Lysosomal Glycogen Accumulation Research Articles

The value of muscle biopsies in Pompe disease: identifying lipofuscin inclusions in juvenile- and adult-onset patients

BackgroundPompe disease, an inherited deficiency of lysosomal acid alpha-glucosidase (GAA), is a metabolic myopathy with heterogeneous clinical presentations. Late-onset Pompe disease (LOPD) is a debilitating progressive muscle disorder that can occur anytime from early childhood to late adulthood. Enzyme replacement therapy (ERT) with recombinant human GAA is currently available for Pompe patients. Although ERT shows some benefits, the reversal of skeletal muscle pathology - lysosomal glycogen accumulation and autophagic buildup - remains a challenge. In this study, we examined the clinical status and muscle pathology of 22 LOPD patients and one atypical infantile patient on ERT to understand the reasons for muscle resistance to ERT.ResultsThe patients were divided into three groups for analysis, based on the age of onset and diagnosis: adult-onset patients, juvenile-onset patients, and those identified through newborn screening (NBS). The areas of autophagic buildup found in patients’ biopsies of all three groups, contained large autofluorescent inclusions which we show are made of lipofuscin, an indigestible intralysosomal material typically associated with ageing. These inclusions, analysed by staining, spectral analysis, time-resolved Fluorescence Lifetime Imaging (FLIM), and Second Harmonic Generation (SHG) imaging, were the major pathology remaining in many fibers after ERT. The best outcome of ERT both clinically and morphologically was observed in the NBS patients.ConclusionsThe muscle biopsy, in spite of its shortcomings, allowed us to recognize an underreported, ERT-resistant pathology in LOPD; numerous lysosomes and autolysosomes loaded with lipofuscin appear to be a hallmark of LOPD skeletal muscle. Lipofuscin accumulation - a result of inefficient lysosomal degradation - may in turn exacerbate both lysosomal and autophagic abnormalities.

Antisense Oligonucleotide-mediated Suppression of Muscle Glycogen Synthase 1 Synthesis as an Approach for Substrate Reduction Therapy of Pompe Disease.

Pompe disease is an autosomal recessive disorder caused by a deficiency of acid α-glucosidase (GAA; EC 3.2.1.20) and the resultant progressive lysosomal accumulation of glycogen in skeletal and cardiac muscles. Enzyme replacement therapy using recombinant human GAA (rhGAA) has proven beneficial in addressing several aspects of the disease such as cardiomyopathy and aberrant motor function. However, residual muscle weakness, hearing loss, and the risks of arrhythmias and osteopenia persist despite enzyme therapy. Here, we evaluated the relative merits of substrate reduction therapy (by inhibiting glycogen synthesis) as a potential adjuvant strategy. A phosphorodiamidate morpholino oligonucleotide (PMO) designed to invoke exon skipping and premature stop codon usage in the transcript for muscle specific glycogen synthase (Gys1) was identified and conjugated to a cell penetrating peptide (GS-PPMO) to facilitate PMO delivery to muscle. GS-PPMO systemic administration to Pompe mice led to a dose-dependent decrease in glycogen synthase transcripts in the quadriceps, and the diaphragm but not the liver. An mRNA response in the heart was seen only at the higher dose tested. Associated with these decreases in transcript levels were correspondingly lower tissue levels of muscle specific glycogen synthase and activity. Importantly, these reductions resulted in significant decreases in the aberrant accumulation of lysosomal glycogen in the quadriceps, diaphragm, and heart of Pompe mice. Treatment was without any overt toxicity, supporting the notion that substrate reduction by GS-PPMO-mediated inhibition of muscle specific glycogen synthase represents a viable therapeutic strategy for Pompe disease after further development.

Doença de Pompe Juvenil: Estudo Retrospetivo de Casuística Clínica

Pompe disease or glycogen storage disease type II is an autosomal recessive disorder due to acid maltase deficiency. It is a rare disease with a prevalence of 1/40.000 in the dutch and african-american populations and 1/46000 in the australian population. There are three forms of clinical presentation (infantile-onset, childhood-onset and adult-onset), although the disease presents as a continuum of clinical phenotypes. Enzyme replacement therapy is available in Portugal since 2006. The clinical files of four patients (two sisters) were analyzed retrospectively. In all, disease presented in the second year of life and the time to diagnosis ranged from two to eleven years. At diagnosis, all presented myopathic features with a delay in motor skills achievement and two had myocardium hypertrophy. Clinical suspicion arose as the result of respiratory failure during infection in two patients. Plasma creatine kinase and aminotransferases levels were increased in all. All patients progressed to pulmonary restrictive syndrome with chronic respiratory failure. The diagnosis was based on reduced activity of acid maltase in fibroblasts: 0 to 1.5% of the lower normal value. Muscle biopsy, performed in three patients, showed lysosomal glycogen accumulation. A c.1064T > C mutation in exon 6 of GAA (glucosidase-alpha-acid) gene was found in all patients, in homozygosity in one. In the sisters, it was associated to c.1666A > G and c.2065G > A mutations in exons 12 and 15, respectively and in the youngest patient, to c.380G > T mutation in exon 2. All patients started enzyme replacement therapy as soon as it became available, with good tolerance. The youngest patient died. The surviving patients maintain ventilatory support measures and physiotherapy. The oldest patient is wheelchair ridden and her sister keeps independent walking. The youngest uses a walking aid. Our cases are clinically included in the juvenile form of Pompe Disease. Pompe disease should be suspected in progressive myopathies at any age, especially those involving limb-girdle and respiratory muscles and in small infants with cardiomyopathy. High creatine kinase is a sensitive, although nonspecific, marker. Given the great variability of the genetic findings, demonstration of reduced activity of acid maltase (in leukocytes or other tissues) remains the diagnosis cornerstone of this rare disorder.

P.17.8 The co-formulation of pharmacological chaperone AT2220 with recombinant human acid alpha-glucosidase improves enzyme uptake and glycogen reduction in a mouse model of Pompe disease

Pompe disease is an inherited lysosomal storage disease that results from deficiency in acid alpha-glucosidase (GAA) activity, and is characterized by progressive accumulation of lysosomal glycogen in heart and skeletal muscles. Enzyme replacement therapy using recombinant human GAA (rhGAA) is the only approved treatment available for Pompe disease. While rhGAA provides some clinical benefits, the infused enzyme tends to be unstable at neutral pH/body temperature, shows insufficient uptake in key tissues, and can elicit immune responses that affect tolerability and efficacy. We have shown previously that oral pre-administration of the pharmacological chaperone AT2220 (1-deoxynojirimycin HCl, duvoglustat HCl) improves the pharmacological properties of rhGAA via binding and stabilization, leading to increased enzyme uptake and glycogen reduction in GAA knock-out (KO) mice. In this study we tested the effects of intravenous (IV) and subcutaneous (SQ) administration of co-formulated AT2220 and rhGAA (AT2220 + rhGAA) as an alternative to giving AT2220 orally prior to rhGAA IV. In rats, IV or SQ administration of co-formulated AT2220 + rhGAA increased the circulating half-life of rhGAA up to twofold, though the maximal rhGAA plasma levels achieved via the SQ route were significantly lower than those seen following IV administration. In GAA KO mice, four IV administrations of co-formulated AT2220 + rhGAA resulted in up to 2.5-fold greater enzyme uptake and glycogen reduction in disease-relevant tissues compared to rhGAA alone; histological staining confirmed reduced skeletal muscle glycogen. Interestingly, four SQ administrations of co-formulated AT2220 + rhGAA increased rhGAA uptake in GAA KO mice, which was not significantly different from those seen following four IV administrations of rhGAA alone. Collectively, these data highlight the potential effects of co-formulated AT2220 + rhGAA using IV or SQ administration, thus warranting further preclinical investigation.

P.17.9 AAV9 vector encoding hGAA improves synaptic pathology of the neuromuscular junction in Pompe disease

Pompe disease is caused by the deficiency of the lysosomal enzyme acid alpha-glucosidase (GAA) and results in a progressive accumulation of lysosomal glycogen. This impacts cellular architecture and function and leads to progressive respiratory insufficiency and motoneuron dysfunction. We have developed an adeno-associated virus (AAV) vector strategy to express long-term therapeutic levels of human GAA in cardiac, skeletal muscle, and the CNS resulting in improved cardiorespiratory function. However, an incomplete restoration of skeletal muscle function or activation still remains. We tested the hypothesis that an AAV9 vector encoding hGAA would result in retrograde transport and restore neuromuscular junction integrity and reverse axonal pathology in Pompe disease. 20 month old Pompe mice (Gaa−/−) were randomized to the following groups: untreated (Gaa−/−), AAV9-CMV-hGAA, or AAV9-DES-hGAA. AAV9 treated animals received a single injection of 1 × 1011vg in the right tibialis anterior (TA) muscle. One month post injection, the TA muscle and lumbar spinal cord were analyzed for vector genome copy number and GAA activity. Significant levels of vector genomes were detected in the TA (AAV9-DES-hGAA 1.5 × 105 ± 3.1 × 104 vg/ug DNA; AAV9-CMV-hGAA 8.4 × 104 ± 1.7 × 104 vg/ug DNA) and lumbar spinal cord (AAV9-DES-hGAA 1.5 × 103 ± 1.7 × 102 vg/ug DNA; AAV9-CMV-hGAA 2.5 × 103 ± 1.3 × 103 vg/ug DNA) suggesting efficient transduction of skeletal muscle and retrograde transport of AAV9. Activity of GAA in TA lysates was 2396% and 1770% above wild-type in AAV9-DES-hGAA and AAV9-CMV-hGAA animals, respectively ( p

P.17.4 Magnetic Resonance Imaging (MRI) to evaluate the effect of enzyme replacement therapy in Late Onset Pompe Disease (LOPD)

Pompe disease is a rare, progressive and often fatal neuromuscular disorder resulting from mutations in the gene for acid alpha-glucosidase (GAA), an enzyme necessary to degrade glycogen. The deficiency or dysfunction of the GAA enzyme leads to lysosomal glycogen accumulation in multiple tissues and results in cardiac and skeletal muscle dysfunction. Glycogen content may vary between muscle groups and in regions of the same muscle. Most importantly, glycogen content may change during the course of the disease. Enzyme replacement therapy (ERT) is the only FDA approved treatment for Pompe disease. However, translation from preclinical to clinical studies is challenged by the lack of reliable, noninvasive outcome measures suitable for repeated evaluations. Effect of treatment is commonly measured by muscle glycogen clearance using biochemical assays, which rely on muscle biopsy. In this study, we used Magnetic Resonance Imaging (MRI) of the thigh and lower leg muscles to evaluate the effect of 12 and 24 weeks of ERT in individuals affected by Late Onset Pompe Disease (LOPD) (n = 4; age 35–50). Maximum cross-sectional area (CSAmax) – an index of muscle mass – increased in average by 1.16- and 1.09-fold at 12 weeks, and by 1.13- and 1.09-fold at 24 weeks in the lower and upper legs respectively. In addition, we studied the MR proton traverse relaxation time (T2) – an index of muscle damage and edema – by applying a mono exponential decay equation to four echo times (TE). At 24 weeks, T2 values reduced by 1.20- and 1.10-fold in the lower and upper legs, respectively. These data demonstrate an increase in CSAmax associated accompanied by a reduction of T2 value, which may suggest a reduction of glycogen accumulation and an increase of contractile myofibers. In support of this hypothesis, biochemical assay from muscle biopsy specimens showed a reduction of glycogen concentration by 2.21-fold at week 24 compare to baseline.

Skeletal muscle pathology of infantile Pompe disease during long-term enzyme replacement therapy

BackgroundPompe disease is an autosomal recessive metabolic neuromuscular disorder caused by a deficiency of the lysosomal enzyme acid alpha-glucosidase (GAA). It has long been believed that the underlying pathology leading to tissue damage is caused by the enlargement and rupture of glycogen-filled lysosomes. Recent studies have also implicated autophagy, an intracellular lysosome-dependent degradation system, in the disease pathogenesis. In this study, we characterize the long-term impact of enzyme replacement therapy (ERT) with recombinant human GAA (rhGAA) on lysosomal glycogen accumulation and autophagy in some of the oldest survivors with classic infantile Pompe disease (IPD).MethodsMuscle biopsies from 8 [4 female, 4 male; 6 cross-reactive immunologic material (CRIM)-positive, 2 CRIM-negative] patients with a confirmed diagnosis of classic IPD were examined using standard histopathological approaches. In addition, muscle biopsies were evaluated by immunostaining for lysosomal marker (lysosomal-associated membrane protein-2; LAMP2), autophagosomal marker (microtubule-associated protein 1 light chain 3; LC3), and acid and alkaline ATPases. All patients received rhGAA by infusion at cumulative biweekly doses of 20–40 mg/kg.ResultsMedian age at diagnosis of classic IPD was 3.4 months (range: 0 to 6.5 months; n = 8). At the time of muscle biopsy, the patients’ ages ranged from 1 to 103 months and ERT duration ranged from 0 (i.e., baseline, pre-ERT) to 96 months. The response to therapy varied considerably among the patients: some patients demonstrated motor gains while others experienced deterioration of motor function, either with or without a period of initial clinical benefit. Skeletal muscle pathology included fiber destruction, lysosomal vacuolation, and autophagic abnormalities (i.e., buildup), particularly in fibers with minimal lysosomal enlargement. Overall, the pathology reflected clinical status.ConclusionsThis is the first study to investigate the impact of rhGAA ERT on lysosomal glycogen accumulation and autophagic buildup in patients with classic IPD beyond 18 months of treatment. Our findings indicate that ERT does not fully halt or reverse the underlying skeletal muscle pathology in IPD. The best outcomes were observed in the two patients who began therapy early, namely at 0.5 and 1.1 months of age.

Glucosa tetrasacárido como biomarcador diagnóstico de la enfermedad de Pompe: estudio en 35 pacientes

Background and objectivesPompe disease is a disorder originating from an acid alpha-glycosidase (AAG) enzyme deficiency. This disease produces an accumulation of lysosomal glycogen in different tissues, whereby the skeletal and heart muscles are especially involved. The established diagnosis is achieved through the identification of the AAG deficiency. There are also other secondary diagnostic biomarkers, such as tetra-saccharide glucose (Glc4), which shows high levels in the urine of these patients. In this study it is highlighted the usefulness of Glc4 as a diagnostic biomarker for Pompe disease in its different forms of presentation, using a high-performance liquid chromatography with ultraviolet detection (HPLC/UV) adapted to the study. Patients and methodsA total of 75 individuals have been analyzed: 40 healthy controls and 35 patients diagnosed with Pompe disease. Twenty-four hour samples of urine were collected from all of the patients and their Glc4 levels were determined by means of HPLC/UV. ResultsThe evaluation of the urinary Glc4 shows a high discrimination ability between healthy/sick individuals. In addition, the results obtained have allowed to establish the most appropriate level of decision or cut-off point for the identification of sick people. ConclusionsGlc4 urinary levels are found to be high in patients suffering from Pompe disease and even though increased levels are also found in other conditions, the existence of a AAG deficiency together with a compatible clinical symptoms, prove very helpful for a correct diagnosis of this serious disease.

Stbd1 is highly elevated in skeletal muscle of Pompe disease mice but suppression of its expression does not affect lysosomal glycogen accumulation

Previous studies strongly suggest that starch binding domain containing protein 1 (Stbd1) plays an important role in intracellular glycogen trafficking into lysosomes. We report here that Stbd1 expression is markedly increased in skeletal muscles but not in heart and liver of GAA-KO mice. An AAV2/9 vector expressing a Stbd1-specific shRNA effectively suppressed Stbd1 expression but did not alter lysosomal glycogen accumulation in the affected tissues of GAA-KO mice. Our results indicate that inhibition of Stbd1 does not appear to be an effective therapeutic approach for Pompe disease.

Evaluation of muscle biopsy in late-onset GSDII patients before and after enzyme replacement therapy (ERT)

Glycogen storage disease, glycogenosis type II (GSDII), or Pompe disease (OMIM 23230), is an autosomal recessive lysosomal storage disorder that results from a deficiency in the acid alpha glucosidase (GAA) enzyme. The disease is characterized by progressive accumulation of lysosomal glycogen in various tissues, primarily in cardiac and skeletal muscles. The histopathological hallmarks in the muscle are fiber vacuolization and autophagy. GSDII is clinically classified as a severe infantile, or an attenuated, later-onset form affecting children and adults. Recombinant human GAA (rhGAA) is the only approved enzyme replacement therapy (ERT) available for the treatment of Pompe disease, and it is effective in infantile patients, whereas in adults, improvements are more variable among different patients. Our project aims to assess the effects of ERT in 19 late-onset patients using both biochemical and morphological (histological, histochemical, and immunohistochemical) evaluations of skeletal muscle biopsies before and after 6 months to 1 year on ERT.

Pompe disease: the role of MRI

Glycogen storage disease type II (Pompe disease) is a rare, progressive muscle disorder with a wide range of phenotypic presentations. It is caused by an inherited deficiency of acid α-glucosidase (GAA), which leads to lysosomal glycogen accumulation in various tissues, most notably cardiac, skeletal, and smooth muscle. The gradual pathologic storage of GAA in muscle cells causes irreversible muscle damage, with different signs and symptoms, including respiratory insufficiency and muscle weakness. In Pompe disease, defining severity grades is essential for prognosis and for monitoring responses to enzyme replacement therapy (available since 2006). The purpose of this analysis was to describe the MR-imaging findings of patients with Pompe disease being treated in our institution between 2010 and 2012 (n=10).

Dysregulation of Multiple Facets of Glycogen Metabolism in a Murine Model of Pompe Disease

Pompe disease, also known as glycogen storage disease (GSD) type II, is caused by deficiency of lysosomal acid α-glucosidase (GAA). The resulting glycogen accumulation causes a spectrum of disease severity ranging from a rapidly progressive course that is typically fatal by 1 to 2 years of age to a slower progressive course that causes significant morbidity and early mortality in children and adults. The aim of this study is to better understand the biochemical consequences of glycogen accumulation in the Pompe mouse. We evaluated glycogen metabolism in heart, triceps, quadriceps, and liver from wild type and several strains of GAA−/− mice. Unexpectedly, we observed that lysosomal glycogen storage correlated with a robust increase in factors that normally promote glycogen biosynthesis. The GAA−/− mouse strains were found to have elevated glycogen synthase (GS), glycogenin, hexokinase, and glucose-6-phosphate (G-6-P, the allosteric activator of GS). Treating GAA−/− mice with recombinant human GAA (rhGAA) led to a dramatic reduction in the levels of glycogen, GS, glycogenin, and G-6-P. Lysosomal glycogen storage also correlated with a dysregulation of phosphorylase, which normally breaks down cytoplasmic glycogen. Analysis of phosphorylase activity confirmed a previous report that, although phosphorylase protein levels are identical in muscle lysates from wild type and GAA−/− mice, phosphorylase activity is suppressed in the GAA−/− mice in the absence of AMP. This reduction in phosphorylase activity likely exacerbates lysosomal glycogen accumulation. If the dysregulation in glycogen metabolism observed in the mouse model of Pompe disease also occurs in Pompe patients, it may contribute to the observed broad spectrum of disease severity.

WITHDRAWN: Clearance of lysosomal glycogen accumulation by transcription factor EB (TFEB) in muscle cells from lysosomal alpha-glucosidase deficient mice

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A novel m.12908T>A mutation in the mitochondrial ND5 gene in patient with infantile-onset Pompe disease

Pompe disease is a progressive metabolic myopathy caused by deficiency in lysosomal acid α-glucosidase and results in cellular lysosomal and cytoplasmic glycogen accumulation. A wide spectrum of clinical phenotypes exists from hypotonia and severe cardiac hypertrophy in the first few months of life to a milder form with the onset of symptoms in adulthood. The disease is typically due to severe mutations in GAA gene. In the present study, we described a newborn boy with clinical features of Pompe disease particularly with hypertrophic cardiomyopathy, hypotonia and hepatomegaly. This case was at first misdiagnosed as mitochondrial disorder. Accordingly, we performed a mitochondrial mutational analysis that revealed a novel mutation m.12908T>A in the ND5 gene. Secondary structure analysis of the ND5 protein further supported the deleterious role of the m.12908T>A mutation, as it was found to involve an extended imbalance in its hydrophobicity and affect its function.

Effects of exercise training during infusion on late-onset Pompe disease patients receiving enzyme replacement therapy

Pompe disease is an autosomal recessive disorder caused by the deficiency of acid α-glucosidase resulting in lysosomal accumulation of glycogen and abnormal autophagic function. The late-onset form of the disease is characterized by progressive skeletal and respiratory muscle dysfunction. Enzyme replacement therapy (ERT, Genzyme Corporation, Cambridge, MA, USA) was recently introduced and resulted in significant prolongation of the life expectancy of the patients with the infantile form while the results were less significant for the late-onset form. It has been postulated that the weak influence of ERT in late-onset patients might be due to a non-effective delivery of the recombinant enzyme to the skeletal muscles perhaps due to the relatively low blood flow to the resting skeletal muscles during infusion. Exercise training acutely increases the blood flow to the exercising muscles. Thus, it was hypothesized that exercise training during enzyme infusion might increase the effectiveness of the ERT therapy. Five late-onset Pompe disease patients receiving ERT and following regular exercise training for approximately 10months, followed a 6-month period of exercise training during infusion of the recombinant enzyme. Before and after this period, body composition, isometric strength and 6minute walking distance were determined. Analysis of the results revealed that none of these parameters changed significantly after the 6-month intervention period (e.g. 6minute walking distance before: 532±31m, vs. after: 527±29m, P=0.246). These results suggest that exercise training during infusion may not add significant functional changes in late-onset Pompe patients receiving ERT and undergoing regular exercise training.

T.P.46 A novel phase 2a study design to investigate drug-drug interactions between escalating doses of AT2220 (duvoglustat hydrochloride) and acid alpha-glucosidase in subjects with Pompe disease

Pompe disease is an inherited lysosomal storage disease that results from a deficiency in acid alpha-glucosidase (GAA) activity, and is characterized by progressive accumulation of lysosomal glycogen primarily in heart and skeletal muscles. Recombinant human GAA (rhGAA, Genzyme) is the only approved enzyme replacement therapy for Pompe, and is administered biweekly via intravenous infusion. While rhGAA provides clinical benefit, drawbacks as low stability at neutral pH/body temperature, modest tissue uptake and glycogen reduction, and immune responses affect tolerability and efficacy. AT2220 (1-deoxynojirimycin; duvoglustat HCl) is a pharmacological chaperone that selectively and reversibly binds and stabilizes endogenous GAA, facilitating proper folding and trafficking to lysosomes. AT2220-010 is an open-label, single ascending dose, fixed-sequence, two-period, dose-finding study comprised of four cohorts of 4–6 subjects with Pompe disease. Study objectives: (1) evaluate the safety of single ascending doses of AT2220 co-administered with rhGAA, and (2) characterize the pharmacokinetic interaction of AT2220 and rhGAA on activity and protein levels in plasma, muscle, and peripheral blood mononuclear cells, and on urine Hex four levels. Period 1, subjects receive rhGAA alone, then Period 2 they receive a single oral dose of AT2220 with rhGAA. Safety data from each completed dose cohort is reviewed by a Data Safety Monitoring Board. An innovative muscle biopsy technique, which requires specialized investigator training, is used to sample quadricep muscle either on Day 3 or 7 of each period. This unique crossover study design allows for meaningful evaluation of safety and pharmacokinetic data in the rare disease field by allowing each subject to serve as their own control and reducing variability as opposed to parallel designs requiring larger sample sizes for statistical comparisons.

T.P.45 An ongoing phase 2a study to investigate drug–drug interactions between escalating doses of AT2220 (duvoglustat hydrochloride) and acid alpha glucosidase in subjects with Pompe disease – Preliminary results

Pompe disease is an inherited lysosomal storage disease that results from a deficiency in acid alpha-glucosidase (GAA) activity, and is characterized by progressive accumulation of lysosomal glycogen primarily in heart and skeletal muscles. Recombinant human GAA (rhGAA, Genzyme) is the only approved enzyme replacement therapy for Pompe, and is administered biweekly via intravenous infusion. While rhGAA provides clinical benefit, drawbacks as low stability at neutral pH/body temperature, modest tissue uptake and glycogen reduction, and immune responses affect tolerability and efficacy. AT2220 (duvoglustat HCl) is a pharmacological chaperone which selectively and reversibly binds and stabilizes endogenous GAA, facilitating proper folding and trafficking to lysosomes. AT2220-010 is an open-label, single ascending dose, fixed-sequence, two-period, dose-finding study comprised of four cohorts of 4–6 subjects with Pompe disease. During the first period, subjects receive their regularly scheduled intravenous infusion of rhGAA 20mg/kg alone. During the second period they receive a single oral dose of AT2220 with rhGAA. The objectives of the study are (1) evaluate the safety of single ascending doses of AT2220 (50mg, 100mg, 250mg, and 600mg) and (2) characterize the pharmacokinetic interaction of AT2220 on rhGAA. Preliminary plasma, peripheral blood mononuclear cells, and muscle GAA activity, total GAA protein, and AT2220 concentrations as well as safety data including urine Hex 4 concentrations are available for the first and second cohorts. Anti-GAA antibodies are also being evaluated.

The Pharmacological Chaperone AT2220 Increases Recombinant Human Acid α-Glucosidase Uptake and Glycogen Reduction in a Mouse Model of Pompe Disease

Pompe disease is an inherited lysosomal storage disease that results from a deficiency in the enzyme acid α-glucosidase (GAA), and is characterized by progressive accumulation of lysosomal glycogen primarily in heart and skeletal muscles. Recombinant human GAA (rhGAA) is the only approved enzyme replacement therapy (ERT) available for the treatment of Pompe disease. Although rhGAA has been shown to slow disease progression and improve some of the pathophysiogical manifestations, the infused enzyme tends to be unstable at neutral pH and body temperature, shows low uptake into some key target tissues, and may elicit immune responses that adversely affect tolerability and efficacy. We hypothesized that co-administration of the orally-available, small molecule pharmacological chaperone AT2220 (1-deoxynojirimycin hydrochloride, duvoglustat hydrochloride) may improve the pharmacological properties of rhGAA via binding and stabilization. AT2220 co-incubation prevented rhGAA denaturation and loss of activity in vitro at neutral pH and 37°C in both buffer and blood. In addition, oral pre-administration of AT2220 to rats led to a greater than two-fold increase in the circulating half-life of intravenous rhGAA. Importantly, co-administration of AT2220 and rhGAA to GAA knock-out (KO) mice resulted in significantly greater rhGAA levels in plasma, and greater uptake and glycogen reduction in heart and skeletal muscles, compared to administration of rhGAA alone. Collectively, these preclinical data highlight the potentially beneficial effects of AT2220 on rhGAA in vitro and in vivo. As such, a Phase 2 clinical study has been initiated to investigate the effects of co-administered AT2220 on rhGAA in Pompe patients.

Autopsy findings in late-onset Pompe disease: A case report and systematic review of the literature

BackgroundLate-onset Pompe disease (LOPD) is a rare cause of declining proximal muscle strength and respiratory function that can also affect other organ systems. The development of enzyme replacement therapy has made it one of the few inherited muscle disorders with treatment, but clinical response is difficult to assess due to the variable and often slow progression of illness. A better understanding of the disease's systemic effects can be gleaned through autopsy findings. PurposeThe purpose of this study was to: (1) describe the histological findings observed in LOPD, (2) provide correlations between reported histological and clinical findings, and (3) review the literature on autopsy findings in LOPD. MethodsHistological evaluation of autopsy tissues from a 62-year-old woman with LOPD was conducted. A clinical history was obtained by review of the medical records. The literature was reviewed for previously reported histological and clinical findings in LOPD. Based on this case report and information from prior publications, histological and clinical findings for the disease were correlated. ResultsHistologic examination revealed mostly mild vacuolar myopathy typical of glycogen accumulation within skeletal and smooth muscle cells. The most prominent vacuolar myopathy was in quadriceps muscle, which also exhibited chronic myositis with degenerating and regenerating muscle fibers. Transmission electron microscopy disclosed lysosomal glycogen accumulation within skeletal, cardiac, and vascular smooth muscle cells, correlating with published case reports of basilar artery and ascending aortic aneurysms and carotid artery dissection. Organs containing smooth muscle cells (the bladder, intestine, and esophagus) were also affected, explaining reports of symptoms such as urinary incontinence and dysphagia. In addition to glycogen accumulation, there was obvious damage to the contraction apparatus of myofibrils within cardiac and skeletal muscle cells. These histological and ultrastructural findings correlate with the clinical manifestations of LOPD. ConclusionsThis study is the first to describe histological findings of LOPD utilizing both traditional paraffin-processed tissues and epoxy resin embedded tissues for high-resolution light microscopy. The findings are similar to those seen in previous studies, but with improved morphological detail and glycogen preservation. This patient exhibited histological involvement of multiple organs, correlating with the clinical features of LOPD. With the advent of definitive therapy for Pompe disease, it is important to be aware of these findings and use them to develop methods for tracking therapeutic response.

The impact of antibodies in late-onset Pompe disease: A case series and literature review

Pompe disease (glycogen storage disease type II, GSD II) is an autosomal recessive disease caused by a deficiency of acid α-glucosidase (GAA), leading to lysosomal glycogen accumulation in various tissues, most notably cardiac, skeletal and smooth muscle. While both infantile and late-onset patients have benefited greatly from alglucosidase alfa (Myozyme®) enzyme replacement therapy (ERT), a subgroup of patients does not demonstrate as pronounced a response as others. Various factors have been identified which may help predict the response to ERT in infantile Pompe disease patients. High, sustained antibody titers (HSAT) have been correlated with poor response to ERT in infantile Pompe cases. However, the literature on the role of antibodies in the late-onset Pompe disease (LOPD) population is limited. Our literature review highlights the need for studies to explore the potential impact of antibodies in LOPD. Further supporting the importance of this issue, our retrospective chart review of sixty LOPD patients revealed that six of these sixty (10%) LOPD patients developed HSAT of ≥1:51,200 on two or more occasions at or beyond 6months on ERT. Here, we present a series of three of these six LOPD patients for whom detailed antibody data and clinical data were available for greater than 1year on ERT. These three patients developed HSAT corresponding with clinical decline as demonstrated by pulmonary function, quality of life, and motor function testing, affirming the development of HSAT in a subset of patients with LOPD, and its potentially negative impact on clinical response to ERT. The findings of our study and literature review lead us to conclude that there is a strong indication for systematic studies to accurately delineate the potential impact of antibodies in LOPD.