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Structures of 6S-NBI-DGJ (the E-configuration about the C=N imine bound is represented) and NOEV.NOEV, N-octyl-4- epi-β-valienamine.
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Lysosomal β-galactosidase (β-Gal) deficiency causes a group of disorders that include neuronopathic GM(1) gangliosidosis and non-neuronopathic Morquio B disease. We have previously proposed the use of small molecule ligands of β-Gal as pharmacological chaperones (PCs) for the treatment of GM(1) gangliosidosis brain pathology. Although it is still u...
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Gangliosides are sialic acid containing glycosphingolipids, which are abundant in mammalian brain tissue. Several fatal human diseases are caused by defects in glycolipid metabolism. Defects in their degradation lead to an accumulation of metabolites upstream of the defective reactions, whereas defects in their biosynthesis lead to diverse problems...
We have identified and characterized three missense mutations in a patient with type 1G(M1) gangliosidosis, namely a substitution of G for A at nucleotide position 1044 (G1044 --> A; in exon 10) on one allele, which converts Asp(332) into asparagine, and both a mutation (C492 --> A in exon 4, leading to the amino acid change of Arg(148), Ser) ana a...
Haploinsufficiency of GRN causes frontotemporal dementia (FTD). The GRN locus produces progranulin (PGRN), which is cleaved to lysosomal granulin polypeptides. The function of lysosomal granulins and why their absence causes neurodegeneration are unclear. Here we discover that PGRN-deficient human cells and murine brains, as well as human frontal l...
Objectives
To study the clinical spectrum of inherited gray matter degenerative brain disorders (DBD) in children.
Methods
This cross-sectional study evaluated children up to 12 y of age, diagnosed with an inherited gray matter DBD in a tertiary care pediatric hospital between July 2019 and December 2020.
Results
A total of 314 children with prog...
GM1 gangliosidosis (GM1) is a fatal neurodegenerative lysosomal storage disease that occurs most commonly in young children, with no effective treatment available. Long-term follow-up of GM1 cats treated by bilateral thalamic and deep cerebellar nuclei (DCN) injection of adeno-associated virus (AAV)-mediated gene therapy has increased lifespan to 8...
Citations
... However, significant progress has been made in several directions. For example, enzyme replacement therapies (ERT) based on ICV injection or efficient enzyme delivery systems such as "Trojan horse" proteins and nanoparticles to address the blood brain barrier (BBB) problem have been explored [266][267][268][269][270]. Chaperon-mediated therapies (CMT) employing small molecule chaperons, such as Gal [271], iminosugars and valienamine derivatives [272][273][274][275][276][277][278][279][280], have been utilized to treat diseases caused by unstable or misfolded mutant enzymes [281][282][283][284]. Some of the therapies result in significant increases in βGal-ase activity [284] and decreases in the GM1 content of neonatal brains [279]. ...
GM1 is one of the major glycosphingolipids (GSLs) on the cell surface in the central nervous system (CNS). Its expression level, distribution pattern, and lipid composition are dependent upon cell and tissue type, developmental stage, and disease state, which suggests a potentially broad spectrum of functions of GM1 in various neurological and neuropathological processes. The major focus of this review is the roles that GM1 plays in the development and activities of brains, such as cell differentiation, neuritogenesis, neuroregeneration, signal transducing, memory, and cognition, as well as the molecular basis and mechanisms for these functions. Overall, GM1 is protective for the CNS. Additionally, this review has also examined the relationships between GM1 and neurological disorders, such as Alzheimer’s disease, Parkinson’s disease, GM1 gangliosidosis, Huntington’s disease, epilepsy and seizure, amyotrophic lateral sclerosis, depression, alcohol dependence, etc., and the functional roles and therapeutic applications of GM1 in these disorders. Finally, current obstacles that hinder more in-depth investigations and understanding of GM1 and the future directions in this field are discussed.
... Measurements of blood β-galactosidase activity were acquired from platelet-poor plasma and sera of wild-type C57BL/6 J mice at the pH 4.3 optimum of β-galactosidase activity and at physiological pH 7.4 in assays using the substrate 4-methylumbelliferyl-β-D-galactopyranoside and the galactosidase inhibitor N-(n-nonyl)deoxygalactonojirimycin (DGJ; Figure 1A) (22). The fluorescence of standardized 4-MU concentrations was used in calculating specific activity among plasma and sera samples ( Figure 1B). ...
Glycosidases are hydrolytic enzymes studied principally in the context of intracellular catabolism within the lysosome. Therefore, glycosidase activities are classically measured in experimentally acidified assay conditions reflecting their low pH optima. However, glycosidases are also present the bloodstream where they may retain sufficient activity to participate in functions including the regulation of glycoprotein half-lives, proteostasis, and disease pathogenesis. We have herein established at physiological pH 7.4 in blood plasma and sera the normal ranges of four major glycosidase activities essential for blood glycoprotein remodeling in healthy mice and humans. These activities included β-galactosidase, β-N-acetylglucosaminidase, α-mannosidase, and α-fucosidase. We have identified their origins to include the mammalian genes Glb1, HexB, Man2a1, and FucA1. In experimental sepsis, excursions of glycosidase activities occurred with differences in host responses to discrete bacterial pathogens. Among similar excursions in human sepsis, the elevation of β-galactosidase activity was a prognostic indicator of increased likelihood of patient death.
... Several N-alkylated DGJ derivatives were synthesized with enhanced specificity and affinity to galactosidases, such as N-butyl-DGJ (NB-DGJ (2); Figure 1) and N-nonyl-DGJ (NN-DGJ (3); Figure 1) [13,14]. The bicyclic sp 2 -iminosugar isothiourea derivative 6S-NBI-DGJ (5N,6S-[N'-butyliminomethylidene)-6-thio-1-deoxygalactonojirimycin (4); Figure 1) resulted in a sixfold β-Gal activity enhancement in the fibroblasts from patients homozygous for the juvenile GM1 mutation p.Arg201Cys [9,15,16] and showed promising properties as PC in a GM1 gangliosidosis mouse model [17]. A significant contribution was also provided by the Graz group, starting with the compound coined as DLHEX-DGJ (5), which showed significant activity enhancement (18-fold) with chaperone-sensitive p.Arg201Cys and p.Arg201His cell lines at 500 µM [18]. ...
... Several N-alkylated DGJ derivatives were synthesized with enhanced specificity and affinity to galactosidases, such as N-butyl-DGJ (NB-DGJ (2); Figure 1) and Nnonyl-DGJ (NN-DGJ (3); Figure 1) [13,14]. The bicyclic sp 2 -iminosugar isothiourea derivative 6S-NBI-DGJ (5N,6S-[N -butyliminomethylidene)-6-thio-1-deoxygalactonojirimycin (4); Figure 1) resulted in a sixfold β-Gal activity enhancement in the fibroblasts from patients homozygous for the juvenile GM1 mutation p.Arg201Cys [9,15,16] and showed promising properties as PC in a GM1 gangliosidosis mouse model [17]. A significant contribution was also provided by the Graz group, starting with the compound coined as DLHEX-DGJ (5), which showed significant activity enhancement (18-fold) with chaperonesensitive p.Arg201Cys and p.Arg201His cell lines at 500 µM [18]. ...
GM1 gangliosidosis is a rare lysosomal disease caused by the deficiency of the enzyme β-galactosidase (β-Gal; GLB1; E.C. 3.2.1.23), responsible for the hydrolysis of terminal β-galactosyl residues from GM1 ganglioside, glycoproteins, and glycosaminoglycans, such as keratan-sulfate. With the aim of identifying new pharmacological chaperones for GM1 gangliosidosis, the synthesis of five new trihydroxypiperidine iminosugars is reported in this work. The target compounds feature a pentyl alkyl chain in different positions of the piperidine ring and different absolute configurations of the alkyl chain at C-2 and the hydroxy group at C-3. The organometallic addition of a Grignard reagent onto a carbohydrate-derived nitrone in the presence or absence of a suitable Lewis Acid was exploited, providing structural diversity at C-2, followed by the ring-closure reductive amination step. An oxidation-reduction process allowed access to a different configuration at C-3. The N-pentyl trihydroxypiperidine iminosugar was also synthesized for the purpose of comparison. The biological evaluation of the newly synthesized compounds was performed on leucocyte extracts from healthy donors and identified two suitable β-Gal inhibitors, namely compounds 10 and 12. Among these, compound 12 showed chaperoning properties since it enhanced β-Gal activity by 40% when tested on GM1 patients bearing the p.Ile51Asn/p.Arg201His mutations.
... pFPhT-DGJ) 57,58 , b-galactosidase (e.g. 5 N,6S-[N'butyliminomethylidene]-1-deoxy-6-thiogalactonojirimycin; 6S-NBI-DGJ) 59,60 , and a-mannosidase (e.g. 6-[tert-butoxycarbonylamino]hexyl 5 N,6O-[oxamethylidene]mannonojirimycin-1-yl amine; BocNHex-N-OMJ) 61 in fibroblasts from patients suffering of Gaucher disease, Fabry disease, G M1 gangliosidosis and a-mannosidosis, respectively ( Figure 1). ...
The late-onset form of Tay-Sachs disease displays when the activity levels of human β-hexosaminidase A (HexA) fall below 10% of normal, due to mutations that destabilise the native folded form of the enzyme and impair its trafficking to the lysosome. Competitive inhibitors of HexA can rescue disease-causative mutant HexA, bearing potential as pharmacological chaperones, but often also inhibit the enzyme O-glucosaminidase (GlcNAcase; OGA), a serious drawback for translation into the clinic. We have designed sp²-iminosugar glycomimetics related to GalNAc that feature a neutral piperidine-derived thiourea or a basic piperidine-thiazolidine bicyclic core and behave as selective nanomolar competitive inhibitors of human Hex A at pH 7 with a ten-fold lower inhibitory potency at pH 5, a good indication for pharmacological chaperoning. They increased the levels of lysosomal HexA activity in Tay-Sachs patient fibroblasts having the G269S mutation, the highest prevalent in late-onset Tay-Sachs disease.
... Ortiz Mellet and Garcia Fernandez prepared novel iminosugar-isothiourea hybrids such as compound 5, which was found to effect a 6-fold enhancement of β-galactosidase activity in the R201C cell line favourably comparing to NOEV (4) but above concentrations of 100 μM and without toxic side effects up to 640 μM. [86] (Scheme 3, Table 1, Supporting Information) Chaperone 5 was found a considerably weaker inhibitor than NOEV (4), however was active with an impressively broader spectrum of G M1 -gangliosidosis mutants than the former including 24 of 88 screened human mutants, [87] in particular with I51T, a frequent mutation in Japan, as well as with G438T which both are known to be not responsive to NOEV (4). [71,72] Gratifyingly, this chaperone was also capable of crossing the blood-brain barrier. ...
... [71,72] Gratifyingly, this chaperone was also capable of crossing the blood-brain barrier. [87] An in-depth investigation into the differences between NOEV (4), and 6S-NBI-DGJ, 5 has recently been conducted revealing a possibly advantageous, more flexible binding mode of the latter, lacking interaction with Tyr133 in the catalytic site. [65] At Graz, in early 2012 it became clear that despite all efforts and many years of experience in the iminosugar area, none of the DGJ-analogs prepared and screened by us thus far were able to match the reported properties of NOEV (4) or would be a viable lead into that direction. ...
A short survey on selected β‐galactosidase inhibitors as potential pharmacological chaperones for GM1‐gangliosidosis and Morquio B associated mutants of human lysosomal β‐galactosidase is provided highlighting recent developments in this particular area of lysosomal storage disorders and orphan diseases. Neurodegenerative GM1‐gangliosidosis and bone‐damaging Morquio B disease are caused by transportation‐challenged or catalytically incompetent β‐d‐galactosidase mutants. For both lysosomal storage disorders there is currently no cure available. Great hope is focused on the concept of chaperone mediated therapy. Herein, we provide a short survey on selected iminosugar based β‐galactosidase inhibitors as potential pharmacological chaperones for GM1‐gangliosidosis and Morquio B associated mutants of human lysosomal β‐galactosidase highlighting recent developments in this particular area of lysosomal storage disorders and orphan diseases.
... Glb1 −/− or Glb1 tm1Adz /knock-out (Hahn et al., 1997) Glb1 −/− or Glb1 tm1Jmat /knock-out (Matsuda et al., 1997a,b) Glb1 −/− or β -gal −/− /knock-out (Przybilla et al., 2019) Glb1 −/− /knock-out (Eikelberg et al., 2020) Glb1 ( Inflammatory response and abnormal autophagy in the brains: CNS inflammation with activated microglia and abnormal autophagy Therapy • ERT (Chen et al., 2020) • SRT (Jeyakumar et al., 2003;Kasperzyk et al., 2004Kasperzyk et al., , 2005Elliot-Smith et al., 2008) • Ex vivo gene therapy (Sano et al., 2005) • In vivo gene therapy (Broekman et al., 2007;Baek et al., 2010;Weismann et al., 2015;Hinderer et al., 2020) • ERT (Matsuda et al., 2003;Suzuki, 2006Suzuki, , 2008Suzuki et al., 2007Suzuki et al., , 2012Takai et al., 2013) • In vivo gene therapy (Takaura et al., 2003) • ERT (Przybilla et al., 2021) n/a n/a Frontiers in Genetics | www.frontiersin.org that the authors attribute to increased neuronal cell size and reduced membrane resistance. ...
... Several pharmacological chaperones including galactose, N-octyl-4-epi-β-valienamine (NOEV)alkylated or fluorinated derivates of N-butyldeoxynojirimycin (NB-DNJ), and (5aR)-5a-C-Pentyl-4-epiisofagomine have been tested against numerous GLB1 mutant enzymes (Matsuda et al., 2003;Suzuki et al., 2007Suzuki et al., , 2012Suzuki, 2008Suzuki, , 2014Fantur et al., 2010;Takai et al., 2013;Parenti et al., 2015;Thonhofer et al., 2016;Front et al., 2017). ...
GM1 gangliosidosis is a progressive, neurosomatic, lysosomal storage disorder caused by mutations in the GLB1 gene encoding the enzyme β-galactosidase. Absent or reduced β-galactosidase activity leads to the accumulation of β-linked galactose-containing glycoconjugates including the glycosphingolipid (GSL) GM1-ganglioside in neuronal tissue. GM1-gangliosidosis is classified into three forms [Type I (infantile), Type II (late-infantile and juvenile), and Type III (adult)], based on the age of onset of clinical symptoms, although the disorder is really a continuum that correlates only partially with the levels of residual enzyme activity. Severe neurocognitive decline is a feature of Type I and II disease and is associated with premature mortality. Most of the disease-causing β-galactosidase mutations reported in the literature are clustered in exons 2, 6, 15, and 16 of the GLB1 gene. So far 261 pathogenic variants have been described, missense/nonsense mutations being the most prevalent. There are five mouse models of GM1-gangliosidosis reported in the literature generated using different targeting strategies of the Glb1 murine locus. Individual models differ in terms of age of onset of the clinical, biochemical, and pathological signs and symptoms, and overall lifespan. However, they do share the major abnormalities and neurological symptoms that are characteristic of the most severe forms of GM1-gangliosidosis. These mouse models have been used to study pathogenic mechanisms, to identify biomarkers, and to evaluate therapeutic strategies. Three GLB1 gene therapy trials are currently recruiting Type I and Type II patients (NCT04273269, NCT03952637, and NCT04713475) and Type II and Type III patients are being recruited for a trial utilizing the glucosylceramide synthase inhibitor, venglustat (NCT04221451).
... Oral administration of this compound in mice with GM1 gangliosidosis led to increased enzyme activity and reduced substrate levels. Other imino sugar derivatives have shown increased enzyme activity, facilitated localization of b-galactosidase to lysosomes in the fibroblasts of affected individuals [35], and improved enzyme activity in a mouse model of GM1 gangliosidosis [36]. ...
... 1,4 Also, chaperone effects are often mutationspecific, so evaluation of each chaperone in the context of each genotype is critical (Table 2). 55,[93][94][95]97,98,178,181 Most chaperones are imino-sugar, 4-epi-isofagomine, or valienamine derivatives. 91,92,97,101,182 The most heavily studied small molecules, NB-DNJ and NB-DGJ, are imino-sugar derivatives of glucose and galactose, respectively. ...
... In contrast to NOEV (discussed below), 6S-NBI-DGJ significantly enhanced β-gal activity in fibroblasts also expressing p.I51T or p.G438E mutations. 181 Of the 88 mutations examined by transient transfection of β-galdeficient COS7 cells, 24 responded positively to EET with 6S-NBI-DGJ. Further, studies in mice expressing human p.R201C mutant β-gal showed that 6S-NBI-DGJ is capable of crossing the BBB, enhancing β-gal activity throughout the CNS, and reducing GM1 ganglioside content. ...
... Further, studies in mice expressing human p.R201C mutant β-gal showed that 6S-NBI-DGJ is capable of crossing the BBB, enhancing β-gal activity throughout the CNS, and reducing GM1 ganglioside content. 178,181 Valienamine Derivatives as Chaperones N-Octyl-4-Epi-β -Valienamine (NOEV) 92 Modification of a glucocerebrosidase inhibitor through removal of its ceramide moiety and substitution with aliphatic chains led to the identification of N-octyl-4-epi-βvalienamine (NOEV) as a potent competitive inhibitor of β-gal and a candidate for EET. In human and mouse GM1 fibroblasts having p.R201H or p.R201C mutations, respectively, NOEV administration led to a marked increase in βgal activity and subsequent reduction in GM1 ganglioside content. ...
Allisandra K Rha,1 Anne S Maguire,1,2 Douglas R Martin1,2 1Scott-Ritchey Research Center, Auburn University, Auburn, AL, 36849, USA; 2Department of Anatomy, Physiology, and Pharmacology, Auburn University College of Veterinary Medicine, Auburn, AL, 36849, USACorrespondence: Douglas R MartinScott-Ritchey Research Center, Auburn University, 1265 H.C. Morgan Drive, Auburn, AL, 36849, USATel +1 334 844 5951Email martidr@auburn.eduAbstract: The lysosomal storage disorder, GM1 gangliosidosis (GM1), is a neurodegenerative condition resulting from deficiency of the enzyme β-galactosidase (β-gal). Mutation of the GLB1 gene, which codes for β-gal, prevents cleavage of the terminal β-1,4-linked galactose residue from GM1 ganglioside. Subsequent accumulation of GM1 ganglioside and other substrates in the lysosome impairs cell physiology and precipitates dysfunction of the nervous system. Beyond palliative and supportive care, no FDA-approved treatments exist for GM1 patients. Researchers are critically evaluating the efficacy of substrate reduction therapy, pharmacological chaperones, enzyme replacement therapy, stem cell transplantation, and gene therapy for GM1. A Phase I/II clinical trial for GM1 children is ongoing to evaluate the safety and efficacy of adeno-associated virus-mediated GLB1 delivery by intravenous injection, providing patients and families with hope for the future.Keywords: GM1 gangliosidosis, gene therapy, biomarkers, chaperone, GLB1, LSD
... Immunoblotting was performed with rabbit polyclonal anti-LC3 Ab (MBL, Nagoya, Japan), and polyclonal anti-tubulin Ab (Santa Cruz Biotech, Santa Cruz, CA). Immuno-positive signals were visualized by ECL plus reagent (GE Healthcare, Buckinghamshire, UK), and images were obtained using LAS-4000 image analyzer (Fujifilm, Tokyo, Japan) (Takai et al. 2013). ...
Xeroderma pigmentosum (XP) involves a defect in the initial step of nucleotide excision repair (NER) and consists of eight genetic complementation groups (groups A–G and a variant). XP group A (XPA) patients have a high incidence of UV-induced skin tumors, immature testicular development, and neurological symptoms. In an earlier study, we have shown that XP group A (Xpa) gene-knockout mice (Xpa−/− mice) were highly sensitive to UV-induced skin carcinogenesis with a defect in NER and were highly susceptibility to spontaneous tumorigenesis with impaired spermatogenesis. However, the pathology of impaired spermatogenesis in Xpa−/− mice is unknown. To unravel the underlying pathology, we made a concerted effort using the testis of 3-month-old Xpa−/− mice. We found many large vacuoles in the seminiferous tubules of 3-month old Xpa−/− mice, while there were no large vacuoles in that of Xpa+/+ mice. Immunohistochemistry of microtubule-associated protein 1 light chain 3 (LC3), an autophagosome marker, showed degenerating cells with intense signal of LC3 in the seminiferous tubules, and immunoblotting revealed induction of LC3-II in the 3-month-old Xpa−/− mice. The results of the present study suggest autophagy induction as the possible mechanism underlying the impaired spermatogenesis in Xpa−/− mice. Therefore, Xpa−/− mice could be a useful model for investigating aging and male infertility with low expression of XPA.
... Compounds 3-8 show several of the most active N-substituent-variants in the literature, which have interesting properties as experimental pharmacological chaperones. [22,[37][38][39][40][41][42][43][44]. ...
Glycosidase inhibitors have shown great potential as pharmacological chaperones for lysosomal storage diseases. In light of this, a series of new cyclopentanoid β-galactosidase inhibitors were prepared and their inhibitory and pharmacological chaperoning activities determined and compared with those of lipophilic analogs of the potent β-d-galactosidase inhibitor 4-epi-isofagomine. Structure-activity relationships were investigated by X-ray crystallography as well as by alterations in the cyclopentane moiety such as deoxygenation and replacement by fluorine of a "strategic" hydroxyl group. New compounds have revealed highly promising activities with a range of β-galactosidase-compromised human cell lines and may serve as leads towards new pharmacological chaperones for GM1-gangliosidosis and Morquio B disease.
