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BMJ Case Reports Jun 2017
Topics: Brain; Developmental Disabilities; Fatal Outcome; Humans; Infant; Magnetic Resonance Imaging; Male; Microcephaly; Multimodal Imaging; Sandhoff Disease; Tomography, X-Ray Computed; beta-Hexosaminidase alpha Chain; beta-Hexosaminidase beta Chain
PubMed: 28647713
DOI: 10.1136/bcr-2017-220912 -
Neuroscience Letters Nov 2021
Topics: Age of Onset; Animals; Cerebellum; Disease Models, Animal; Enzyme Replacement Therapy; G(M2) Ganglioside; Gangliosidoses, GM2; Genetic Therapy; Glucosyltransferases; Glycoside Hydrolase Inhibitors; Gray Matter; Humans; Mutation; Retina; Spinal Cord; White Matter; beta-Hexosaminidase alpha Chain; beta-Hexosaminidase beta Chain
PubMed: 34450229
DOI: 10.1016/j.neulet.2021.136195 -
The Journal of Clinical Investigation May 2016GM2 gangliosidoses, including Tay-Sachs and Sandhoff diseases, are neurodegenerative lysosomal storage diseases that are caused by deficiency of β-hexosaminidase A,...
GM2 gangliosidoses, including Tay-Sachs and Sandhoff diseases, are neurodegenerative lysosomal storage diseases that are caused by deficiency of β-hexosaminidase A, which comprises an αβ heterodimer. There are no effective treatments for these diseases; however, various strategies aimed at restoring β-hexosaminidase A have been explored. Here, we produced a modified human hexosaminidase subunit β (HexB), which we have termed mod2B, composed of homodimeric β subunits that contain amino acid sequences from the α subunit that confer GM2 ganglioside-degrading activity and protease resistance. We also developed fluorescent probes that allow visualization of endocytosis of mod2B via mannose 6-phosphate receptors and delivery of mod2B to lysosomes in GM2 gangliosidosis models. In addition, we applied imaging mass spectrometry to monitor efficacy of this approach in Sandhoff disease model mice. Following i.c.v. administration, mod2B was widely distributed and reduced accumulation of GM2, asialo-GM2, and bis(monoacylglycero)phosphate in brain regions including the hypothalamus, hippocampus, and cerebellum. Moreover, mod2B administration markedly improved motor dysfunction and a prolonged lifespan in Sandhoff disease mice. Together, the results of our study indicate that mod2B has potential for intracerebrospinal fluid enzyme replacement therapy and should be further explored as a gene therapy for GM2 gangliosidoses.
Topics: Amino Acid Substitution; Animals; Brain; Disease Models, Animal; Gangliosidoses, GM2; Humans; Mice; Mice, Knockout; Mutation, Missense; Peptide Hydrolases; Proteolysis; beta-Hexosaminidase beta Chain
PubMed: 27018595
DOI: 10.1172/JCI85300 -
Neurobiology of Disease Feb 2020The favorable outcome of in vivo and ex vivo gene therapy approaches in several Lysosomal Storage Diseases suggests that these treatment strategies might equally benefit...
Novel bicistronic lentiviral vectors correct β-Hexosaminidase deficiency in neural and hematopoietic stem cells and progeny: implications for in vivo and ex vivo gene therapy of GM2 gangliosidosis.
The favorable outcome of in vivo and ex vivo gene therapy approaches in several Lysosomal Storage Diseases suggests that these treatment strategies might equally benefit GM2 gangliosidosis. Tay-Sachs and Sandhoff disease (the main forms of GM2 gangliosidosis) result from mutations in either the HEXA or HEXB genes encoding, respectively, the α- or β-subunits of the lysosomal β-Hexosaminidase enzyme. In physiological conditions, α- and β-subunits combine to generate β-Hexosaminidase A (HexA, αβ) and β-Hexosaminidase B (HexB, ββ). A major impairment to establishing in vivo or ex vivo gene therapy for GM2 gangliosidosis is the need to synthesize the α- and β-subunits at high levels and with the correct stoichiometric ratio, and to safely deliver the therapeutic products to all affected tissues/organs. Here, we report the generation and in vitro validation of novel bicistronic lentiviral vectors (LVs) encoding for both the murine and human codon optimized Hexa and Hexb genes. We show that these LVs drive the safe and coordinate expression of the α- and β-subunits, leading to supranormal levels of β-Hexosaminidase activity with prevalent formation of a functional HexA in SD murine neurons and glia, murine bone marrow-derived hematopoietic stem/progenitor cells (HSPCs), and human SD fibroblasts. The restoration/overexpression of β-Hexosaminidase leads to the reduction of intracellular GM2 ganglioside storage in transduced and in cross-corrected SD murine neural progeny, indicating that the transgenic enzyme is secreted and functional. Importantly, bicistronic LVs safely and efficiently transduce human neurons/glia and CD34+ HSPCs, which are target and effector cells, respectively, in prospective in vivo and ex vivo GT approaches. We anticipate that these bicistronic LVs may overcome the current requirement of two vectors co-delivering the α- or β-subunits genes. Careful assessment of the safety and therapeutic potential of these bicistronic LVs in the SD murine model will pave the way to the clinical development of LV-based gene therapy for GM2 gangliosidosis.
Topics: Animals; Gangliosidoses, GM2; Genetic Therapy; Genetic Vectors; Hematopoietic Stem Cells; Humans; Lentivirus; Mice; Neural Stem Cells; beta-Hexosaminidase alpha Chain; beta-Hexosaminidase beta Chain
PubMed: 31682993
DOI: 10.1016/j.nbd.2019.104667 -
Science Translational Medicine Sep 2016Lysosomal storage diseases (LSDs) often manifest with severe systemic and central nervous system (CNS) symptoms. The existing treatment options are limited and have no...
Lysosomal storage diseases (LSDs) often manifest with severe systemic and central nervous system (CNS) symptoms. The existing treatment options are limited and have no or only modest efficacy against neurological manifestations of disease. We demonstrate that recombinant human heat shock protein 70 (HSP70) improves the binding of several sphingolipid-degrading enzymes to their essential cofactor bis(monoacyl)glycerophosphate in vitro. HSP70 treatment reversed lysosomal pathology in primary fibroblasts from 14 patients with eight different LSDs. HSP70 penetrated effectively into murine tissues including the CNS and inhibited glycosphingolipid accumulation in murine models of Fabry disease (Gla(-/-)), Sandhoff disease (Hexb(-/-)), and Niemann-Pick disease type C (Npc1(-/-)) and attenuated a wide spectrum of disease-associated neurological symptoms in Hexb(-/-) and Npc1(-/-) mice. Oral administration of arimoclomol, a small-molecule coinducer of HSPs that is currently in clinical trials for Niemann-Pick disease type C (NPC), recapitulated the effects of recombinant human HSP70, suggesting that heat shock protein-based therapies merit clinical evaluation for treating LSDs.
Topics: Administration, Intravenous; Animals; Blood-Brain Barrier; Bone Morphogenetic Proteins; Disease Models, Animal; Disease Progression; Fabry Disease; Fibroblasts; Glycosphingolipids; Heat-Shock Proteins; Humans; Hydroxylamines; Injections, Intraperitoneal; Intracellular Signaling Peptides and Proteins; Lysosomes; Niemann-Pick C1 Protein; Niemann-Pick Disease, Type C; Proteins; Recombinant Proteins; Sphingolipidoses; Tissue Distribution
PubMed: 27605553
DOI: 10.1126/scitranslmed.aad9823 -
Scientific Reports Feb 2021This study evaluates the genetic spectrum of leukodystrophies and leukoencephalopathies in Iran. 152 children, aged from 1 day to 15 years, were genetically tested for...
This study evaluates the genetic spectrum of leukodystrophies and leukoencephalopathies in Iran. 152 children, aged from 1 day to 15 years, were genetically tested for leukodystrophies and leukoencephalopathies based on clinical and neuroradiological findings from 2016 to 2019. Patients with a suggestive specific leukodystrophy, e. g. metachromatic leukodystrophy, Canavan disease, Tay-Sachs disease were tested for mutations in single genes (108; 71%) while patients with less suggestive findings were evaluated by NGS. 108 of 152(71%) had MRI patterns and clinical findings suggestive of a known leukodystrophy. In total, 114(75%) affected individuals had (likely) pathogenic variants which included 38 novel variants. 35 different types of leukodystrophies and genetic leukoencephalopathies were identified. The more common identified disorders included metachromatic leukodystrophy (19 of 152; 13%), Canavan disease (12; 8%), Tay-Sachs disease (11; 7%), megalencephalic leukodystrophy with subcortical cysts (7; 5%), X-linked adrenoleukodystrophy (8; 5%), Pelizaeus-Merzbacher-like disease type 1 (8; 5%), Sandhoff disease (6; 4%), Krabbe disease (5; 3%), and vanishing white matter disease (4; 3%). Whole exome sequencing (WES) revealed 90% leukodystrophies and genetic leukoencephalopathies. The total diagnosis rate was 75%. This unique study presents a national genetic data of leukodystrophies; it may provide clues to the genetic pool of neighboring countries. Patients with clinical and neuroradiological evidence of a genetic leukoencephalopathy should undergo a genetic analysis to reach a definitive diagnosis. This will allow a diagnosis at earlier stages of the disease, reduce the burden of uncertainty and costs, and will provide the basis for genetic counseling and family planning.
Topics: Adolescent; Canavan Disease; Child; Child, Preschool; Female; Genetic Testing; Hereditary Central Nervous System Demyelinating Diseases; Humans; Infant; Infant, Newborn; Iran; Leukodystrophy, Globoid Cell; Leukodystrophy, Metachromatic; Leukoencephalopathies; Male; Mutation
PubMed: 33547378
DOI: 10.1038/s41598-021-82778-0 -
Glycobiology Mar 2022We recently discovered that human neutrophils express immunomodulatory glycoproteins carrying unusual and highly truncated paucimannosidic N-glycans...
We recently discovered that human neutrophils express immunomodulatory glycoproteins carrying unusual and highly truncated paucimannosidic N-glycans (Man1-3GlcNAc2Fuc0-1), but their biosynthesis remains elusive. Guided by the well-characterized truncation pathway in invertebrates and plants in which the N-acetyl-β-D-hexosaminidase (Hex) isoenzymes catalyze paucimannosidic protein (PMP) formation, we here set out to test if the homologous human Hex α and β subunits encoded by HEXA and HEXB drive a similar truncation pathway in human neutrophils. To this end, we performed quantitative glycomics and glycoproteomics of several CRISPR-Cas9-edited Hex-disrupted neutrophil-like HL-60 mutants (HEXA-KO and HEXB-KO) and matching unedited cell lines. Hex disruption was validated using next-generation sequencing, enzyme-linked immunosorbent assay (ELISA), quantitative proteomics and Hex activity assays. Excitingly, all Hex-disrupted mutants displayed significantly reduced levels of paucimannosylation, particularly Man2-3GlcNAc2Fuc1, relative to unedited HL-60 suggesting that both HEXA and HEXB contribute to PMP formation via a hitherto unexplored truncation pathway in neutrophils. Quantitative N-glycomics indeed demonstrated reduced utilization of a putative noncanonical truncation pathway in favor of the canonical elongation pathway in all Hex-disrupted mutants relative to unedited controls. Quantitative glycoproteomics recapitulated the truncation-to-elongation switch in all Hex-disrupted mutants and showed a greater switch for N-glycoproteins cotrafficking with Hex to the azurophilic granules of neutrophils such as myeloperoxidase. Finally, we supported the Hex-PMP relationship by documenting that primary neutrophils isolated from an early-onset Sandhoff disease patient (HEXB-/-) displayed dramatically reduced paucimannosylation relative to neutrophils from an age-matched unaffected donor. We conclude that both human Hex α and β mediate PMP formation via a putative noncanonical truncation pathway in neutrophils.
Topics: Hexosaminidase A; Hexosaminidase B; Hexosaminidases; Humans; Neutrophils; beta-N-Acetylhexosaminidases
PubMed: 34939086
DOI: 10.1093/glycob/cwab108 -
Journal of Lipid Research May 2015Bis(monoacylglycero)phosphate (BMP) is a negatively charged glycerophospholipid with an unusual sn-1;sn-1' structural configuration. BMP is primarily enriched in...
Bis(monoacylglycero)phosphate (BMP) is a negatively charged glycerophospholipid with an unusual sn-1;sn-1' structural configuration. BMP is primarily enriched in endosomal/lysosomal membranes. BMP is thought to play a role in glycosphingolipid degradation and cholesterol transport. Elevated BMP levels have been found in many lysosomal storage diseases (LSDs), suggesting an association with lysosomal storage material. The gangliosidoses are a group of neurodegenerative LSDs involving the accumulation of either GM1 or GM2 gangliosides resulting from inherited deficiencies in β-galactosidase or β-hexosaminidase, respectively. Little information is available on BMP levels in gangliosidosis brain tissue. Our results showed that the content of BMP in brain was significantly greater in humans and in animals (mice, cats, American black bears) with either GM1 or GM2 ganglioside storage diseases, than in brains of normal subjects. The storage of BMP and ganglioside GM2 in brain were reduced similarly following adeno-associated viral-mediated gene therapy in Sandhoff disease mice. We also found that C22:6, C18:0, and C18:1 were the predominant BMP fatty acid species in gangliosidosis brains. The results show that BMP accumulates as a secondary storage material in the brain of a broad range of mammals with gangliosidoses.
Topics: Animals; Brain; Cat Diseases; Cats; Female; Gangliosidosis, GM1; Humans; Lipid Metabolism; Lysophospholipids; Male; Mice, 129 Strain; Mice, Knockout; Monoglycerides; Sandhoff Disease; Ursidae
PubMed: 25795792
DOI: 10.1194/jlr.M057851 -
BMC Medical Genetics Dec 2019Neurological disorders are a common cause of morbidity and mortality within Pakistani populations. It is one of the most important challenges in healthcare, with...
BACKGROUND
Neurological disorders are a common cause of morbidity and mortality within Pakistani populations. It is one of the most important challenges in healthcare, with significant life-long socio-economic burden.
METHODS
We investigated the cause of disease in three Pakistani families in individuals with unexplained autosomal recessive neurological conditions, using both genome-wide SNP mapping and whole exome sequencing (WES) of affected individuals.
RESULTS
We identified a homozygous splice site variant (NM_000521:c.445 + 1G > T) in the hexosaminidase B (HEXB) gene confirming a diagnosis of Sandhoff disease (SD; type II GM2-gangliosidosis), an autosomal recessive lysosomal storage disorder caused by deficiency of hexosaminidases in a single family. In two further unrelated families, we identified a homozygous frameshift variant (NM_024298.3:c.758_778del; p.Glu253_Ala259del) in membrane-bound O-acyltransferase family member 7 (MBOAT7) as the likely cause of disease. MBOAT7 gene variants have recently been identified as a cause of intellectual disability (ID), seizures and autistic features.
CONCLUSIONS
We identified two metabolic disorders of lipid biosynthesis within three Pakistani families presenting with undiagnosed neurodevelopmental conditions. These findings enabled an accurate neurological disease diagnosis to be provided for these families, facilitating disease management and genetic counselling within this population. This study consolidates variation within MBOAT7 as a cause of neurodevelopmental disorder, broadens knowledge of the clinical outcomes associated with MBOAT7-related disorder, and confirms the likely presence of a regionally prevalent founder variant (c.758_778del; p.Glu253_Ala259del) in Pakistan.
Topics: Acyltransferases; Consanguinity; Electroencephalography; Female; Genes, Recessive; Homozygote; Humans; Infant; Magnetic Resonance Imaging; Male; Membrane Proteins; Mutation; Nervous System Diseases; Pakistan; Polymorphism, Single Nucleotide; Exome Sequencing; beta-Hexosaminidase beta Chain
PubMed: 31852446
DOI: 10.1186/s12881-019-0907-7 -
Molecular Therapy : the Journal of the... Oct 2020The GM2 gangliosidoses, Tay-Sachs disease (TSD) and Sandhoff disease (SD), are fatal lysosomal storage disorders caused by mutations in the HEXA and HEXB genes,...
The GM2 gangliosidoses, Tay-Sachs disease (TSD) and Sandhoff disease (SD), are fatal lysosomal storage disorders caused by mutations in the HEXA and HEXB genes, respectively. These mutations cause dysfunction of the lysosomal enzyme β-N-acetylhexosaminidase A (HexA) and accumulation of GM2 ganglioside (GM2) with ensuing neurodegeneration, and death by 5 years of age. Until recently, the most successful therapy was achieved by intracranial co-delivery of monocistronic adeno-associated viral (AAV) vectors encoding Hex alpha and beta-subunits in animal models of SD. The blood-brain barrier crossing properties of AAV9 enables systemic gene therapy; however, the requirement of co-delivery of two monocistronic AAV vectors to overexpress the heterodimeric HexA protein has prevented the use of this approach. To address this need, we developed multiple AAV constructs encoding simultaneously HEXA and HEXB using AAV9 and AAV-PHP.B and tested their therapeutic efficacy in 4- to 6-week-old SD mice after systemic administration. Survival and biochemical outcomes revealed superiority of the AAV vector design using a bidirectional CBA promoter with equivalent dose-dependent outcomes for both capsids. AAV-treated mice performed normally in tests of motor function, CNS GM2 ganglioside levels were significantly reduced, and survival increased by >4-fold with some animals surviving past 2 years of age.
Topics: Animals; Dependovirus; Disease Management; Disease Models, Animal; G(M2) Ganglioside; Gene Expression; Genetic Predisposition to Disease; Genetic Therapy; Genetic Vectors; Mice; Mutation; Sandhoff Disease; Tay-Sachs Disease; Transgenes; beta-N-Acetylhexosaminidases
PubMed: 32592687
DOI: 10.1016/j.ymthe.2020.06.021