A NOVEL BIOTIN-SENSITIVE LEUKODYSTROPHY (BSL) Sedel F 1 , Challe G 1 , Vignal C 1 , Assouad R 1 , Bellanger A 1 , Galanaud D 1 1 Pitié-Salpêtrière Hospital, Paris, France Biotin (vitamin H) is a water soluble vitamin which serves as a cofactor for (1) pyruvate carboxylase (gluconeogenesis), (2) 3-methylcrotonyl CoA carboxylase, (3) propionyl CoA carboxylase and (4) acetyl CoA carboxylase. In addition, biotin regulates in vitro and in vivo the expression of numerous genes. In humans, two inherited metabolic diseases directly affect the metabolism of biotin: biotinidase and holocarboxylase synthetase deficiencies. A third disease, Biotin responsive basal ganglia disease (BBGD) was identified in patients from the Middle East presenting with encephalopathies responding to high doses of biotin. The gene responsible for the disease, SLC19A3, encodes a second thiamine transporter (ThTr-2). Here we report the identification of a novel biotin-responsive disease in four unrelated individuals, three of them originated from North Algeria. All patients display a characteristic leukodystrophy that involves the periventricular white matter, corticospinal tracts, cerebellar peduncles, and optic radiations. Clinical features encompass relapsing episodes of cerebellar ataxia and optic neuropathy. In between episodes, patients may present psychiatric problems, cerebellar ataxia and/or optic atrophy. Visual evoked potentials show absence of P100 or increased latency of the P100, consistent with involvement of optic nerves. Clinical, electrophysiological, radiological and spectroscopic parameters improve after treatment with very high doses of biotin. Background: CYP7B1 encodes the cytochrome P450 alpha-hydroxylase and plays a role in the alternate/acidic pathway for primary bile acid production. Mutations in CYP7B1 were identified in children with severe liver disease and, recently, in adult SPG5 patients presenting with hereditary spastic paraplegias (HSP) (Tsaousidou, 2008) . Increased levels of 27-hydroxycholesterol (27-OHC) were reported in the plasma of 4 SPG5 patients (Schüle, 2010). Besides its role in hepatobiliary metabolism, in vitro studies suggest that 27-OHC may decrease bone mineral density, inhibit the cardiovascular protective effects of estrogens and increase oxidative stress in retinal pigment epithelial cells. Methods: We investigated 9 SPG5 patients from 6 families in order (i) to determine whether plasma 27-OHC can be used as a biomarker to screen HSP patients, and (ii) to explore the non-neurological manifestations that may result from the altered oxysterols metabolism in SPG5 patients-i.e. hepatobiliary and cardiovascular functions, bone homeostasis and retina. Results: The marked elevation of plasma 27-OHC was associated with altered liver functions, reduced bone density and optic atrophy in SPG5 patients. Conclusion: Following these investigations, we are now designing a clinical trial to determine the best candidate drugs-and doses-to lower 27-OHC in SPG5 patients. Background: Availability of sophisticated techniques for selective screening for inborn errors of metabolism is not available in Guatemala. Since 2002 the authors have a close collaboration between Guatemala and Germany/Switzerland. Methods: Aminoacids and Acylcarnitines were extracted from dried blood spots (DBS), and measured by MS/MS. If necessary for confirmatory diagnostics, organic acids were determined after extraction of urine dried on filterpaper. Results: Between 2002 and 2011 we have analysed 444 DBS samples from Guatemalan children. So far we found 11 children with a confirmed inborn error of metabolism (1*PKU; 1*GA-I; 1*Cit-type 2; 1*MCADD; 2*Arginase-def.; 1*VLCADD; 1*PA; 2*MSUD; 1*CAH), and 5 further cases with a presumptive, not yet confirmed, diagnosis (1*MADD; 2*OCTN2; 1*LCHADD, 1*CPT-II). The calculated effectivity index (confirmed cases only) is 2.4% (95% C.I.:1.2-4.4%) or 1 case in about 40 requests. Conclusions: Although the total number of samples is still quite low, the high rate of positive cases indicates that: (1) The overall incidence of inborn errors of metabolism is (at least) as high as in other countries. (2) The preselection of patients to be investigated, is at least as good as in other countries. International cooperation is a simple and effective way to provide state-of-the-art diagnostics for developing countries. Delivery of the best possible genetic healthcare to those with genetic disease requires access to all data of all individuals with their mutations and phenotype. The reasons and the utility of such data has been published recently (Cotton et al Genetics in Medicine, 11:843-9, 2009 ). Collection of such data has been suggested to be best by two complementary but partially redundant routes: (a) collection of all instances of each disease in each country and (b) collection of this data into gene or disease specific databases. There are examples of each of these: country (Australia) at www.hvpaustralia.org and colon cancer www.insight-group.org (Kohonen Corish et al Hum Mutat 2010; 31: 1374-81) . The latter is curated by representatives of a society, InSiGHT (www.insight-group.org). A similar grouping is emerging in inherited neurological disease (Haworth A et al Neurogenetics, 2011; in press) and being initiated in Mitochondrial disease. Such groupings have advantages such as load sharing, more capacity to raise funds, etc. There is a case for a similar initiative in Inborn Errors of Metabolism supported by the Human Variome Project. Those interested could contact Nenad Blau at: nenad.blau@kispi.uzh.ch Background: Metabolomics becomes an important tool in clinical research and diagnosing human diseases. In this work we focused on diagnosing of inborn errors of metabolism (IEMs) in plasma samples using a targeted metabolomic approach. Methods: Plasma samples were analysed using the AbsoluteIDQ p 150 Kit (BIOCRATES Life Sciences AG, Austria). The standard flow injection method of the kit comprising two subsequent 10 ul injections (one for positive and one for negative detection mode) was applied for all measurements. All experiments were performed on an QTRAP 5500 tandem mass spectrometer (AB SCIEX, USA) with electrospray ionization. Multiple reaction monitoring detection was used for quantification. Results: We analyzed 50 control samples and 34 samples with amino acids defects (phenylketonuria, maple syrup urine disease, tyrosinemia I, argininemia, homocystinuria, carbamoyl phosphate synthetase deficiency, ornithine transcarbamylase deficiency, non-ketotic hyperglycinemia) and with acylcarnitine defects (methylmalonic acidemia, propionic acidemia, glutaric aciduria I, 3-hydroxy-3-methylglutaric aciduria, isovaleric acidemia, medium-chain acyl-coenzyme A dehydrogenase deficiency and carnitine palmitoyltransferase II deficiency A screening method for 36 metabolites related to inborn errors of metabolism (IEMs) of purines, pyrimidines, creatine, organic acid and galactose metabolism is presented. The method utilises ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-TMS) for analysis of the compounds in urine. Selected compounds were separated on a Phenomenex Luna 3 μ NH2 column in hydrophilic interaction mode with gradient elution. Mobile phase consisted of 20 mM ammonium acetate, pH 9.45 (A) and acetonitrile (B). Total analysis time was 13 min. Detection was performed by multiple reaction monitoring (MRM) in positive and/or negative mode.Two MRM transitions were chosen for each compound (with the exception of four compounds) and retention times were used for peak confirmation. Detector response was linear in the range of expected concentrations, except for pyruvate and uric acid. Limits of detection (LOD) lie below or within the normal concentration range of the metabolites. The chosen metabolites were analysed in normal urine, elevated concentrations of typical metabolites were found in urines of patients with IEMs. The presented UHPLC-TMS method is fast, sensitive and requires only minimal sample preparation. This work was supported by grants CZ. Background: 1H NMR spectroscopy has successfully been applied to the field of inborn errors of metabolism (IEM). However, body fluid NMR spectra lead to a wealth of data and it can be a daunting task to find the relevant information. In this study, chemometric methods were used to analyze urine NMR data. Objectives: To demonstrate the use of chemometric methods in NMRbased urine analyses to diagnose patients with IEM. Alkaptonuria (ALK) and methylmalonic aciduria (MMA) will be used as examples. Material and Methods: 1H NMR spectroscopy was performed on urine samples from 58 healthy volunteers, 5 patients with ALK and 6 patients with MMA. Principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) were applied in order to establish models for discrimination between diseased and normal urine samples. Results: Discrimination of ALK and MMA urine from healthy urine was achieved by PLS-DA with excellent results. Not only is the technique able to find perfect separation of the groups, also the resonances of the diagnostic metabolites for both diseases can easily be found. Conclusion/Discussion: Chemometrics analyses can assist the interpretation of complex body fluid NMR data and may be useful in the rapid screening of patients with IEM.