Introduction: Prognosis of neuroblastoma patients is very diverse, indicating the need for more accurate prognostic parameters. The excretion of catecholamine metabolites by most neuroblastomas is used for diagnostic purposes, but their correlation with prognosis has hardly been investigated. Therefore, we performed an in-depth analysis of a panel of elevated urinary catecholamine metabolites at diagnosis and their correlation with prognosis. Patients and methods: Retrospective study of eight urinary catecholamine metabolites in a test (n = 96) and validation (n = 205) cohort of patients with neuroblastoma (all stages) at diagnosis. Results: Multivariate analyses, including risk factors such as stage and MYCN amplification, revealed that 3-methoxytyramine (3MT) was an independent risk factor for event-free survival (EFS) and overall survival (OS). Furthermore, only 3MT appeared to be an independent risk factor for both EFS and OS in high-risk patients, which was independent of modern high-risk therapy and immunotherapy. Among high-risk patients, those with elevated 3MT and older than 18 months had an extremely poor prognosis compared to patients with non-elevated 3MT and younger than 18 months (5-year EFS of 14.3% +/- 4% and 66.7% +/- 18%, respectively, p = 0.001; 5-year OS of 21.8% +/- 5% and 87.5% +/- 12%, respectively, p < 0.001). Conclusions: Elevated 3MT at diagnosis was associated with high-risk disease and poor prognosis. For high-risk patients, elevated 3MT at diagnosis was the only significant risk factor for EFS and OS. 3MT was also able to identify subgroups of high-risk patients with favourable and extremely poor prognosis. (C) 2017 Elsevier Ltd. All rights reserved.

BackgroundPhenylketonuria (PKU), a genetic metabolic disorder that is characterized by the inability to convert phenylalanine to tyrosine, leads to severe intellectual disability and other cerebral complications if left untreated. Dietary treatment, initiated soon after birth, prevents most brain-related complications. A leading hypothesis postulates that a shortage of brain monoamines may be associated with neurocognitive deficits that are observable even in early-treated PKU. However, there is a paucity of evidence as yet for this hypothesis.MethodsWe therefore assessedin vivostriatal dopamine D2/3receptor (D2/3R) availability and plasma monoamine metabolite levels together with measures of impulsivity and executive functioning in 18 adults with PKU and average intellect (31.2 ± 7.4 years, nine females), most of whom were early and continuously treated. Comparison data from 12 healthy controls that did not differ in gender and age were available.ResultsMean D2/3R availability was significantly higher (13%;p= 0.032) in the PKU group (n= 15) than in the controls, which may reflect reduced synaptic brain dopamine levels in PKU. The PKU group had lower plasma levels of homovanillic acid (p< 0.001) and 3-methoxy-4-hydroxy-phenylglycol (p< 0.0001), the predominant metabolites of dopamine and norepinephrine, respectively. Self-reported impulsivity levels were significantly higher in the PKU group compared with healthy controls (p= 0.033). Within the PKU group, D2/3R availability showed a positive correlation with both impulsivity (r= 0.72,p= 0.003) and the error rate during a cognitive flexibility task (r= 0.59,p= 0.020).ConclusionsThese findings provide further support for the hypothesis that executive functioning deficits in treated adult PKU may be associated with cerebral dopamine deficiency.

β-Ureidopropionase deficiency (OMIM #613161) is a rare autosomal recessive inborn error of metabolism due to mutations in the UPB1 gene, which encodes the third enzyme involved in the pyrimidine degradation pathway. A total of 28 cases have been reported, mainly presenting with seizures, microcephaly, and intellectual disabilities. However, 11 of them were asymptomatic cases (Nakajima et al., J Inherit Metab Dis 37(5):801-812, 2014). We report on a 9-year-old female presenting with intractable epilepsy, microcephaly, and global developmental delay. She was homozygous for p.R326Q (c.977G>A) and heterozygous for p.G31S (c.91G>A) in the UPB1 gene, detected by targeted next-generation sequencing test and subsequently confirmed by biochemical analysis of urine, plasma, and cerebrospinal fluid (CSF) using reversed-phase HPLC, combined with electrospray tandem mass spectrometry. We report a first Korean female case with β-ureidopropionase deficiency.

A patient with hematuria was shown to have thymine-uraciluria. The dihydropyrimidine dehydrogenase (DPD) activity in peripheral blood mononuclear cells was 0.16 nmol/mg/h; controls: 9.9 +/- 2.8 nmol/mg/h. Analysis of DPYD showed that the patient was compound heterozygous for the novel mutations 237C > A (C79X) in exon 4 and 704G > A (R235Q) in exon 7. The nonsense mutation (C79X) leads to premature termination of translation and thus to a non-functional protein. Analysis of the crystal structure of pig DPD suggested that the R235Q mutation might interfere with the binding of FAD and the electron flow between the NADPH and the pyrimidine substrate site of DPD.

Mutations in the N-linked glycosylation pathway cause rare autosomal recessive defects known as Congenital Disorders of Glycosylation (CDG). A previously reported mutation in the Conserved Oligomeric Golgi complex gene, COG7, defined a new subtype of CDG in a Tunisian family. The mutation disrupted the hetero-octomeric COG complex and altered both N- and O-linked glycosylation. Here we present clinical and biochemical data from a second family with the same mutation.

Dihydropyrimidine dehydrogenase (DPD) constitutes the first step of the pyrimidine degradation pathway in which the pyrimidine bases uracil and thymine are catabolised to beta-alanine and beta-aminoisobutyric acid (beta-AIB), respectively. The mean concentration of beta-AIB was approximately 5- to 8-fold lower in urine of patients with a DPD deficiency, when compared to age-matched controls. Comparable levels of 8-hydroxydeoxyguanosine (8-OHdG) were present in urine from controls and DPD patients at the age <2 year. In contrast, slightly elevated levels of 8-OHdG were detected in urine from DPD patients with an age >2 year, suggesting the presence of increased oxidative stress.

beta-Ureidopropionase is the third enzyme of the pyrimidine degradation pathway and it catalyses the irreversible hydrolysis of N-carbamyl-ss-aminoisobutyric acid or N-carbamyl-ss-alanine to beta-aminoisobutyric acid or ss-alanine, ammonia, and CO2. Analysis of the beta-ureidopropionase gene (UPB1) of the first 4 patients presenting with a complete enzyme deficiency, revealed the presence of 2 splice-site mutations (IVS1-2A>G and IVS8-1G>A) and one missense mutation (A85E). RT-PCR analysis of the complete beta-ureidopropionase cDNA suggested that both splice-site mutations lead to a variety of alternative splice variants, with deletions of a single or several exons. The alanine at position 85 was not conserved in other eukaryotic beta-ureidopropionase protein sequences.

Methylmalonic aciduria (MMA-uria) is an autosomal recessive inborn error of amino acid metabolism, involving valine, threonine, isoleucine, and methionine. This organic aciduria may present in the neonatal period with life-threatening metabolic acidosis, hyperammonemia, feeding difficulties, pancytopenia, and coma. Most affected patients have mutations in the methylmalonyl-coenzyme A (methylmalonyl-CoA) mutase gene. Mildly affected patients may present in childhood with failure to thrive and recurrent attacks of metabolic acidosis. Both a higher residual activity of methylmalonyl-CoA mutase as well as the vitamin B12-responsive defects (cblA and cblB) may form the basis of the mild disorder. A few patients with moderate MMA-uria are known in whom no defect could be identified. Here we present a 16-year-old female patient with persisting moderate MMA-uria (approximately 50 mmol/mol creatinine). She was born to consanguineous Caucasian parents. Her fibroblast mutase activity was normal and no effect of vitamin B12 supplementation could be established. Reduced incorporation of 14C-propionate into macromolecules suggested a defect in the propionate-to-succinate pathway. We found a homozygous nonsense mutation (c.139C>T) in the methylmalonyl-CoA epimerase gene (MCEE), resulting in an early terminating signal (p.R47X). Both parents were heterozygous for this mutation; they were found to excrete normal amounts of methylmalonic acid (MMA). This is the first report of methylmalonyl-CoA epimerase deficiency, thereby unequivocally demonstrating the biochemical role of this enzyme in human metabolism.

Aromatic L-amino acid decarboxylase (AADC) deficiency in most cases is a treatable defect in the biosynthesis of the biogenic amine neurotransmitters dopamine and serotonin. Patients are usually detected in infancy due to developmental delay, hypotonia and extrapyramidal movements, but the clinical presentation is variable and can initially be quite mild and/or aspecific. Until now the only way to detect this disorder in general metabolic screening of urine was to identify vanillactic acid (VLA) in GC-MS analysis of organic acids (see figure 1A). Because of the sometimes only small increase of VLA, and/or insufficient analytical sensitivity, AADC deficiency is probably seriously underdiagnosed. We present new findings in recently diagnosed patients that have yielded promising additional diagnostic parameters.

Major depressive disorders (MDD) and cardiovascular disease are mutually associated. They share signs and symptoms of the "metabolic syndrome". Two observations that may be causally related with the metabolic syndrome and therefore with both MDD and cardiovascular disease are a decrease in omega-3 polyunsaturated fatty acids (PUFAs) and a rise in plasma homocysteine (tHcy) levels. Both the rise in tHcy and the decrease in omega-3 PUFAs may be associated with enhanced lipid peroxidation. We exploratively studied 44 randomly chosen patients out of a cohort of 134 patients with the recurrent form of MDD (MDD-R). We measured tHcy levels together with saturated FAs, monounsaturated fatty acids (MUFAs) and PUFAs of the omega-3, omega-6 and omega-9 series in plasma and erythrocytes. Levels were compared with laboratory reference values. The main findings were a decrease in the erythrocytes of C22:5omega-3, C22:6omega-3, C24:1omega-9 and C20:3omega-9 and in the plasma a decrease in C24:1omega-9 and C20:3omega-9. The only significant association we found was between the total of omega-6 fatty acids and plasma tHcy. The FA alterations were found in patients although most of them were clinically recovered, suggesting that the alterations may represent a biological" trait" marker for recurrent depression.

BACKGROUND:Sialic acid storage diseases (SSDs) are severe autosomal recessive neurodegenerative disorders caused by a transport defect across the lysosomal membrane, which leads to accumulation of sialic acid in tissues, fibroblasts, and urine. Defective free sialic acid transport can be established by quantification of free sialic acid in urine.METHODS:Urine sample size was adjusted to the equivalent of 100 nmol of creatinine. After addition of 2-keto-3-deoxy-d-glycero-d-galactonononic acid as internal standard, samples were diluted with water to an end volume of 250 microL. We used 10 microL for HPLC-tandem mass spectrometric analysis in the negative electrospray ionization mode, monitoring transitions m/z 308.3-->m/z 86.9 (sialic acid) and m/z 267.2-->m/z 86.9 (internal standard). The overall method was validated and studied for ion suppression, interfering compounds, and pH effects. Samples from controls (n = 72) and SSD patients (n = 3) were analyzed.RESULTS:The limit of detection was 3 micromol/L. Intraassay imprecision (CV; n = 10) was 6%, 3%, and 2% at 30, 130, and 1000 mmol/mol creatinine, respectively; corresponding interassay CV (n = 10) were 5%, 5%, and 2%. Recovery was 109% (100-1000 mmol/mol creatinine). The mean (SD) [range] excretion rates (mmol/mol creatinine) were 31.3 (16.6) [0.7-56.9] at 0-1 year (n = 20), 21.2 (9.8) [6.3-38.3] at 1-3 years (n = 15), 14.4 (8.2) [1.7-32.9] at 3-10 years (n = 25), and 4.6 (2.6) [0-9.8] above age 10 years (n = 12). SSD patients 1.2, 3.9, and 12 years of age had concentrations of 111.5, 54.2, and 36.1 mmol/mol creatinine, respectively.CONCLUSIONS:The HPLC-tandem MS method for free sialic acid in urine is more rapid, accurate, sensitive, selective, and robust than earlier methods and may serve as a candidate reference method for free sialic acid in diagnosis of SSD.

Het vakgebied klinische chemie van erfelijke metabole ziekten (EMZ) is voortgekomen uit de algemene klinische chemie. Al in de vijftiger jaren van de vorige eeuw werd door enkele Nederlandse klinisch chemici pionierswerk op dit terrein verricht, toen er nog maar een handvol ziekten bekend was. Een en ander werd mogelijk gemaakt door vorderingen in de analytische chemie, met name de ontwikkeling van chromatografische scheidingsmethoden. De zeer vroege beschikbaarheid van aminozuuranalyse (vanaf ongeveer 1960) en suikeranalyse maakte het opsporen van fenylketonurie (PKU) en galactosemie, beide goed behandelbare aandoeningen, mogelijk. Dieetbehandeling was het terrein van enthousiaste kinderartsen, die in de verschillende universitaire kinderklinieken sub-afdelingen Metabole Ziekten oprichtten. Anno 2003 is het vakgebied erfelijke metabole ziekten tot volle wasdom gekomen; internationale congressen trekken gemakkelijk 500 - 1000 deelnemers. Algemeen wordt gerekend met ongeveer 400 bekende erfelijke stofwisselingsziekten, waarvoor diagnostiek mogelijk is. Maandelijks worden hieraan nieuwe defecten toegevoegd. Trefwoorden: erfelijke metabole ziekte; stofwisselingsziekte; diagnostiek; algemene klinische chemie In Nederland wordt het vak klinische chemie van erfelijke metabole ziekten (EMZ) fulltime in alle acht academische ziekenhuizen beoefend (met 4 hoogleraren) en op parttime basis in een toenemend aantal algemene ziekenhuizen. Naast inbedding in de NVKC zijn de beroepsbeoefenaren verenigd in de Vereniging Erfelijke Stofwisselingsziekten Nederland (ESN), die nauw gelieerd is aan de internationale Society for the Study of Inborn Errors of Metabolism (SSIEM, www.ssiem.org.uk).