Introduction: The AB-variant form of GM2-gangliosidosis is due to adeficiency of GM2-activator protein (GM2AP). Only five mutations havebeen described in the GM2AP gene (GM2A). This report describes a childof Indian non-consanguinous parentage homozygous for a novel GM2Amutation.

Case history: A 15-month-old girl was born with oligohydramnios fourweeks premature by C-section. She sat unsupported at 7 months andcrawled at 11 months but failed to develop further. Seizures began at 15months. She had easy startle and repeated URIs. The head circumferencewas at the 5th percentile, and a macula cherry-red spot and generalizedhypotonia were present. Lysosomal enzymes including hexosaminidasesA and B, 2-galactosidase and 1-neuraminidase were normal. Electronmicroscopy of a skin biopsy showed lysosomal storage.

Methods: Total RNA was isolated from cultured skin fibroblasts andcDNA isolated by RT-PCR. Purified DNA fragments were directlysequenced with either sense or anti-sense primers and ‘‘big dye terminatorchemistry’’ (Applied Biosystems, CA) using an ABI 3730 DNA analyzer.

Results: A missense mutation (CTGCGG) at nucleotide base 521 ofthe GM2A gene was found in both alleles of the patient. This is predictedto create a leucine to arginine substitution in codon 174.

Conclusion: The new mutation is in a phylogenetically conservedregion of the gene and therefore a likely cause of GM2-gangliosidosis inthis child. Further proof of this sequence alteration as the disease-causingmutation will require expression studies for GM2AP.

doi:10.1016/j.ymgme.2007.10.067

56. GM1-gangliosidosis in an American Black Bear

Edwin Kolodny a, Brian Frankel a, Paola Torres a, Joseph Alroy b,

Srinivasa Raghavan a, a New York University School of Medicine, New

York, New York, USA, b Tufts University School of Medicine and

Veternary Medicine

Introduction: A female bear approximately 15 months old was broughtto the Tufts Cummings School of Veternary Medicine because of lethargy,dehydration, body tremors and ataxia. Blood analyses disclosed hypercho-lesterolemia and cytoplasmic vacuoles in monocytes and lymphocytes. Abrain MRI revealed dysmyelination. The animal was euthanized andpost-mortem studies performed showing enlarged neurons staining posi-tive with PAS and Luxol fast blue.

Hypothesis: A neuronopathic form of lipid storage disease wassuspected.

Methods: Total lipid extraction was performed on formalin-fixed brainof the affected animal and an age-matched control. Total gangliosideswere isolated on a BondElutC18 cartridge, total lipid sialic acid deter-mined, then separated by HPTLC. Individual ganglioside bands were visu-alized with resorcinol and quantitated by densitometry. Neutral lipidswere separated by silicic acid chromatography and the glycosphingolipidsfurther fractionated by HPTLC.

Results: Lipid-bound sialic acid in affected brain was elevated 6.6-fold with a prominent band of resorcinol-positive material co-migrat-ing with GM1-ganglioside. TLC of the neutral glycosphingolipid frac-tion showed reduction of cerebroside and sulfatide in the affected bear.No difference in free cholesterol was seen between the affected andcontrol specimens. Frozen material was not available to determine 2-galactosidase activity; however, DNA analysis of the bear 2-galactosi-dase gene is in progress.

Conclusion: GM1-gangliosidosis has been reported in cats, dogs,sheep, and calves. This is the first documented case of bear GM1-gangli-osidosis. Two additional cases of cerebral lipid storage among New Eng-land black bears known to J.A. suggest a possible founder effect withinthis population.

doi:10.1016/j.ymgme.2007.10.068

57. Quantifying the rate of neurologic decline in Batten disease in clinical

trials

Jennifer Kwon, Frederick Marshall, Heather Adams, Paul Rothberg, Amy

Vierhile, Elisabeth deBlieck, David Pearce, Jonathan Mink, University of

Rochester Medical Center, Rochester, NY, USA

Introduction: The Unified Batten Disease Rating Scale (UBDRS) wasdeveloped to measure physical, behavioral and functional capabilitydomains in patients suffering from Batten disease (neuronal ceroid lipofus-cinosis, NCL) a neurodegenerative lysosomal storage disorder. The UBDRShas been evaluated in subjects with NCL since 2002 and has been refined overthe years to improve its reliability and validity. Now that therapies are beingproposed to prevent, slow, or reverse the course of NCL, the UBDRS willplay an important role in assessing clinical outcomes in research trials.

Hypothesis: The UBDRS can be used to predict the rate of decline injuvenile-onset NCL (JNCL).

Methods: The UBDRS has been administered to nearly 70 patients withBatten disease of whom 55 have JNCL. Most subjects have had serialresearch evaluations. From these data, the rate of physical and motor declinehas been calculated in the 48 subjects whose JNCL has been genetically con-firmed. Of these, 20, 9, 8, 1, 5, and 5 subjects have been evaluated over 1, 2, 3,4, 5, and 6 years, respectively. The rate of decline over time was quantifiedusing multivariate linear regression and repeated measures analysis.

Results: The UBDRS physical disability subscales show decline overtime that proceeds at a quantifiable linear rate in the years following initialonset of clinical symptoms.

Conclusions: The UBDRS is a reliable and valid instrument that mea-sures clinical progression in JNCL. Our data support use of the UBDRSin clinical trials of novel therapies in JNCL.

doi:10.1016/j.ymgme.2007.10.069

58. Reproductive fitness in individuals affected by Fabry disease

Dawn Laney a, Daniel Gruskin b, Ami Mehta c, a Emory University,

Decatur, GA, USA, b Genzyme Corporation, c Sarah Lawrence College

Objective: To compare the reproductive fitness of males and femalesaffected by Fabry disease as defined by production of liveborn offspring.

Methods: A retrospective review of pedigrees constructed during clin-ical appointments by genetic counselors and medical geneticists was con-ducted for males and females affected by Fabry disease and of optimalchildbearing age (15–44 years).

Results: Females affected by Fabry disease (n = 32) reported having 34biological children (17 male and 17 female). Males affected by Fabry disease(n = 25) reported having 19 biological children (10 female and 9 male).

Conclusion: Males of optimal childbearing age affected by Fabry dis-ease have a reduced rate of liveborn children (0.76 children per male onaverage) as compared to females in the same age range affected by Fabrydisease (1.1 children per female on average). While further work is neededto determine if this holds true in a larger population, our data suggeststhat males with Fabry disease have decreased reproductive fitness com-pared to females; this might be due to worse general health status, azosper-mia, depression, reproductive choice, or other undetermined causes. Inaddition to playing a role in patient and family counseling, this findingmight have epidemiologic implications in regards to the expected numberof affected females, and also could provide further impetus to early treat-ment of Fabry disease.

doi:10.1016/j.ymgme.2007.10.070

59. Inhibition of the nicotinic acid adenine dinucleotide phosphate receptor

induces a novel lysosomal storage disease

Alexander Lewis, Emyr Lloyd-Evans, Frances Platt, Grant Churchill,

University of Oxford, Oxford, Oxon., UK

S28 Abstracts / Molecular Genetics and Metabolism 93 (2008) S14–S46

NAADP (nicotinic acid adenine dinucleotide phosphate) is a novelCa2+-releasing second messenger. Although it is the most potent knownCa2+-releasing agent, its receptor is currently unidentified. Nevertheless,there is evidence that NAADP is able to release Ca2+ from lysosomes.We therefore investigated the possibility that the NAADP receptor isimportant in lysosomal function. Inhibition of the NAADP receptor,using the recently identified selective antagonist Ned-14, produced exten-sive dysfunction in the endocytic pathway. This was demonstrated byvastly increased loading of both lysotracker and rhodamine dextran, aswell as by the existence of a remarkably abnormal lysosomal morphologyon electron microscope images, demonstrating extensive lysosomal stor-age. As Ned-14 is fluorescent, it was possible to confirm that Ned-14 bind-ing co-localised with lysosomes loaded with lysotracker or rhodaminedextran. Further analysis of the lysosomal dysfunction by means of fluo-rescent markers, antibody staining and high performance liquid chroma-tography demonstrated that the lysosomal dysfunction includes storageof cholesterol, sphingomyelin and glycosphingolipids. BODIPY-LacCerlabelling of cells treated with Ned-14 demonstrated that the storage dis-ease arose due to inhibition of transport out of lysosomes. This has beenconfirmed with fusion assays. This is the first study to demonstrate a rolefor the NAADP receptor in endocytosis. Furthermore, this work not onlydemonstrates the possibility of a lysosomal storage disease resulting fromNAADP receptor dysfunction, but also raises the possibility that theNAADP receptor may be involved in already identified diseases.

doi:10.1016/j.ymgme.2007.10.071

60. Gaucher Registry: A comparison among Argentina, Brazil and Colombia

2007 country reports

Adriana Linares a, Aurora Feliu b, Elisa Sobreira c, a Universidad Nacional

de Colombia, Bogota, DC, Colombia, b Hospital Garrahan, Buenos Aires,

Argentina, c Santa Casa de Sao Paulo, Brazil

Introduction: The International Collaborative Gaucher Group(ICGG) Gaucher Registry is a global outcomes assessment and diseasemanagement program. Scientific direction for the Gaucher Registry is pro-vided by an independent board of regional and international advisors whoare active participants in the Registry. In 2005, the Board of Latin Amer-ica Registry Coordinators was established. Currently, Argentina, Braziland Colombia have the largest number of enrolled patients in the GaucherRegistry.

Objective and methods: The objective of this analysis was to comparedemographic, clinic and genetic data for these three countries entered intothe Gaucher Registry as of April, 2007.

Results: Most patients were diagnosed with type 1 Gaucher diseaseand there was a higher proportion of females in these three cohorts. Themajority of the patients were diagnosed between birth and age youngerthan 20 years and more than 74% have received enzymatic replacementtherapy (ERT) with imiglucerase.

Genotype data for this population indicate that N370S/Rare allele wasthe most frequent genotype in Brazil and Colombia, followed by N370S/L444P, which was the most frequent genotype in Argentina. Comparing toBrazil and Colombia, Argentina had the largest population (22%) withN370S/N370S genotype.

By the first infusion date, Argentinean and Brazilian cohorts differedfrom Colombian cohort, respectively, in higher proportion of anemia(53% and 53% versus 21%). Brazilian and Colombian cohorts differedfrom Argentinean cohort, respectively, in higher proportion of moderatethrombocytopenia (52% and 64% versus 34%), severe splenomegaly(63% and 62% versus 47%), bone pain (60% and 42% versus 28%), and evi-dence of any bone disease (81% and 94% versus 61%). Pediatric growthretardation was observed in more than 50% of patients in all threecountries.

The majority of Argentinean and Brazilian type 1 patients, who havereceived ERT, started receiving the mean dose of 30 U/kg/2 weeks andthis dose remained relatively stable through 10 years; in contrast, more

than 50% of Colombian type 1 patients received the mean dose of 60 U/kg/2 weeks during the same period. Patients receiving ERT with imiglucer-ase showed increased hemoglobin concentrations after 1 year of treatmentand stable levels were maintained for 5 years. The median plateletresponse to ERT increased into the normal range after 1 year of treatmentand this improvement was sustained for 5–10 years. Organomegaly datashowed that median spleen volumes decreased faster among Argentineanand Colombian patients, from severe splenomegaly to moderate or mildsplenomegaly in 1 year and, among Brazilian patients in 2 years ofERT. The three cohorts improved median liver volumes in the first yearof imiglucerase treatment, from moderate hepatomegaly to mild hepato-megaly or normal volumes, and this response was sustained for 5–10 years.

Conclusion: By comparing data of Gaucher patients from Argentina,Brazil and Colombia as reported to the Gaucher Registry, phenotypicand genetic similarities as well as differences among Gaucher patients fromthese three South American countries were elucidated.

The ICGG Gaucher Registry is supported by Genzyme Corporation.

doi:10.1016/j.ymgme.2007.10.072

61. Smith-Lemli-Opitz syndrome: A closet lysosomal storage disease hiding

within an inheritable metabolic disorder of cholesterol biosynthesis

Emyr Lloyd-Evans a, Jinzhi Zhang a, Forbes D. Porter b, Fran Platt a,a University of Oxford, Oxford, Oxon., UK, b National Institutes of Health,

UK

Smith-Lemli-Opitz syndrome (SLOS) is an inherited metabolic diseasein which the cholesterol biosynthetic enzyme (32-hydroxysterol 7-reduc-tase) is defective. This causes accumulation of the cholesterol precursor,7-dehydrocholesterol (7-DHC). A potential therapy for SLOS patients isto increase dietary cholesterol intake, surprisingly, this is of limited clinicalbenefit.

Recent work has shown that 7-DHC accumulation in SLOS causesdefective transport of exogenous cholesterol from lysosomes to the endo-plasmic reticulum (ER) where it could be utilised. This causes cholesterolstorage in lysosomes, mimicking another lipid transport disorder, Nie-mann-Pick disease type C1 (NPC1). Thus, one reason that SLOS patientscannot correctly utilize dietary cholesterol is a lack of transport to the ER.

7-DHC is a secondary amphiphile, a group of molecules capable ofinducing an NPC1-like lysosomal storage phenotype. We now show thataccumulation of 7-DHC, potentially interfering with NPC1 protein func-tion, in SLOS cells and tissues leads to lysosomal storage of multiplesphingolipids that ultimately inhibits cholesterol efflux from lysosomesto the ER.

Treating SLOS cells with NB-DGJ, to reduce sphingolipid biosynthesis(ultimately reducing sphingolipid storage), clears lysosomal storage bodies(shown by EM) and corrects the SLOS lysosomal transport defect, result-ing in delivery of cholesterol to the ER for utilisation. SLOS patients maytherefore benefit from sphingolipid lowering drugs.

This work indicates that SLOS is a new and novel lysosomal storagedisease with storage resulting from a primary defect in cholesterolbiosynthesis.

doi:10.1016/j.ymgme.2007.10.073

62. Abnormal lysosomal calcium homeostasis in mucolipidosis type IV

Emyr Lloyd-Evans, Ksenia Peterneva, Alexander Lewis, Grant Churchill,

Fran Platt, University of Oxford, Oxford, Oxon., UK

Mucolipidosis type IV (MLIV) is a rare inherited neurodegenerativelysosomal storage disorder, characterised by early onset, psychomotorretardation and developmental delay. Lysosomal storage, primarily ofsphingolipids and phospholipids, is present in all tissues and cells. Nohydrolytic enzyme defects have been associated with MLIV (despite

Abstracts / Molecular Genetics and Metabolism 93 (2008) S14–S46 S29