Haynes BA 1 , Bragat A 1 , Sitaraman-Das S 1 , Schiffmann R 2 1 Amicus Therapeutics, Cranbury, NJ, United States 2 Baylor Research Institute, Dallas, TX, United States Background: Fabry disease (FD) is an X-linked lysosomal storage disorder characterized by deficiency of alpha-galactosidase A with accumulation of globotriaosylceramide (GL-3) in tissues and body fluids, leading to progressive, multi-organ disease. Objective: To demonstrate the utility of a validated GLP assay to quantify two isoforms of urinary . Methods: Two kidney-related isoforms were measured in total 24 h urine in 120 untreated Fabry subjects from the FACETS clinical study and 38 healthy controls. C22:0 and C24:0 isoforms were quantified after urine homogenization and sonication; uGL-3 concentration is expressed relative to creatinine (ng/mg Cr). Preliminary Results: Males with Fabry disease have elevated levels of uGL-3 (n=38; mean+SD=888+865; median=846; range 9.8-3504) when compared to FD females and controls. Levels of uGL-3 in females ranged from 3.8 to1079(n=82; mean=183+192; median=143), overlapping with the range for the affected male population. In females, the mean level of uGL-3 was higher than that of the control group (control group mean= 17.1+8.3; median 14.5; range 7. 3-50.3) . Conclusion: This new analytically validated uGL-3 assay was used as a study screening tool to assess baseline uGL-3 levels in males and females with Fabry disease. It is anticipated that this assay may be useful to monitor treatment effects in FD subjects. Conflict of Interest declared. Background: NP-C diagnosis can be delayed for years due to heterogeneous presentation. A Suspicion Index was developed, ranking specific symptoms within and across domains, and with family history, providing a predictive score to help identify suspected patients with NP-C. Objectives: To construct a Suspicion Index and validate its sensitivity and specificity using patient data. Methods: A retrospective chart review in seven centres in Europe and Australia (N=216). Three patient types were selected: classical or variant filipin staining NP-C-positive cases (n=71); suspected NP-C cases that were filipin staining negative (n=65); or non-cases (without NP-C suspicion) with ≥1 characteristic symptom of NP C (n=80). Signs and symptoms of NP-C were categorized into visceral, neurological or psychiatric domains, and scores assigned according to their relative prediction for NP-C. Results: The Suspicion Index has good discriminatory performance (area under the ROC curve >0.75) with cut-points for grading suspicion of NP-C. Neonatal jaundice/cholestasis, splenomegaly, vertical supranuculear gaze palsy, cataplexy, and cognitive decline/dementia were strong predictors of NP-C, as well as symptoms occurring in multiple domains in individual patients, and siblings/parents with NP-C. Conclusion: The Suspicion Index is a valid tool that can help identify patients who may warrant further investigation for NP-C. Conflict of Interest declared. Methods: Retrospective study and review of laboratory/neuroimaging data were carried out in NPC patients diagnosed in the last 6 years. Results: Twenty-six patients were confirmed to have NPC by filipin staining ( 14 patients required molecular analysis). Regarding clinical form, 7 were perinatal , 16 infantile, 11 juvenile and 8 adults. Prolonged neonatal jaundice was a common feature and five of them were diagnosed with "neonatal hepatitis". Hepatosplenomegaly was present in all perinatal/ infantile patients, but absent in 6 adults. Ocular abnormalities were seen in 38 patients ( mostly, vertical supranuclear gaze paralysis). Leukoencephalopathy and progressive cerebral/cerebellar atrophy were the main MRI features. Other systemic manifestations included dystonia, cataplexia, immunodeficiency, dysphagia. At the time of the study, 13 patients were deceased. Conclusions: Onset of neurological disease in NPC patients is extremely variable, even in the same sibship. Better understanging of the natural history of the disease is crucial for evaluation of potential therapeutic approaches in such devastating disorder. Niemann-Pick type C (NPC) is a rare autosomal-recessive disorder caused by accumulation of unesterified cholesterol and glycosphingolipids in the lysosomes. Clinically is characterized by progressive neurological deterioration and hepato-splenomegaly. The aim of this study was to analyze the activity of sphingomyelinase, beta-glucosidase, beta-galactosidase and chitotriosidase in blood samples from patients with NPC and correlate this activity with that of normal individuals. Our results showed that NPC patients have a decreased sphingomyelinase activity (63%) than normal subjects (NPC=1.52 ±0.52 nmol/h/mgprot; normal=2.39±1.14 nmol/h/mgprot) and this difference was significant (p<0.04). NPC patients have 11.40±2.82 nmol/h/mgprot of beta-glucosidase activity and 1882±2461 nmol/h/mL of chitotriosidase activity compared to 14.8±4.49 nmol/h/mgprot for beta-glucosidase and 58.5±34.6 nmol/h/mL of chitotriosidase from normal individuals. The differences between the two groups are significant for both enzymes (p< 0.02 and p<0.0002 for beta-glucosidase and chitotriosidase, respectively). Beta-galactosidase did not differ between the two groups (NPC=152.6± 52.8 nmol/h/mgprot; normal=125.0±38.6 nmol/h/mgprot). These results allow us to conclude that cholesterol accumulation may be interfering with the metabolism of other sphingolipids such as glucosylceramide and sphingomyelin, affecting the activity of their degrading enzymes. Niemann-Pick disease type C (NPC) is an autosomal recessive condition caused by defects on cholestherol trafikking, which leads to a progressive and usually severe visceral and/or neurologic syndrome. The signs and symptoms overlap with other conditions, and diagnosis is difficult as usually requires a staining test performed on cultured fibroblasts and/or comprehensive molecular studies of the two NPC genes (NPC1 and NPC2). The possibility of treating affected patients with substrate reduction therapy (miglustat) makes more important the correct and timely identification of affected patients. With this aim we set up a comprehensive diagnostic program for NPC in Brazil, which includes: 1) providing information on diagnostic procedures, including a collection and transportation kit (for skin biopsy and blood collection); 2) assay of plasma chitotriosidase; 3) culture of skin fibroblasts and Filipin staining; 4)DNA isolation and molecular analysis of NPC1 and NPC2 gene, depending on results of the Filipin testing. This protocol was performed in 140 patients with suspected NPC, referred from physicians from all Brazilian regions. Diagnosis of NPC was confirmed in 15 cases, being abnormal but not conclusive in further 12 patients. These 27 cases were referred to molecular analysis of both genes to complete investigation (molecular analyses in progress). Conflict of Interest declared. Niemann-Pick C (NPC) disease is an autosomal recessive condition related to cholesterol trafficking defects. We describe a male patient and her aunt with NPC, with different genotypes and phenotype heterogeneity. Case 1: male,born from non-consanguineous parents,presented with normal development until the age of 2 years when started generalized seizures. The brain MRI showed hyperintense periventricular signal. At 3 years of age showed hepatosplenomegaly,umbilical hernia and spastic quadriplegia. Plasma chitotriosidase was increased (1749 nmoles/mL)and Filipin test in fibroblasts was strongly positive. Molecular analysis showed different mutations on the NPC1 gene(G1140V and L1157P). The patient died at 5 years due to respiratory complications. His aunt(father's sister),daughter of non-consanguineous parents,began at the age of 4 years with seizures, jerky movements who progressed with cognitive impairment, motor disorders,hallucinations and aggressiveness. At the age of 26 years she presents ataxia, dysmetria, choreoathetosis and absence of convergence and vertical saccades. The Filippin test in fibroblasts was positive. Molecular analysis of the NPC1 gene showed only one mutation (L1157P),indicating that the patient would be a carrier or would present a complex genotype. Although genetic heterogeneity could explain the different phenotypes observed in this family,further molecular studies(in progress)should help to clarify this interesting situation. (2-13.5y )] were followed in our centre between 2007 and 2011. They fell into 4 subgroups: 5 patients presented only with visceral symptoms (mean age of onset: 1 month) followed by neurological symptoms in the early infantile period in 3 of them (M: 12 m). Four patients (4-12 y) presented with neurovisceral symptoms and 2 patients presented with pure neuropsychological symptoms (3-7y) . Six patients were treated by miglustat for a mean period of 14 months (6-22 m). All patients underwent clinical assessments including neuropsychological tests, Pineda disability scale and MRI with diffusion tensor imaging (DTI) and spectroscopy (MRS). The mean dose of miglustat was 443 mg (457 mg/ m2). In treated patients neurological deterioration was observed in 4 and stabilization in 2. MRI showed retarded myelination but normal MRS. Fractional anisotropy (FA) was low in most patients compared to controls and seemed to correlate to age and disease severity. FA increased in some treated patients. Miglustat was well tolerated except in one patient who interrupted treatment. Response to treatment was variable with a trend for stabilization in patients treated at an early stage of the disease. Morbus Niemann -Pick type C (NPC) is a neurovisceral lysosomal storage disorder caused by mutations of either NPC1 or NPC2 gene. It results in abnormal cellular cholesterol trafficking and secondary accumulation of glycosphingolipids. The precise mechanism leading to clinical symptoms is unknown. One potential mechanism may be altered composition of cellular membrane lipids with subsequent impairment of lipid raft function and protein trafficking. Miglustat (N-butyl-deoxynojirimycin, NB-DNJ), an N-alkylated imino-sugar, inhibits the ceramide-specific glucosyltransferase has been advocated in the treatment of NPC. Methods and results: We examined the effects of NB-DNJ on membrane trafficking in fibroblasts of NPC patients and controls. We found substantial reduction in glycosphingolipid levels in the patient's fibroblasts after treatment with NB-DNJ. Furthermore, NB-DNJ resulted in a partial restoration of cholesterol-and sphingolipid-enriched membrane microdomains (lipid rafts) as assessed by increased levels of flotillin 2 (a protein marker of lipid rafts) in the floating fractions of sucrose density gradients in Triton X-100 lysates of NB DNJ-treated NPC fibroblasts. Finally, the enzymatic activities of mitochondrial inner membrane complexes II and IV were partially restored by NB-DNJ. Conclusion: Our study demonstrates that NB-DNJ as a potential therapeutic agent for NPC patients reverses membrane lipid abnormalities in NPC fibroblasts. Conflict of Interest declared.