Faculty Bibliography

A 12year-old female presented with a seven-year history of progressive muscle weakness, atrophy, tremor and fasciculations. Cognition was normal. Rectal biopsy revealed intracellular storage material and biochemical testing indicated low hexosaminidase activity consistent with juvenile-onset G(M2)-gangliosidosis. Genetic evaluation revealed compound heterozygosity with two novel mutations in the hexosaminidase beta-subunit (c.512-3 C>A and c.1613+15_1613+18dup). Protein analysis was consistent with biochemical findings and indicated only a small portion of beta-subunits were properly processed. These results provide additional insight into juvenile-onset G(M2)-gangliosidoses and further expand the number of beta-hexosaminidase mutations associated with motor neuron disease.

Fabry disease (FD) is an X-linked hereditary defect of glycosphingolipid storage caused by mutations in the gene encoding the lysosomal hydrolase alpha-galactosidase A (GLA, alpha-gal A). To date, over 400 mutations causing amino acid substitutions have been described. Most of these mutations are related to the classical Fabry phenotype. Generally in lysosomal storage disorders a reliable genotype/phenotype correlation is difficult to achieve, especially in FD with its X-linked mode of inheritance. In order to predict the metabolic consequence of a given mutation, we combined in vitro enzyme activity with in vivo biomarker data. Furthermore, we used the pharmacological chaperone (PC) 1-deoxygalactonojirimycin (DGJ) as a tool to analyse the influence of individual mutations on subcellular organelle-trafficking and stability. We analysed a significant number of mutations and correlated the obtained properties to the clinical manifestation related to the mutation in order to improve our knowledge of the identity of functional relevant amino acids. Additionally, we illustrate the consequences of different mutations on plasma lyso-globotriaosylsphingosine (lyso-Gb3) accumulation in the patients' plasma, a biomarker proven to reflect the impaired substrate clearance caused by specific mutations. The established system enables us to provide information for the clinical relevance of PC therapy for a given mutant. Finally, in order to generate reliable predictions of mutant GLA defects we compared the different data sets to reveal the most coherent system to reflect the clinical situation.

Alexander disease (AxD) is a usually fatal astrogliopathy primarily caused by mutations in the gene encoding glial fibrillary acidic protein (GFAP), an intermediate filament protein expressed in astrocytes. We describe three patients with unique characteristics, and whose mutations have implications for AxD diagnosis and studies of intermediate filaments. Patient 1 is the first reported case with a noncoding mutation. The patient has a splice site change producing an in-frame deletion of exon 4 in about 10% of the transcripts. Patient 2 has an insertion and deletion at the extreme end of the coding region, resulting in a short frameshift. In addition, the mutation was found in buccal DNA but not in blood DNA, making this patient the first reported chimera. Patient 3 has a single-base deletion near the C-terminal end of the protein, producing a short frameshift. These findings recommend inclusion of intronic splice site regions in genetic testing for AxD, indicate that alteration of only a small fraction of GFAP can produce disease, and provide caution against tagging intermediate filaments at their C-terminal end for cell biological investigations. Hum Mutat 33:1141-1148, 2012. (c) 2012 Wiley Periodicals, Inc.

Cerebral micro- and macro-vasculopathy have been described in Fabry disease (FD). Neuronal globotriaosylceramide accumulation in selective cortical and brain stem areas including the hippocampus has been reported by autopsy studies in FD, but clinical surrogates as well as the clinical relevance of these findings have not been investigated so far. We measured the hippocampus volumes in a group of clinically affected patients with FD and correlated the findings with the cognitive performance of the patients. Hippocampal volumes were determined manually on T1-weighted MR-images of 25 FD patients (age 36.5 +/- 11.0 years) and 20 age-matched controls. Additionally, individual white matter (WM) and gray matter (GM) volumes were measured using brain segmentation analyses. After controlling for age, white matter lesion (WML) volume, and WM/GM-volumes hippocampal volumes were significantly decreased in FD. These findings were substantially more pronounced in a subgroup of men with FD. WM and WM/GM volumes, and memory function did not significantly differ between patients and controls. In patients with FD hippocampal volumes were neither significantly correlated to WML volume nor to WM or WM/GM volumes. Hippocampus atrophy was not driven by the WML or other brain tissue atrophy and seems to correlate with the neuronal involvement in FD. In this young to middle-aged Fabry cohort the hippocampus degeneration was functionally compensated without memory impairment. Longitudinal studies are needed to determine whether this degenerative component in FD will progress and, in concert with the individual WML-load, predict subsequent cognitive decline

Introduction: To define the phenotypic spectrum of hematological and visceral disease in patients with type 3 Gaucher disease (GD3) and its response to alglucerase/imiglucerase therapy. Description/Results: All patients <18 years of age who were enrolled in the International Collaborative Gaucher Group (ICGG) Gaucher Registry as of July 2010 with a diagnosis of GD3 were included in this analysis. Subsets of patients with baseline status of anemia, thrombocytopenia, splenomegaly, hepatomegaly, or height zscore<=- 1 were identified. Outcomes of hemoglobin concentration, platelet count, and hepatic and splenic volumes and height Z-scores were assessed at baseline and up to 5 years following initiation of therapy using non-linear mixed effects models. A total of 334 GD3 patients were identified; the majority of patients were from the Middle East (34%), Europe (29%) and the USA (16%). Ninety percent of the patients were diagnosed at age <6 years and 81% of patients initiated therapy at age <6 years. GBA1 gene analysis revealed a preponderance of L444P mutations: L444P homozygotes (63%) and L444P compound heterozygotes (16%). Among the subset with height Z-score<=-1 at baseline (n=159), the mean baseline height Z-score was -2.7. Improvement in height was seen after both 1 (mean height Z-score -2.5) and 5 years of treatment (mean height Z-score -1.7). In the subset with anemia at baseline (n=145), anemia was present in 42% within <=1 year of treatment initiation, and declined to 29% after 5 years of treatment. For the subset with thrombocytopenia (platelet counts <120 103/ mm3) at baseline (n=131), thrombocytopenia was present in 57% within less than 1 year, and declined to 11% after 5 years of treatment. Liver and spleen volumes decreased over the 5 years of treatment. Conclusions: There is early onset of prominent visceral and hematologic disease in GD3 patients before the age of 6 years. Moreover, the patients exhibit striking growth failure. These effects are reversed by alglucerase/imiglucerase treatment within 5 years. This cohort represents the largest cohort of children with GD3 ever described to delineate the phenotypic spectrum and its response to alglucerase/imiglucerase therapy

OBJECTIVE: In untreated Fabry patients without overt autonomic dysfunction and normal baroreflex sensitivity (BRS) at rest, BRS is impaired during orthostatic, sympathetic challenge but normalizes after enzyme-replacement therapy (ERT) (Hilz et al., J Hypertens 2010; 28:1438-1448). This study evaluated BRS during parasympathetic challenge with six cycles per minute metronomic deep breathing (MDB) in Fabry patients before and after ERT. METHODS: In 22 Fabry patients (28 +/- 8years), we monitored RR-intervals (RRIs), SBP, and respiratory frequency during spontaneous breathing (spont_breath) and MDB, before and after 18 (11 patients) or 23 months (11 patients) of biweekly ERT (1.0 mg/kg agalsidase beta). We determined spectral powers of mainly sympathetic low-frequency (0.04-0.15 Hz) RRI fluctuations, parasympathetic high-frequency (0.15-0.5 Hz) RRI fluctuations, sympathetically mediated low-frequency powers of SBP and high-frequency powers of SBP. We calculated BRS (ms/mmHg) during spont_breath and MDB as low-frequency-high-frequency alpha index (coherence >0.5). We compared parameters during spont_breath and MDB within and between patients before and after ERT and 15 age-matched (27 +/- 5years) healthy men (RANOVA and posthoc analysis; significance: P < 0.05). RESULTS: During spont_breath and MDB, parameters were similar between groups. Within the three groups, RRIs were lower, whereas RRI low-frequency powers and SBP low-frequency powers were higher during MDB than during spont_breath. BRS was similar during MBD and spont_breath in untreated patients (P > 0.05), but increased significantly with MDB in patients after ERT (P = 0.048) and in controls (P = 0.035). CONCLUSION: In untreated Fabry patients, MDB uncovers impaired BRS. After 18 or 23 months of ERT, MDB-induced BRS increase is similar in Fabry patients and controls, demonstrating that ERT not only restores sympathetic but also parasympathetic baroreflex activation

Lysosomal integral membrane protein type 2 (LIMP-2) is responsible for proper sorting and lysosomal targeting of glucocerebrosidase, the enzyme deficient in Gaucher disease (GD). Mutations in the gene for LIMP-2, SCARB2, are implicated in inherited forms of myoclonic epilepsy, and myoclonic epilepsy is part of the phenotypic spectrum associated with GD. We investigated whether SCARB2 mutations impact the Gaucher phenotype focusing on patients with myoclonic epilepsy, including a pair of siblings with GD who were discordant for myoclonic seizures. Sequencing of SCARB2 genomic and cDNA identified a heterozygous, maternally inherited novel mutation, c.1412A>G (p.Glu471Gly), in the brother with GD and myoclonic epilepsy, absent from his sibling and controls. Glucocerebrosidase activity, Western blots, real-time PCR, and immunofluorescence studies demonstrated markedly decreased LIMP-2 and glucocerebrosidase in cells from the sibling with (p.Glu471Gly) LIMP-2, and diminished glucocerebrosidase in lysosomes. The cells secreted highly glycosylated enzyme and showed mistrafficking of glucocerebrosidase. Sequencing of SCARB2 in 13 other subjects with GD and myoclonic epilepsy and 40 controls failed to identify additional mutations. The study provides further evidence for the association of LIMP-2 and myoclonic epilepsy, explains the drastically different phenotypes encountered in the siblings, and demonstrates that LIMP-2 can serve as a modifier in GD. Hum Mutat 32:1232-1238, 2011. (c)2011 Wiley Periodicals, Inc

A large number of mutations, and several polymorphisms, have been characterized in the GBA gene, encoding the lysosomal enzyme glucocerebrosidase, the activity of which is impaired in Gaucher disease. In this communication we summarize published and new data concerning biochemical characterization of the E326K amino acid change (1093G>A in the GBA1 cDNA) in tissue culture and its association with Parkinson disease, suggesting it is a disease causing mutation and not merely a polymorphism in the GBA gene

The G(M2) gangliosidoses are a group of lysosomal storage diseases caused by defects in the genes coding for the enzyme hexosaminidase or the G(M2) activator protein. Four Jacob sheep from the same farm were examined over a 3-year period for a progressive neurologic disease. Two lambs were 6-month-old intact males and 2 were 8-month-old females. Clinical findings included ataxia in all 4 limbs, proprioceptive deficits, and cortical blindness. At necropsy, the nervous system appeared grossly normal. Histologically, most neurons within the brain, spinal cord, and peripheral ganglia were enlarged, and the cytoplasm was distended by foamy to granular material that stained positively with Luxol fast blue and Sudan black B stains. Other neuropathologic findings included widespread astrocytosis, microgliosis, and scattered spheroids. Electron microscopy revealed membranous cytoplasmic bodies within the cytoplasm of neurons. Biochemical and molecular genetic studies confirmed the diagnosis of G(M2) gangliosidosis. This form of G(M2) gangliosidosis in Jacob sheep is very similar to human Tay-Sachs disease and is potentially a useful animal model