Long Term Bone Marrow Responses, as Measured by Quantitative Chemical Shift Imaging (QCSI) MRI, Following Treatment With Taliglucerase Alfa in Patients with Type 1 Gaucher Disease

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Abstracts / Molecular Genetics and Metabolism 105 (2012) S15–S69

index is significant because it will provide the background against which future therapies can be compared. Our study is unique in that it adds evaluations of neurocognitive and neuroimaging as landmarks of neurodegeneration, enhancing the overall evaluative analysis. Methods: For Tay-Sachs and Sandhoff diseases, we are collecting longitudinal clinical and QOL data that characterize both the developmental and disease course for these patients and aids in the development of a disease stage and severity scale. We are also assessing any therapeutic regimens, mutations, and concomitant medication therapy that may play a role in influencing and/or altering disease course. This longitudinal data will be collated with retrospective QOL data collected at Massachusetts General Hospital to generate statistically meaningful results. For patients with late-onset Tay-Sachs (LOTS) we are evaluating the underlying structural and functional abnormalities of the central nervous system to identify any measured changes in these systems over time. This is accomplished by conducting an annual MRI and neurodevelopmental testing in addition to monitoring clinical changes. Status: The NIH and the DMCC have approved this project; electronic case report forms have been promoted and activated. This study is currently enrolling participants. doi:10.1016/j.ymgme.2011.11.164

MPS I: Monitoring Plasma And Urine Levels Of Dermatan And Heparan Sulfate During Enzyme Replacement Therapy L. van der Tol a, M.H. de Ru a, N. van Vlies b, A.H.A. Wagemans b, L. IJlst b, F.A. Wijburg a, aDepartment of Pediatrics and Amsterdam Lysosome Center ‘Sphinx’, Academic Medical Center, Amsterdam, Noord-Holland, The Netherlands, bLaboratory of Genetic Metabolic Diseases, Academic Medical Center, Amsterdam, The Netherlands

Niemann-Pick Disease Type C: Significant Clinical Discordance Between Siblings Linda van der Tol a, Marc Engelen b, Frits A. Wijburg a, aDepartment of Pediatrics and Amsterdam Lysosome Center ‘Sphinx’, Academic Medical Center, Amsterdam, Noord-Holland, The Netherlands, bPediatric Neurology, Department of Pediatrics, Academic Medical Center, Amsterdam, The Netherlands Niemann-Pick disease type C (NPC) is a rare lysosomal storage disease, caused by impairment of intracellular lipid transport. Age of onset and symptoms are highly variable and include systemic, neurological and psychiatric manifestations. Increased identification of NPC patients revealed significant heterogeneity even within families, which is important for adequate counseling and early diagnosis. We report the discordant manifestations of NPC in a family with 2 affected siblings. The younger sister presented at the age of 15 with frequent falls and developed a cerebellar ataxia, vertical supranuclear gaze palsy (VSGP), dysphagia, dysarthria and seizures. NPC was diagnosed by filipin staining of fibroblast and mutation analysis. Later she developed a psychosis. She showed a continuous neurological decline and died at age 23. The 3 years older brother was not screened following diagnosis of his sister as he was asymptomatic. He experienced social difficulties at age 17 and a diagnosis of Asperger syndrome was made. He started with his studies on English Language at the University; however, social and cognitive difficulties arose more frequently. At the age of 23 he experienced two psychotic episodes. At age 26 he developed ataxia, dysarthria and VSGP, leading to the diagnosis of NPC. Miglustat treatment was started. These cases illustrate the necessity of awareness of significant intra-familial variations in the clinical course of NPC. Further research could lead to the discovery of modifier genes and allow us to understand this remarkable intra-familial heterogeneity. doi:10.1016/j.ymgme.2011.11.166

Introduction: Mucopolysaccharidosis type I (MPS I) is a lysosomal disorder caused by a deficiency of the enzyme alpha-L-Iduronidase, leading to accumulation of the glycosaminoglycans (GAG) heparan sulfate (HS) and dermatan sulfate (DS). Enzyme replacement therapy (ERT) can significantly ameliorate somatic symptoms. Although it has been established that ERT decreases GAG accumulation, possible fluctuation of DS and HS concentration related to the timing of ERT has not been assessed. Methods: Samples were collected at different time points related to weekly ERT, in three non-consecutive weeks in seven MPS I patients (phenotypes: one Hurler, two Hurler-Scheie, four Scheie), treated with ERT for at least 2 years. Urine (early morning voids) was collected daily and plasma was collected immediately before (t = 0), two and four days after ERT. Concentrations of HS and DS were determined after enzymatic digestion to disaccharides by HPLC MS/ MS. Anti-iduronidase antibody levels were measured. Results: There are significant inter-individual differences in HS and DS levels in patients on ERT, mostly distinguishing Hurler and Hurler-Sheie from the Scheie phenotype, where HS likely correlates better with the phenotype than DS. Differences may also be related to antibody levels. Urinary GAG excretion (DMB assay) does not correlate to HS and DS levels in all patients. Hence, a patient with a normal urinary GAG excretion can have high values of HS and DS in plasma. Not all data have been analyzed, final results and conclusions will be presented at the WORLD symposium. Our results may reveal important information for future dose optimization studies. doi:10.1016/j.ymgme.2011.11.165

Long Term Bone Marrow Responses, as Measured by Quantitative Chemical Shift Imaging (QCSI) MRI, Following Treatment With Taliglucerase Alfa in Patients with Type 1 Gaucher Disease Laura van Dussen a, Carla Hollak a, Ari Zimran b, Milan Petakov c, Hanna Rosenbaum d, David Aviezer e, Einat Brill-Almon e, Raul Chertkoff e, Mario Maas a, aUniversity of Amsterdam, Amsterdam, The Netherlands, bGaucher Clinic, Shaare Zedek Medical Center, Hebrew University and Hadassha Medical School, Jerusalem 91031, Israel, c Clinical Center of Serbia, Institute of Endocrinology, Diabetes and Metabolic Disease, Belgrade, Serbia, dHaematology Ambulatory Services, Rambam Medical center, Haifa 31096, Israel, eProtalix Biotherapeutics, Carmiel, Israel Taliglucerase alfa is a carrot-cell-expressed recombinant betaglucocerebrosidase formulation developed as a treatment for Gaucher disease (GD). In a pivotal, double-blind, randomized Phase III study, PB-06-001 and its extension study PB- 06–003 , designed to evaluate safety and efficacy of Taliglucerase alfa, two dose levels, 30 and 60 units/kg, were compared in treatment naïve patients (1). As bone disease is one of the most debilitating features of GD, quantification of bone marrow involvement is of importance for monitoring the response to treatment. For this purpose, bone marrow fat fraction (Ff) measured by Quantitative Chemical Shift Imaging (QCSI) was included as an exploratory parameter in a subpopulation of this cohort. Study objective: To asses bone marrow response by QCSI in GD patients treated with taliglucerase alfa up to 36 months. Method:

Abstracts / Molecular Genetics and Metabolism 105 (2012) S15–S69

Eight GD patients with intact spleens were treated with 60U/kg (n = 4) or 30 U/kg (n = 4) biweekly. QCSI results were compared to results in 15 untreated GD patients with a follow-up interval of 1 year extracted from the Dutch Gaucher database. Results: Five taliglucerase treated patients had a decreased Ff (< 23%) at baseline (median (n = 8): 19%, range 11-35%). Ff significantly increased compared to baseline (p = 0.012) and compared to untreated patients (p = 0.007) already after 1 year of followup with further improvement up to 36 months (median absolute increase at max. follow-up 13.5% (range 5-29%). No difference for the two dose groups was established. Conclusion: Treatment with taliglucerase alfa results in significant increases in bone marrow fat fractions. (1) Zimran A, Brill-Almon E, Chertkoff R, Petakov M, Blanco-Favela F, Terreros ME, et al. Pivotal trial with plant-cell-expressed recombinant glucocerebrosidase, taliglucerase alfa, a novel enzyme replacement therapy for Gaucher disease. Blood 2011 Sep 6. doi:10.1016/j.ymgme.2011.11.167

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glycosaminoglycan (GAG) degradation. The subsequent GAG accumulation in tissues is responsible for various pathological features like mental retardation, short stature, cardiorespiratory complications, joint stiffness and dysostosis multiplex. Although aberrant bone remodeling and growth plate abnormalities with disorganized trabecular morphology were recently described in MPS I mice, the cellular turnover in bone marrow (BM) remains unknown. Thus, the aim of our study was to analyze murine MPS I BM hematopoiesis. Three-month-old MPS I and wild-type (WT) mice were euthanized and BM cells were extracted from right/anterior femora. The cell pellets were resuspended and analyzed through flow citometry for each subpopulation. Myeloid common progenitors and natural-killer cells were significantly decreased in MPS I compared to WT mice (p < 0.01). In addiction, activated B-lymphocytes were also decreased in MPS I group (p < 0.05). However, no alterations were found in peripheral blood cells count (hemogram). These results suggest that bone abnormalities found in murine MPS I can impair effective medullary hematopoiesis and possibly justify bone alterations presented by patients and other pathological processes manifested locally and systemically in this disease. Financial support: FAPESP, CNPq, IGEIM and AFIP.

Genome-Wide RNAi Screen For Lysosomal Storage Disorders Arash Velayati a, Pinar Tuzmen b, Rajarshi Guha b, Scott Martin b, Ehud Goldin a, Ellen Sidransky a, aMedical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD, USA, bRNAi Screening Core Facility, NIH Chemical Genomics Center, National Human Genome Research Institute, National institutes of Health, USA Untangling the web of genes contributing to the phenotypic diversity in monogenic disorders will contribute to our understanding of disease mechanisms, provide insight into complex genetic disorders and lead to new therapeutic options. We performed genome-wide RNAi screening to probe multiple genes in well defined pathways, to assess their effect on function and to identify novel protein interactions for three common lysosomal storage diseases (LSD), Gaucher disease (GD), Fabry disease and Pompe disease. RNAi libraries available from the NIH RNAi screening core facility were printed in 384-well plates and transfected into fibroblast and BE(2)-M17 neuroblastoma cell lines using standard methods. Activity of each selected lysosomal enzyme was assayed in cell lysates 72 hr after transfection using a fluorescent substrate. Standard normalization and data analysis protocols were used for hit selection. Candidate genes identified were confirmed by follow-up screens and further by evaluated to analyze their effect on cell viability, lysosome function and integrity, global gene expression and intracellular transport. The RNAi screening data from distinct LSD were compared to identify shared genes of interest, which may have direct relevance to lysosome biogenesis and function. Specific genes relevant to individual LSD were explored as potential therapeutic targets. The Next, cell based assays using fibroblast lines from patients with GD and other LSD will be utilized to investigate gene function. doi:10.1016/j.ymgme.2011.11.168

Impaired Medullary Hematopoiesis in Murine Mucopolysaccharidosis Type I Gustavo Viana, Edgar Julian Paredes-Gamero, Ana Maria Martins, Vânia D'Almeida, Universidade Federal de São Paulo (UNIFESP), São Paulo, São Paulo, Brazil Mucopolysaccharidosis type I (MPS I) is an autossomal recessive disease caused by alpha-L-iduronidase deficiency, leading to impaired

doi:10.1016/j.ymgme.2011.11.169

Intracerebroventricular (ICV) Recombinant Human Tripeptidyl Peptidase-1 (rhTPP1) Enzyme Replacement Attenuates Disease Progression in a Canine Model of Late Infantile Neuronal Ceroid Lipofuscinosis (LINCL) Brian Vuillemenot a, Martin Katz b, Joan Coates b, Derek Kennedy a, Camille Flournoy b, Christine Sibigtroth b, Rebecca Whiting b, Randal Reed c, Eric Adams c, Laurie Tsuruda a, Donald Musson a, Charles O'Neill a, aBioMarin Pharmaceutical Inc., Novato, CA, USA, bUniversity of Missouri, Columbia, MO, USA, cNorthern Biomedical Research, Inc., Muskegon, MI, USA LINCL is caused by lack of the enzyme TPP1. LINCL patients exhibit accumulation of lysosomal storage in the CNS accompanied by neurodegeneration, loss of function, and death. TPP1-null dachshunds recapitulate many symptoms of the human disease. This study was performed to determine the pharmacology of ICV rhTPP1 in this model. An additional objective was to characterize the pharmacokinetics (PK) and distribution in the CNS. Affected animals (N = 9) and wild-type controls (N = 9) received ICV infusions of 4 or 16 mg rhTPP1 or artificial cerebrospinal fluid (CSF) vehicle. rhTPP1 was administered as two or four hour infusions via a catheter implanted in a lateral ventricle. Animals received approximately 20 biweekly doses starting at age 2 months. Serial plasma and CSF samples were collected during and after dose administration to characterize rhTPP1 PK. Elevated CSF concentrations were observed for at least 48 hours after infusion and were approximately 1000-fold higher than plasma levels. Neurological and clinical examinations, electretinography, measurement of pupillary light reflexes and visual evoked potentials, magnetic resonance imaging, and cognitive testing were performed throughout the study to assess disease progression. Necropsy occurred 48 hours after the final dose at approximately 11 months of age. Animals given 4 mg doses of rhTPP1 exhibited improved clinical signs and attenuated functional decline compared to vehicle treated controls. In-life assessments at the 16 mg dose level, as well as analysis of CNS tissues for rhTPP1 concentration, storage material, and markers of neuronal injury, are ongoing. doi:10.1016/j.ymgme.2011.11.170