Head-to-head trial of pegunigalsidase alfa versus agalsidase beta in patients with Fabry disease and deteriorating renal function: results from the 2-year randomised phase III BALANCE study.

BACKGROUND: Pegunigalsidase alfa is a PEGylated α-galactosidase A enzyme replacement therapy. BALANCE (NCT02795676) assessed non-inferiority of pegunigalsidase alfa versus agalsidase beta in adults with Fabry disease with an annualised estimated glomerular filtration rate (eGFR) slope more negative than -2 mL/min/1.73 m2/year who had received agalsidase beta for ≥1 year. METHODS: Patients were randomly assigned 2:1 to receive 1 mg/kg pegunigalsidase alfa or agalsidase beta every 2 weeks for 2 years. The primary efficacy analysis assessed non-inferiority based on median annualised eGFR slope differences between treatment arms. RESULTS: Seventy-seven patients received either pegunigalsidase alfa (n=52) or agalsidase beta (n=25). At baseline, mean (range) age was 44 (18-60) years, 47 (61%) patients were male, median eGFR was 74.5 mL/min/1.73 m2 and median (range) eGFR slope was -7.3 (-30.5, 6.3) mL/min/1.73 m2/year. At 2 years, the difference between median eGFR slopes was -0.36 mL/min/1.73 m2/year, meeting the prespecified non-inferiority margin. Minimal changes were observed in lyso-Gb3 concentrations in both treatment arms at 2 years. Proportions of patients experiencing treatment-related adverse events and mild or moderate infusion-related reactions were similar in both groups, yet exposure-adjusted rates were 3.6-fold and 7.8-fold higher, respectively, with agalsidase beta than pegunigalsidase alfa. At the end of the study, neutralising antibodies were detected in 7 out of 47 (15%) pegunigalsidase alfa-treated patients and 6 out of 23 (26%) agalsidase beta-treated patients. There were no deaths. CONCLUSIONS: Based on rate of eGFR decline over 2 years, pegunigalsidase alfa was non-inferior to agalsidase beta. Pegunigalsidase alfa had lower rates of treatment-emergent adverse events and mild or moderate infusion-related reactions. TRIAL REGISTRATION NUMBER: NCT02795676.

Dairy Fat Intake, Plasma Pentadecanoic Acid, and Plasma Iso-heptadecanoic Acid Are Inversely Associated With Liver Fat in Children.

OBJECTIVES: We sought to evaluate the relevance of pediatric dairy fat recommendations for children at risk for nonalcoholic fatty liver disease (NAFLD) by studying the association between dairy fat intake and the amount of liver fat. The effects of dairy fat may be mediated by odd chain fatty acids (OCFA), such as pentadecanoic acid (C15:0), and monomethyl branched chain fatty acids (BCFA), such as iso-heptadecanoic acid (iso-C17:0). Therefore, we also evaluated the association between plasma levels of OCFA and BCFA with the amount of liver fat. METHODS: Observational, cross-sectional, community-based sample of 237 children ages 8 to 17. Dairy fat intake was assessed by 3 24-hour dietary recalls. Plasma fatty acids were measured by gas chromatography-mass spectrometry. Main outcome was hepatic steatosis measured by whole liver magnetic resonance imaging proton density fat fraction (MRI-PDFF). RESULTS: Median dairy fat intake was 10.6 grams/day (range 0.0--44.5 g/day). Median liver MRI-PDFF was 4.5% (range 0.9%-45.1%). Dairy fat intake was inversely correlated with liver MRI-PDFF (r = -0.162; P = .012). In multivariable log linear regression, plasma C15:0 and iso-C17:0 were inverse predictors of liver MRI-PDFF (B = -0.247, P = 0.048; and B = -0.234, P = 0.009). CONCLUSIONS: Dairy fat intake, plasma C15:0, and plasma iso-C17:0 were inversely correlated with hepatic steatosis in children. These hypothesis-generating findings should be tested through clinical trials to better inform dietary guidelines.

Cardiac tissue citric acid cycle intermediates in exercised very long-chain acyl-CoA dehydrogenase-deficient mice fed triheptanoin or medium-chain triglyceride.

Anaplerotic odd-chain fatty acid supplementation has been suggested as an approach to replenish citric acid cycle intermediate (CACi) pools and facilitate adenosine triphosphate (ATP) production in subjects with long-chain fatty acid oxidation disorders, but the evidence that cellular CACi depletion exists and that repletion occurs following anaplerotic substrate supplementation is limited. We exercised very long-chain acyl-CoA dehydrogenase-deficient (VLCAD-/-) and wild-type (WT) mice to exhaustion and collected cardiac tissue for measurement of CACi by targeted metabolomics. In a second experimental group, VLCAD-/- and WT mice that had been fed chow prepared with either medium-chain triglyceride (MCT) oil or triheptanoin for 4 weeks were exercised for 60 minutes. VLCAD-/- mice exhibited lower succinate in cardiac muscle at exhaustion than WT mice suggesting lower CACi in VLCAD-/- with prolonged exercise. In mice fed either MCT or triheptanoin, succinate and malate were greater in VLCAD-/- mice fed triheptanoin compared to VLCAD-/- animals fed MCT but lower than WT mice fed triheptanoin. Long-chain odd acylcarnitines such as C19 were elevated in VLCAD-/- and WT mice fed triheptanoin suggesting some elongation of the heptanoate, but it is unknown what proportion of heptanoate was oxidized vs elongated. Prolonged exercise was associated with decreased cardiac muscle succinate in VLCAD-/- mice in comparison to WT mice. VLCAD-/- fed triheptanoin had increased succinate compared to VLCAD-/- mice fed MCT but lower than WT mice fed triheptanoin. Cardiac CACi were higher following dietary ingestion of an anaplerotic substrate, triheptanoin, in comparison to MCT.

Promises and pitfalls of untargeted metabolomics.

Metabolomics is one of the newer omics fields, and has enabled researchers to complement genomic and protein level analysis of disease with both semi-quantitative and quantitative metabolite levels, which are the chemical mediators that constitute a given phenotype. Over more than a decade, methodologies have advanced for both targeted (quantification of specific analytes) as well as untargeted metabolomics (biomarker discovery and global metabolite profiling). Untargeted metabolomics is especially useful when there is no a priori metabolic hypothesis. Liquid chromatography coupled to mass spectrometry (LC-MS) has been the preferred choice for untargeted metabolomics, given the versatility in metabolite coverage and sensitivity of these instruments. Resolving and profiling many hundreds to thousands of metabolites with varying chemical properties in a biological sample presents unique challenges, or pitfalls. In this review, we address the various obstacles and corrective measures available in four major aspects associated with an untargeted metabolomics experiment: (1) experimental design, (2) pre-analytical (sample collection and preparation), (3) analytical (chromatography and detection), and (4) post-analytical (data processing).

p300 is not required for metabolic adaptation to endurance exercise training.

The acetyltransferase, E1a-binding protein (p300), is proposed to regulate various aspects of skeletal muscle development, metabolism, and mitochondrial function,viaits interaction with numerous transcriptional regulators and other proteins. Remarkably, however, the contribution of p300 to skeletal muscle function and metabolism,in vivo, is poorly understood. To address this, we used Cre-LoxP methodology to generate mice with skeletal muscle-specific knockout of E1a-binding protein (mKO). mKO mice were indistinguishable from their wild-type/floxed littermates, with no differences in lean mass, skeletal muscle structure, fiber type, respirometry flux, or metabolites of fatty acid and amino acid metabolism.Ex vivomuscle function in extensor digitorum longus and soleus muscles, including peak stress and time to fatigue, as well asin vivorunning capacity were also comparable. Moreover, expected adaptations to a 20 d voluntary wheel running regime were not compromised in mKO mice. Taken together, these findings demonstrate that p300 is not required for the normal development or functioning of adult skeletal muscle, nor is it required for endurance exercise-mediated mitochondrial adaptations.-LaBarge, S. A., Migdal, C. W., Buckner, E. H., Okuno, H., Gertsman, I., Stocks, B., Barshop, B. A., Nalbandian, S. R., Philp, A., McCurdy, C. E., Schenk, S. p300 is not required for metabolic adaptation to endurance exercise training.

Controversies and research agenda in nephropathic cystinosis: conclusions from a "Kidney Disease: Improving Global Outcomes" (KDIGO) Controversies Conference.

Nephropathic cystinosis is an autosomal recessive metabolic, lifelong disease characterized by lysosomal cystine accumulation throughout the body that commonly presents in infancy with a renal Fanconi syndrome and, if untreated, leads to end-stage kidney disease (ESKD) in the later childhood years. The molecular basis is due to mutations in CTNS, the gene encoding for the lysosomal cystine-proton cotransporter, cystinosin. During adolescence and adulthood, extrarenal manifestations of cystinosis develop and require multidisciplinary care. Despite substantial improvement in prognosis due to cystine-depleting therapy with cysteamine, no cure of the disease is currently available. Kidney Disease: Improving Global Outcomes (KDIGO) convened a Controversies Conference on cystinosis to review the state-of-the-art knowledge and to address areas of controversies in pathophysiology, diagnostics, monitoring, and treatment in different age groups. More importantly, promising areas of investigation that may lead to optimal outcomes for patients afflicted with this lifelong, systemic disease were discussed with a research agenda proposed for the future.

Diagnosis and Monitoring of Cystinosis Using Immunomagnetically Purified Granulocytes.

BACKGROUND: Cystine determination is a critical biochemical test for the diagnosis and therapeutic monitoring of the lysosomal storage disease cystinosis. The classical mixed-leukocyte cystine assay requires prompt specialized recovery/isolation following blood drawing, providing cystine concentrations normalized to total protein from assorted types of white blood cells, each with varying cystine content. METHODS: We present a new workflow for cystine determination using immunomagnetic granulocyte purification, and new reference ranges established from 47 patient and 27 obligate heterozygote samples assayed. Samples were collected in acid-citrate dextrose tubes and their stability was proven to allow for overnight shipping before analysis. Cystine was quantified by LC-MS/MS. RESULTS: The new method was reproducible (<15% root mean square error) and specific, assaying purified granulocytes from blood samples that no longer required immediate preparation and therefore allowing for up to 30 h before processing. There was a nearly a 2-fold increase in the therapeutic target (1.9 nmol half-cystine/mg protein) range, established using distributions of patient, obligate heterozygote, and control samples. The 2.5-97.5 percentile ranges (-2 SD to +2 SD around mean) for these cohorts were 0.67-6.05 nmol/mg protein for patients, 0.33-1.35 nmol/mg protein for obligate heterozygotes, and 0.09-0.35 nmol/mg protein for controls. CONCLUSIONS: The intracellular cystine determination method using immunopurified granulocytes followed by LC-MS/MS analysis improves the inherent variability of mixed leukocyte analysis and eliminates the need for immediate sample preparation following blood draw.

In vivo monitoring of urea cycle activity with (13)C-acetate as a tracer of ureagenesis.

BACKGROUND: The hepatic urea cycle is the main metabolic pathway for detoxification of ammonia. Inborn errors of urea cycle function present with severe hyperammonemia and a high case fatality rate. Long-term prognosis depends on the residual activity of the defective enzyme. A reliable method to estimate urea cycle activity in-vivo does not exist yet. The aim of this study was to evaluate a practical method to quantify (13)C-urea production as a marker for urea cycle function in healthy subjects, patients with confirmed urea cycle defect (UCD) and asymptomatic carriers of UCD mutations. METHODS: (13)C-labeled sodium acetate was applied orally in a single dose to 47 subjects (10 healthy subjects, 28 symptomatic patients, 9 asymptomatic carriers). RESULTS: The oral (13)C-ureagenesis assay is a safe method. While healthy subjects and asymptomatic carriers did not differ with regards to kinetic variables for urea cycle flux, symptomatic patients had lower (13)C-plasma urea levels. Although the (13)C-ureagenesis assay revealed no significant differences between individual urea cycle enzyme defects, it reflected the heterogeneity between different clinical subgroups, including male neonatal onset ornithine carbamoyltransferase deficiency. Applying the (13)C-urea area under the curve can differentiate between severe from more mildly affected neonates. Late onset patients differ significantly from neonates, carriers and healthy subjects. CONCLUSION: This study evaluated the oral (13)C-ureagenesis assay as a sensitive in-vivo measure for ureagenesis capacity. The assay has the potential to become a reliable tool to differentiate UCD patient subgroups, follow changes in ureagenesis capacity and could be helpful in monitoring novel therapies of UCD.

Consanguinity and rare mutations outside of MCCC genes underlie nonspecific phenotypes of MCCD.

PURPOSE: 3-Methylcrotonyl-CoA carboxylase deficiency (MCCD) is an autosomal recessive disorder of leucine catabolism that has a highly variable clinical phenotype, ranging from acute metabolic acidosis to nonspecific symptoms such as developmental delay, failure to thrive, hemiparesis, muscular hypotonia, and multiple sclerosis. Implementation of newborn screening for MCCD has resulted in broadening the range of phenotypic expression to include asymptomatic adults. The purpose of this study was to identify factors underlying the varying phenotypes of MCCD. METHODS: We performed exome sequencing on DNA from 33 cases and 108 healthy controls. We examined these data for associations between either MCC mutational status, genetic ancestry, or consanguinity and the absence or presence/specificity of clinical symptoms in MCCD cases. RESULTS: We determined that individuals with nonspecific clinical phenotypes are highly inbred compared with cases that are asymptomatic and healthy controls. For 5 of these 10 individuals, we discovered a homozygous damaging mutation in a disease gene that is likely to underlie their nonspecific clinical phenotypes previously attributed to MCCD. CONCLUSION: Our study shows that nonspecific phenotypes attributed to MCCD are associated with consanguinity and are likely not due to mutations in the MCC enzyme but result from rare homozygous mutations in other disease genes.Genet Med 17 8, 660-667.

Perturbations of tyrosine metabolism promote the indolepyruvate pathway via tryptophan in host and microbiome.

The drug nitisinone (NTBC) is used to treat tyrosinemia type I, and more recently has been also used for the treatment of another disorder of tyrosine metabolism, alkaptonuria. While studying the dose effects of NTBC treatment on alkaptonuria, untargeted metabolomics revealed perturbations in a completely separate pathway, that of tryptophan metabolism. Significant elevations in several indolic compounds associated with the indolepyruvate pathway of tryptophan metabolism were present in NTBC-treated patient sera and correlated with elevations of an intermediate of tyrosine metabolism. Indolic compounds of this pathway have long been associated with commensal bacterial and plant metabolism. These exogenous sources of indoles have been more recently implicated in affecting mammalian cell function and disease. We studied the correlation of these indolic compounds in other disorders of tyrosine metabolism including tyrosinemia types I and II as well as transient tyrosinemia, and demonstrated that 4-hydroxyphenylpyruvate (4-HPP) was directly responsible for the promotion of this pathway. We then investigated the regulation of the indolepyruvate pathway and the role of 4-HPP further in both mammalian cells and intestinal microbial cultures. We demonstrated that several of the indolic products, including indolepyruvate and indolelactate, were in fact generated by human cell metabolism, while the downstream indole metabolite, indolecarboxaldehyde, was produced exclusively by microbial cultures of human gut flora. This study describes a symbiotic perturbation in host and microbiome tryptophan metabolism in response to elevations related to defects of tyrosine metabolism and concomitant drug treatment.

Cysteamine modulates oxidative stress and blocks myofibroblast activity in CKD.

Therapy to slow the relentless expansion of interstitial extracellular matrix that leads to renal functional decline in patients with CKD is currently lacking. Because chronic kidney injury increases tissue oxidative stress, we evaluated the antifibrotic efficacy of cysteamine bitartrate, an antioxidant therapy for patients with nephropathic cystinosis, in a mouse model of unilateral ureteral obstruction. Fresh cysteamine (600 mg/kg) was added to drinking water daily beginning on the day of surgery, and outcomes were assessed on days 7, 14, and 21 after surgery. Plasma cysteamine levels showed diurnal variation, with peak levels similar to those observed in patients with cystinosis. In cysteamine-treated mice, fibrosis severity decreased significantly at 14 and 21 days after unilateral ureteral obstruction, and renal oxidized protein levels decreased at each time point, suggesting reduced oxidative stress. Consistent with these results, treatment of cultured macrophages with cysteamine reduced cellular generation of reactive oxygen species. Furthermore, treatment with cysteamine reduced α-smooth muscle actin-positive interstitial myofibroblast proliferation and mRNA levels of extracellular matrix proteins in mice and attenuated myofibroblast differentiation and proliferation in vitro, but did not augment TGF-ß signaling. In a study of renal ischemia reperfusion, cysteamine therapy initiated 10 days after injury and continued for 14 days decreased renal fibrosis by 40%. Taken together, these data suggest previously unrecognized antifibrotic actions of cysteamine via TGF-ß-independent mechanisms that include oxidative stress reduction and attenuation of the myofibroblast response to kidney injury and support further investigation into the potential benefit of cysteamine therapy in the treatment of CKD.

Infants suspected to have very-long chain acyl-CoA dehydrogenase deficiency from newborn screening.

Very long-chain acyl-CoA dehydrogenase deficiency (VLCADD) is a fatty acid oxidation disorder with widely varying presentations that has presented a significant challenge to newborn screening (NBS). The Western States Regional Genetics Services Collaborative developed a workgroup to study infants with NBS positive for VLCADD. We performed retrospective analysis of newborns with elevated C14:1-acylcarnitine on NBS in California, Oregon, Washington, and Hawai'i including available confirmatory testing and clinical information. Overall, from 2,802,504 children screened, there were 242 cases screen-positive for VLCADD. There were 34 symptomatic true positive cases, 18 asymptomatic true positives, 112 false positives, 55 heterozygotes, 11 lost to follow-up, and 12 other disorders. One in 11,581 newborns had an abnormal NBS for suspected VLCADD. Comparison of analytes and analyte ratios from the NBS demonstrated statistically significant differences between true positive and false positive groups for C14:1, C14, C14:1/C2, and C14:1/C16. The positive predictive value for all true positive cases was 94%, 54%, and 23% when C14:1 was ≥2.0 µM, ≥1.0 µM, and ≥0.7 µM, respectively. Sequential post-analytical analysis could reduce the referral rate in 25.8% of cases. This study is the largest reported follow-up of infants with NBS screen-positive results for suspected VLCADD and demonstrates the necessity of developing comprehensive and consistent long-term follow-up NBS systems. Application of clinical information revealed differences between symptomatic and asymptomatic children with VLCADD. Comparison of NBS analytes and analyte ratios may be valuable in developing more effective diagnostic algorithms.

A model-driven quantitative metabolomics analysis of aerobic and anaerobic metabolism in E. coli K-12 MG1655 that is biochemically and thermodynamically consistent.

The advent of model-enabled workflows in systems biology allows for the integration of experimental data types with genome-scale models to discover new features of biology. This work demonstrates such a workflow, aimed at establishing a metabolomics platform applied to study the differences in metabolomes between anaerobic and aerobic growth of Escherichia coli. Constraint-based modeling was utilized to deduce a target list of compounds for downstream method development. An analytical and experimental methodology was developed and tailored to the compound chemistry and growth conditions of interest. This included the construction of a rapid sampling apparatus for use with anaerobic cultures. The resulting genome-scale data sets for anaerobic and aerobic growth were validated by comparison to previous small-scale studies comparing growth of E. coli under the same conditions. The metabolomics data were then integrated with the E. coli genome-scale metabolic model (GEM) via a sensitivity analysis that utilized reaction thermodynamics to reconcile simulated growth rates and reaction directionalities. This analysis highlighted several optimal network usage inconsistencies, including the incorrect use of the beta-oxidation pathway for synthesis of fatty acids. This analysis also identified enzyme promiscuity for the pykA gene, that is critical for anaerobic growth, and which has not been previously incorporated into metabolic models of E coli.

Metabolomics reveals signature of mitochondrial dysfunction in diabetic kidney disease.

Diabetic kidney disease is the leading cause of ESRD, but few biomarkers of diabetic kidney disease are available. This study used gas chromatography-mass spectrometry to quantify 94 urine metabolites in screening and validation cohorts of patients with diabetes mellitus (DM) and CKD(DM+CKD), in patients with DM without CKD (DM-CKD), and in healthy controls. Compared with levels in healthy controls, 13 metabolites were significantly reduced in the DM+CKD cohorts (P≤0.001), and 12 of the 13 remained significant when compared with the DM-CKD cohort. Many of the differentially expressed metabolites were water-soluble organic anions. Notably, organic anion transporter-1 (OAT1) knockout mice expressed a similar pattern of reduced levels of urinary organic acids, and human kidney tissue from patients with diabetic nephropathy demonstrated lower gene expression of OAT1 and OAT3. Analysis of bioinformatics data indicated that 12 of the 13 differentially expressed metabolites are linked to mitochondrial metabolism and suggested global suppression of mitochondrial activity in diabetic kidney disease. Supporting this analysis, human diabetic kidney sections expressed less mitochondrial protein, urine exosomes from patients with diabetes and CKD had less mitochondrial DNA, and kidney tissues from patients with diabetic kidney disease had lower gene expression of PGC1α (a master regulator of mitochondrial biogenesis). We conclude that urine metabolomics is a reliable source for biomarkers of diabetic complications, and our data suggest that renal organic ion transport and mitochondrial function are dysregulated in diabetic kidney disease.

Changes in plasma and urine globotriaosylceramide levels do not predict Fabry disease progression over 1 year of agalsidase alfa.

PURPOSE: Globotriaosylceramide concentrations were assessed as potential predictors of change from baseline after 12 months by estimated glomerular filtration rate and left-ventricular mass index using pooled data from three randomized, placebo-controlled agalsidase alfa trials and open-label extensions of patients with Fabry disease. METHODS: Males (aged 18 years or older) with Fabry disease received agalsidase alfa (0.2 mg/kg every other week for 12 months). A backward-elimination approach evaluated potential predictors (baseline estimated glomerular filtration rate and left-ventricular mass index; age at first dose; baseline and change from baseline at 12 months of globotriaosylceramide (urine, plasma); urine protein excretion; and systolic and diastolic blood pressure). Subgroups included patients randomized to placebo or agalsidase alfa (double-blind phase), then to agalsidase alfa (open-label extensions; placebo→agalsidase alfa or agalsidase alfa→agalsidase alfa, respectively) and stage 2/3 chronic kidney disease patients. RESULTS: Baseline estimated glomerular filtration rate, age at first dose, baseline urine globotriaosylceramide excretion, and baseline and change from baseline urine protein excretion significantly predicted change from baseline estimated glomerular filtration rate in the analysis population (N = 73; all P<0.05), although not in all subgroups. Change from baseline urine and plasma globotriaosylceramide (baseline and change from baseline) concentrations did not predict change from baseline estimated glomerular filtration rate. No predictors of left-ventricular mass index were significant. CONCLUSION: Changes in globotriaosylceramide concentrations do not appear to be useful biomarkers for prediction of Fabry disease-related changes in estimated glomerular filtration rate or left-ventricular mass index.

Elevation of guanidinoacetate in newborn dried blood spots and impact of early treatment in GAMT deficiency.

Guanidinoacetate methyltransferase (GAMT) deficiency is a good candidate disorder for newborn screening because early treatment appears to improve outcomes. We report elevation of guanidinoacetate in archived newborn dried blood spots for 3 cases (2 families) of GAMT deficiency compared with an unaffected carrier and controls. We also report a new case of a patient treated from birth with normal developmental outcome at the age of 42 months.

Biochemical, molecular, and clinical characteristics of children with short chain acyl-CoA dehydrogenase deficiency detected by newborn screening in California.

BACKGROUND: Short-chain acyl-CoA dehydrogenase deficiency (SCADD) is an autosomal recessive inborn error of mitochondrial fatty acid oxidation with highly variable biochemical, genetic, and clinical characteristics. SCADD has been associated with accumulation of butyryl-CoA byproducts, including butyrylcarnitine (C4), butyrylglycine, ethylmalonic acid (EMA), and methylsuccinic acid (MS) in body fluid and tissues. Differences in genotype frequencies have been shown between patients diagnosed clinically versus those diagnosed by newborn screening. Moreover, while patients diagnosed clinically have a variable clinical presentation including developmental delay, ketotic hypoglycemia, epilepsy and behavioral disorders, studies suggest patients diagnosed by newborn screening are largely asymptomatic. Scant information is published about the biochemical, genetic and clinical outcome of SCADD patients diagnosed by newborn screening. METHODS: We collected California newborn screening, follow-up biochemical levels, and ACADS mutation data from September, 2005 through April, 2010. We retrospectively reviewed available data on SCADD cases diagnosed by newborn screening for clinical outcomes. RESULTS: During the study period, 2,632,058 newborns were screened and 76 confirmed SCADD cases were identified. No correlations between initial C4 value and follow-up biochemical markers (C4, EMA or MS levels) were found in the 76 cases studied. We found significant correlation between urine EMA versus MS, and correlation between follow-up C4 versus urine EMA. Of 22 cases where ACADS gene sequencing was performed: 7 had two or more deleterious mutations; 8 were compound heterozygotes for a deleterious mutation and common variant; 7 were homozygous for the common variant c.625G>A; and 1 was heterozygous for c.625G>A. Significant increases in mean urine EMA and MS levels were noted in patients with two or more deleterious mutations versus mutation heterozygotes or common polymorphism homozygotes. Clinical outcome data was available in 31 patients with follow-up extending from 0.5 to 60 months. None developed epilepsy or behavioral disorders, and three patients had isolated speech delay. Hypoglycemia occurred in two patients, both in the neonatal period. The first patient had concomitant meconium aspiration; the other presented with central apnea, poor feeding, and hypotonia. The latter, a c.625G>A homozygote, has had persistent elevations in both short- and medium-chain acylcarnitines; diagnostic workup in this case is extensive and ongoing. CONCLUSIONS: This study examines the largest series to date of SCADD patients identified by newborn screening. Our results suggest that confirmatory tests may be useful to differentiate patients with common variants from those with deleterious mutations. This study also provides evidence to suggest that, even when associated with deleterious mutations, SCADD diagnosed by newborn screening presents largely as a benign condition.

Novel mutations in the human MCCA and MCCB gene causing methylcrotonylglycinuria.

Methylcrotonylglycinuria (MCG) is an inborn error of leucine catabolism and has a recessive pattern of inheritance that results from the deficiency of 3-methylcrotonyl-CoA carboxylase (MCC). The clinical phenotypes are highly variable ranging from neonatal onset with severe neurological involvement to asymptomatic adults. Here we identified two novel MCCA (exon 3: c.137G>A; p.46G>E), (IVS7-1G>A splice site mutation), and four novel MCCB (exon 11: c.1065A>T; p.355L>F), (exon 15: c.1430A>G; p.477Q>R), (exon 16: c.1549G>A; p.517G>R), (exon 16: c.1559A>C; p.520Y>S) mutant alleles from five MCC-deficient patients.

45-Year-old female with propionic acidemia, renal failure, and premature ovarian failure; late complications of propionic acidemia?

We describe a 45-year-old patient who was diagnosed with propionic acidemia in infancy, who experienced an unstable first two years of life but who eventually had a good developmental outcome. She developed severe renal failure requiring renal transplantation in her forties and premature ovarian failure. Renal failure and premature ovarian failure have not previously been associated with propionic acidemia. We hypothesize that propionic acidemia may have contributed to these complications, and discuss several possible mechanisms for this, emphasizing mainly the electron transport chain/mitochondrial energy deficiency hypothesis.