate, 50 or 250 μmol/L ferric citrate, or 50 μmol/L hemin. High resolution responses of the microbiota were examined by 16S rDNA pyrosequencing, microarray analysis, and metagenomic sequencing. The metabolome was assessed by fatty acid quantification, gas chromatography-mass spectrometry (GC-MS), and (1)H-NMR spectroscopy. Cultured intestinal epithelial Caco-2 cells were used to assess fecal water toxicity. Microbiome analysis showed, among others, that supplementary iron induced decreased levels of Bifidobacteriaceae and Lactobacillaceae, while it caused higher levels of Roseburia and Prevotella. Metagenomic analyses showed an enrichment of microbial motility-chemotaxis systems, while the metabolome markedly changed from a saccharolytic to a proteolytic profile in response to iron. Branched chain fatty acids and ammonia levels increased significantly, in particular with ferrous sulfate. Importantly, the metabolite-containing effluent from iron-rich conditions showed increased cytotoxicity to Caco-2 cells. Our explorations indicate that in the absence of host influences, iron induces a more hostile environment characterized by a reduction of microbes that are generally beneficial, and increased levels of bacterial metabolites that can impair the barrier function of a cultured intestinal epithelial monolayer.
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http://dx.doi.org/10.3389/fmicb.2015.01481DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4701948PMC
January 2016

Expanding the phenotype in aminoacylase 1 (ACY1) deficiency: characterization of the molecular defect in a 63-year-old woman with generalized dystonia.

Metab Brain Dis 2016 06 19;31(3):587-92. Epub 2015 Dec 19.

Department of Laboratory Medicine, Translational Metabolic Laboratory, Radboud University Medical Center, Nijmegen, The Netherlands.

Aminoacylase 1 (ACY1) deficiency is an organic aciduria due to mutations in the ACY1 gene. It is considered much underdiagnosed. Most individuals known to be affected by ACY1 deficiency have presented with neurologic symptoms. We report here a cognitively normal 63-year-old woman who around the age of 12 years had developed dystonic symptoms that gradually evolved into generalized dystonia. Extensive investigations, including metabolic diagnostics and diagnostic exome sequencing, were performed to elucidate the cause of dystonia. Findings were only compatible with a diagnosis of ACY1 deficiency: the urinary metabolite pattern with N-acetylated amino acids was characteristic, there was decreased ACY1 activity in immortalized lymphocytes, and two compound heterozygous ACY1 mutations were detected, one well-characterized c.1057C>T (p.Arg353Cys) and the other novel c.325A>G (p.Arg109Gly). Expression analysis in HEK293 cells revealed high residual activity of the enzyme with the latter mutation. However, following co-transfection of cells with stable expression of the c.1057C>T variant with either wild-type ACY1 or the c.325A>G mutant, only the wild-type enhanced ACY1 activity and ACY1 presence in the Western blot, suggesting an inhibiting interference between the two variants. Our report extends the clinical spectrum of ACY1 deficiency to include dystonia and indicates that screening for organic acidurias deserves consideration in patients with unexplained generalized dystonia.
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http://dx.doi.org/10.1007/s11011-015-9778-6DOI Listing
June 2016

Urinary excretion of polyols and sugars in children with chronic kidney disease.

Pediatr Nephrol 2015 Sep 9;30(9):1537-40. Epub 2015 Jun 9.

Department of Pediatric Nephrology & Growth and Regeneration, University Hospitals Leuven & Katholieke Universiteit Leuven, Herestraat 49, 3000, Leuven, Belgium,

Background: The urinary concentrations of monosaccharides and polyols are used for diagnosing inborn errors of metabolism and renal tubular disorders. Reference values are age-related and depend on the method of detection. However, the influence of the renal function is often still neglected. In this study we examined the urinary excretion of monosaccharides and polyols in children with various degrees of chronic kidney disease (CKD), but with no known metabolic or renal tubular disorders.

Case Diagnosis/treatment: In 25 patients with CKD stage 1-5, urinary concentrations of 18 monosaccharides and polyols were measured by gas chromatography-mass spectrometry (GC-MS) in random urinary samples and were compared with age-related reference values. Serum creatinine was measured at the time of the urine sample, and the height-independent estimated glomerular filtration rate (eGFR-Pottel) was calculated. Urinary excretions of monosaccharides and polyols were above the reference values in 8-88% of all patients. A significant difference between CKD stage 1-2 compared with CKD stage 3-5 was found for allose, arabitol and sorbitol (p < 0.05) and for arabinose, fucose, myoinositol, ribitol, xylitol, and xylose (p < 0.01).

Conclusions: We show that the excretion of polyols and sugars depends on eGFR, which warrants a cautious interpretation of the results in patients with CKD.
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http://dx.doi.org/10.1007/s00467-015-3135-1DOI Listing
September 2015

Leucine Loading Test is Only Discriminative for 3-Methylglutaconic Aciduria Due to AUH Defect.

JIMD Rep 2014 23;16:1-6. Epub 2014 Apr 23.

Department of Pediatrics, Nijmegen Centre for Mitochondrial Disorders (NCMD), Amalia Children's Hospital, Radboud University Medical Centre, 9101, 6500 HB, Nijmegen, The Netherlands,

Currently, six inborn errors of metabolism with 3-methylglutaconic aciduria as discriminative feature are known. The "Primary 3-methylglutaconic aciduria," 3-methylglutaconyl-CoA hydratase deficiency or AUH defect, is a disorder of leucine catabolism. For all other subtypes, also denoted "Secondary 3-methylglutaconic acidurias" (TAZ defect or Barth syndrome, SERAC1 defect or MEGDEL syndrome, OPA3 defect or Costeff syndrome, DNAJC19 defect or DCMA syndrome, TMEM70 defect, "not otherwise specified (NOS) 3-MGA-uria"), the origin of 3-methylglutaconic aciduria remains enigmatic but is hypothesized to be independent from leucine catabolism. Here we show the results of leucine loading test in 21 patients with different inborn errors of metabolism who present with 3-methylglutaconic aciduria. After leucine loading urinary 3-methylglutaconic acid levels increased only in the patients with an AUH defect. This strongly supports the hypothesis that 3-methylglutaconic aciduria is independent from leucine breakdown in other inborn errors of metabolism with 3-methylglutaconic aciduria and also provides a simple test to discriminate between primary and secondary 3-methylglutaconic aciduria in regular patient care.
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http://dx.doi.org/10.1007/8904_2014_309DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4221300PMC
November 2014