A novel GBE1 gene variant in a child with glycogen storage disease type IV.

Glycogen storage disease type IV is an autosomal recessive disorder of carbohydrates caused by deficiency of amylo-1-4-glycanoglycosyltransferase, which leads to accumulation of amylopectin-like polysaccharides in tissues including liver, heart and neuromuscular system. More than 40 different mutations in the glycogen branching enzyme gene (GBE1) have been described. In this study, we report a 2-year-old boy who presented with developmental delay and muscle weakness. He subsequently was diagnosed with glycogen storage disease type IV based on a liver biopsy histology and electron microscopy. Glycogen branching enzyme activity was in the low range. Genetic analysis demonstrated a novel heterozygous variant (c.760A>G; p.Thr254Ala) in exon 6 of the GBE1 gene, which is believed to be pathogenic. This variant was inherited from the patient's mother who was asymptomatic with normal glycogen branching enzyme activity. Whole-exome sequencing failed to reveal additional variations in the GBE1 gene.

Structural basis of glycogen branching enzyme deficiency and pharmacologic rescue by rational peptide design.

Glycogen branching enzyme 1 (GBE1) plays an essential role in glycogen biosynthesis by generating α-1,6-glucosidic branches from α-1,4-linked glucose chains, to increase solubility of the glycogen polymer. Mutations in the GBE1 gene lead to the heterogeneous early-onset glycogen storage disorder type IV (GSDIV) or the late-onset adult polyglucosan body disease (APBD). To better understand this essential enzyme, we crystallized human GBE1 in the apo form, and in complex with a tetra- or hepta-saccharide. The GBE1 structure reveals a conserved amylase core that houses the active centre for the branching reaction and harbours almost all GSDIV and APBD mutations. A non-catalytic binding cleft, proximal to the site of the common APBD mutation p.Y329S, was found to bind the tetra- and hepta-saccharides and may represent a higher-affinity site employed to anchor the complex glycogen substrate for the branching reaction. Expression of recombinant GBE1-p.Y329S resulted in drastically reduced protein yield and solubility compared with wild type, suggesting this disease allele causes protein misfolding and may be amenable to small molecule stabilization. To explore this, we generated a structural model of GBE1-p.Y329S and designed peptides ab initio to stabilize the mutation. As proof-of-principle, we evaluated treatment of one tetra-peptide, Leu-Thr-Lys-Glu, in APBD patient cells. We demonstrate intracellular transport of this peptide, its binding and stabilization of GBE1-p.Y329S, and 2-fold increased mutant enzymatic activity compared with untreated patient cells. Together, our data provide the rationale and starting point for the screening of small molecule chaperones, which could become novel therapies for this disease.

Adult polyglucosan body disease: a rare presentation with chronic liver disease and ground-glass hepatocellular inclusions.

Liver involvement in genetic and metabolic disorders may result in intrahepatic accumulation of specific precursors or byproducts, which have distinctive features on light microscopy. The "polyglucosan disorders" are diseases in which polyglucosan (abnormal glycogen with decreased branching) is formed and deposited in various tissues because of decreased or absent glycogen branching enzyme activity. These disorders include Lafora disease (myoclonus epilepsy) and type IV glycogen storage disease. Polyglucosan deposits in both conditions result in ground-glass hepatocellular inclusions resembling those seen in chronic hepatitis B virus infection. In the present report, we describe a case of the rare, adulthood form of glycogen branching enzyme deficiency, adult polyglucosan body disease (APBD), in which abnormal serum liver tests prompted a liver biopsy. The pathologic findings of periportal ground-glass hepatocellular inclusions, mild chronic portal inflammation, and periportal fibrosis are not well described in APBD, but resemble the chronic changes that have been reported in Lafora disease. The differential diagnosis of ground-glass hepatocytes and the genetic basis of APBD are discussed.

Branching enzyme deficiency/glycogenosis storage disease type IV presenting as a severe congenital hypotonia: muscle biopsy and autopsy findings, biochemical and molecular genetic studies.

The fatal infantile neuromuscular presentation of branching enzyme deficiency (glycogen storage disease type IV) due to mutations in the gene encoding the glycogen branching enzyme, is a rare but probably underdiagnosed cause of congenital hypotonia. We report an infant girl with severe generalized hypotonia, born at 33 weeks gestation who required ventilatory assistance since birth. She had bilateral ptosis, mild knee and foot contractures and echocardiographic evidence of cardiomyopathy. A muscle biopsy at 1 month of age showed typical polyglucosan storage. The autopsy at 3.5 months of age showed frontal cortex polymicrogyria and polyglucosan bodies in neurons of basal ganglia, thalamus, substantia innominata, brain stem, and myenteric plexus, as well as liver involvement. Glycogen branching enzyme activity in muscle was virtually undetectable. Sequencing of the GBE1 gene revealed a homozygous 28 base pair deletion and a single base insertion at the same site in exon 5. This case confirms previous observations that GBE deficiency ought to be included in the differential diagnosis of congenital hypotonia and that the phenotype correlates with the 'molecular severity' of the mutation.

Neuropathological study of skeletal muscle, heart, liver, and brain in a neonatal form of glycogen storage disease type IV associated with a new mutation in GBE1 gene.

Glycogen storage disease type IV (GSD IV, or Andersen disease) is an autosomal recessive disorder due to the deficiency of 1,4-alpha-glucan branching enzyme (or glycogen branching enzyme, GBE1), resulting in an accumulation of amylopectin-like polysaccharide in muscle, liver, heart and central and peripheral nervous system. Typically, the presentation is in childhood with liver involvement up to cirrhosis. The neuromuscular form varies in onset (congenital, perinatal, juvenile and adult) and in severity. Congenital cases are rare, and fewer than 20 cases have been described and genetically determined so far. This form is characterized by polyhydramnios, neonatal hypotonia, and neuronal involvement; hepatopathy is uncommon, and the babies usually die between 4 weeks and 4 months of age. We report the case of an infant who presented severe hypotonia, dilatative cardiomyopathy, mild hepatopathy, and brain lateral ventricle haemorrhage, features consistent with the congenital form of GSD IV. He died at one month of life of cardiorespiratory failure. Muscle biopsy and heart and liver autoptic specimens showed many vacuoles filled with PAS-positive diastase-resistant materials. Electron-microscopic analysis showed mainly polyglucosan accumulations in all the tissues examined. Postmortem examination showed the presence of vacuolated neurons containing this abnormal polysaccharide. GBE1 biochemical activity was virtually absent in muscle and fibroblasts, and totally lacking in liver and heart as well as glycogen synthase activity. GBE1 gene sequence analysis revealed a novel homozygous nonsense mutation, p.E152X, in exon 4, correlating with the lack of enzyme activity and with the severe neonatal involvement. Our findings contribute to increasing the spectrum of mutation associated with congenital GSD IV.

Multisystem involvement in a patient due to accumulation of amylopectin-like material with diminished branching enzyme activity.

We report a 13-year-old boy with multisystem involvement secondary to accumulation of amylopectin-like material. He was born to consanguineous parents at full term without any complications and his maternal perinatal history was uneventful. His parents were cousins. He had normal growth and development except for his weight. His sister died from an unexplained cardiomyopathy at the age of 8 years. Our patient's initial symptom was severe heart failure. Since he also had a complaint of muscle weakness, electromyography was performed which showed muscle involvement. The diagnosis was suggested by tissue biopsy of skeletal muscle showing intracellular, basophilic, diastase-resistant, periodic acid-Schiff-positive inclusion bodies and was confirmed by the presence of a completed branching enzyme deficiency. Similar intracytoplasmic inclusion-like bodies were also found in liver biopsy, but very few in number compared with the skeletal muscle. The patient died from an intercurrent infection. Postmortem endomyocardial biopsy revealed the same intracytoplasmic inclusions as described above affecting almost all myocardial cells. Ultrastructural examination of liver biopsy was nondiagnostic; however, myocardium showed prominent, large, intracytoplasmic deposits. Glycogen branching enzyme gene sequence was normal, and thus classical branching enzyme deficiency was excluded. Our patient represents the first molecular study performed on a patient in whom there was multiple system involvement secondary to accumulation of amylopectin-like material. We suggest that this is an as yet undefined and different phenotype of glycogen storage disease associated with multisystemic involvement.

Fetal type IV glycogen storage disease: clinical, enzymatic, and genetic data of a pure muscular form with variable and early antenatal manifestations in the same family.

We report on a family of three consecutive fetuses affected by type IV glycogen storage disease (GSD IV). In all cases, cervical cystic hygroma was observed on the 12-week-ultrasound examination. During the second trimester, fetal hydrops developed in the first pregnancy whereas fetal akinesia appeared in the second pregnancy. The diagnosis was suggested by microscopic examination of fetal tissues showing characteristic inclusions exclusively in striated fibers, then confirmed by enzymatic studies on frozen muscle. Antenatal diagnosis was performed on the third and fourth pregnancies: cervical cystic hygroma and low glycogen branching enzyme (GBE) activity on chorionic villi sample (CVS) were detected in the third pregnancy whereas ultrasound findings were normal and GBE activity within normal range on CVS in the fourth pregnancy. Molecular analysis showed that the mother was heterozygous for a c.1471G > C mutation in exon 12, leading to the replacement of an alanine by a tyrosine at codon 491 (p.A491T); the father was heterozygous for a c.895G > T mutation in exon 7, leading to the creation of a stop codon at position 299 (p.G299X). GSD IV has to be considered in a context of cervical cystic hygroma with normal karyotype, particularly when second trimester hydrops or akinesia develop. Enzymatic analysis of GBE must be performed on CVS or amniotic cells to confirm the diagnosis. Characteristic intracellular inclusions are specific to the disease and should be recognized, even in macerated tissues after fetal death. Genetic analysis of the GBE gene may help to shed some light on the puzzling diversity of GSD IV phenotypes.

Adult polyglucosan body disease: a case report of a manifesting heterozygote.

A 62-year-old man developed progressive gait instability, bladder dysfunction, proximal weakness, distal sensory loss, and mild cognitive impairment over 6 years. Neurologic examination revealed upper and lower motor neuron dysfunction in the lower extremities, with distal sensory loss. Electrodiagnostic studies, magnetic resonance imaging of the brain, and sural nerve biopsy were consistent with adult polyglucosan body disease. Biochemical and genetic analyses demonstrated reduced glycogen brancher enzyme levels associated with a heterozygous point mutation (Tyr329Ser or Y329S) in the glycogen brancher enzyme gene on chromosome 3. Mutational heterozygosity in the glycogen brancher enzyme gene has not been previously reported as a cause for this rare disease. A review of the clinical presentation, pathogenesis, etiology, and diagnosis of this disease is presented.

Novel missense mutations in the glycogen-branching enzyme gene in adult polyglucosan body disease.

We describe the first non-Ashkenazi patient with adult polyglucosan body disease and decreased glycogen-branching enzyme (GBE) activity in leukocytes. Gene analysis revealed compound heterozygosity for two novel missense mutations Arg515His and Arg524Gln in the GBE gene. Both missense mutations are predicted to impair GBE activity. This is the first identification of GBE mutations underlying adult polyglucosan body disease in a non-Ashkenazi family, and confirms that adult glycogen storage disease type IV can manifest clinically as adult polyglucosan body disease.

Concomitant branching enzyme and phosphorylase deficiencies. An unusual glycogenosis with extensive neuronal polyglucosan storage.

A baby girl was born hypotonic and was respirator-dependent until death at 43 days of age. A muscle biopsy revealed PAS-positive, diastase-resistant sarcoplasmic inclusions with a vaguely fibrillar structure by electron microscopy. Biochemical studies at autopsy disclosed complete absence of branching enzyme in skeletal muscle and heart, and a deficiency of phosphorylase activity in skeletal muscle with a modest reduction in myocardium. Storage material was present in glia and perikarya of neurons, increasing in amount in the rostrocaudal direction, involving most severely the motor neurons in the brain stem and spinal cord, dorsal root ganglia and myenteric plexi. Inclusions were also present in most organs, especially liver and skeletal muscle. Ultrastructurally, the inclusions ranged from granular aggregates of membrane-bound material concentrated in the region of Golgi apparatus to large filamentous bodies similar to polyglucosan bodies. This baby differs from other patients with infantile glycogenosis IV by the severity and onset of symptoms at birth, involvement of neuronal perikarya and widespread extraneural deposits. The combined deficiencies of branching enzyme and phosphorylase may have accounted for the unique clinical and neuropathological findings.

Hereditary branching enzyme dysfunction in adult polyglucosan body disease: a possible metabolic cause in two patients.

We describe 2 unrelated patients with adult polyglucosan body disease (APBD) diagnosed by sural nerve biopsy. Both patients were offspring of consanguineous marriages. They presented clinically with late onset pyramidal tetraparesis, micturition difficulties, peripheral neuropathy, and mild cognitive impairment. Magnetic resonance imaging of the brain revealed extensive white matter abnormalities in both. In search of a possible metabolic defect, we evaluated glycogen metabolism in these patients and their clinically unaffected children. Branching enzyme activity in the patients' polymorphonuclear leukocytes was about 15% of control values, whereas their children displayed values of 50 to 60%, suggesting a possible autosomal recessive mode of transmission. This is the first report of an inherited metabolic defect in patients with adult polyglucosan body disease. We suggest that branching enzyme dysfunction may be implicated in the pathogenesis of some patients with adult polyglucosan body disease.

A new variant of type IV glycogenosis: deficiency of branching enzyme activity without apparent progressive liver disease.

Type IV glycogenosis is due to branching enzyme deficiency and is usually manifested clinically by progressive liver disease with cirrhosis and hepatic failure between the second and fourth years of life. We describe a 5-year-old boy who, following an acute febrile illness at 2 years of age, was first noted to have hepatomegaly with mildly elevated serum transaminase levels. Liver biopsy revealed hepatic fibrosis with periodic-acid Schiff-positive, diastase-resistant inclusions in hepatocytes and fibrillar inclusions characteristic of amylopectin by electron microscopy. Enzymatic assay revealed deficient hepatic branching enzyme activity with normal activity of glucose-6-phosphatase, debranching enzyme and phosphorylase activities. During the succeeding 3 years, he grew and developed normally with apparent resolution of any clinical evidence of liver disease and only intermittent elevation in serum transaminase levels associated with fever and prolonged fasting. Repeat liver biopsy at 4 years of age showed persistence of scattered hepatocellular periodic-acid Schiff-positive, diastase-resistant inclusions, but no progression of hepatic fibrosis in spite of persistent deficiency of hepatic branching enzyme activity. Skeletal muscle and skin fibroblasts from the patient also showed deficient enzyme activity. Skin fibroblasts from both parents exhibited half the normal control activity, suggesting a heterozygote state. This is the first documented patient with deficiency of branching enzyme but without evidence of progressive hepatic disease. This patient, coupled with reports of other patients with late onset hepatic or muscle disease with branching enzyme deficiency, suggests that the defect resulting in Type IV glycogen storage disease is more heterogenous and possibly more common than previously suspected.