G. Peter A. Smit

Enzyme replacement therapy in late-onset Pompe's disease : A three-year follow-up

Pompes disease is an autosomal recessive myopathy. The characteristic lysosomal storage of glycogen is caused by acid α‐glucosidase deficiency. Patients with late‐onset Pompes disease present with progressive muscle weakness also affecting pulmonary function. In search of a treatment, we investigated the feasibility of enzyme replacement therapy with recombinant human α‐glucosidase from rabbit milk. Three patients (aged 11, 16, and 32 years) were enrolled in the study. They were all wheelchair‐bound and two of them were ventilator dependent with a history of deteriorating pulmonary function. After 3 years of treatment with weekly infusions of α‐glucosidase, the patients had stabilized pulmonary function and reported less fatigue. The youngest and least affected patient showed an impressive improvement of skeletal muscle strength and function. After 72 weeks of treatment, he could walk without support and finally abandoned his wheelchair. Our findings demonstrate that recombinant human α‐glucosidase from rabbit milk has a therapeutic effect in late‐onset Pompes disease. There is good reason to continue the development of enzyme replacement therapy for Pompes disease and to explore further the production of human therapeutic proteins in the milk of mammals. Ann Neurol 2004;55:000–000

Glycogen storage disease type I: diagnosis, management, clinical course and outcome. Results of the European study on glycogen storage disease type I (ESGSD I)

Abstract. Glycogen storage disease type I (GSD I) is a relatively rare metabolic disease and therefore, no metabolic centre has experience of large numbers of patients. To document outcome, to develop guidelines about (long-term) management and follow-up, and to develop therapeutic strategies, the collaborative European Study on GSD I (ESGSD I) was initiated. This paper is an descriptive analysis of data obtained from the retrospective part of the ESGSD I. Included were 231 GSD Ia and 57 GSD Ib patients. Median age of data collection was 10.4 years (range 0.4–45.4 years) for Ia and 7.1 years (0.4–30.6 years) for Ib patients. Data on dietary treatment, pharmacological treatment, and outcome including mental development, hyperlipidaemia and its complications, hyperuricaemia and its complications, bleeding tendency, anaemia, osteopenia, hepatomegaly, liver adenomas and carcinomas, progressive renal disease, height and adult height, pubertal development and bone maturation, school type, employment, and pregnancies are presented. Data on neutropenia, neutrophil dysfunction, infections, inflammatory bowel disease, and the use of granulocyte colony-stimulating factor are presented elsewhere (Visser et al. 2000, J Pediatr 137:187–191; Visser et al. 2002, Eur J Pediatr DOI 10.1007/s00431-002-1010-0). Conclusion: there is still wide variation in methods of dietary and pharmacological treatment of glycogen storage disease type I. Intensive dietary treatment will improve, but not correct completely, clinical and biochemical status and fewer patients will die as a direct consequence of acute metabolic derangement. With ageing, more and more complications will develop of which progressive renal disease and the complications related to liver adenomas are likely to be two major causes of morbidity and mortality.

Molecular characterization of hepatocellular adenomas developed in patients with glycogen storage disease type I

BACKGROUND & AIMS Hepatocellular adenomas (HCA) are benign liver tumors mainly related to oral contraception and classified into 4 molecular subgroups: inflammatory (IHCA), HNF1A-inactivated (H-HCA), β-catenin-activated (bHCA) or unclassified (UHCA). Glycogen storage disease type I (GSD) is a rare hereditary metabolic disease that predisposes to HCA development. The aim of our study was to characterize the molecular profile of GSD-associated HCA. METHODS We characterized a series of 25 HCAs developed in 15 patients with GSD by gene expression and DNA sequence of HNF1A, CTNNB1, IL6ST, GNAS, and STAT3 genes. Moreover, we searched for glycolysis, gluconeogenesis, and fatty acid synthesis alterations in GSD non-tumor livers and compared our results to those observed in a series of sporadic H-HCA and various non-GSD liver samples. RESULTS GSD adenomas were classified as IHCA (52%) mutated for IL6ST or GNAS, bHCA (28%) or UHCA (20%). In contrast, no HNF1A inactivation was observed, showing a different molecular subtype distribution in GSD-associated HCA from that observed in sporadic HCA (p = 0.0008). In non-tumor GSD liver samples, we identified glycolysis and fatty acid synthesis activation with gluconeogenesis repression. Interestingly, this gene expression profile was similar to that observed in sporadic H-HCA. CONCLUSIONS Our study showed a particular molecular profile in GSD-related HCA characterized by a lack of HNF1A inactivation. This exclusion could be explained by similar metabolic defects observed with HNF1A inactivation and glucose-6-phosphatase deficiency. Inversely, the high frequency of β-catenin mutations could be related to the increased frequency of malignant transformation in hepatocellular carcinoma.

A novel defect of peroxisome division due to a homozygous non-sense mutation in the PEX11β gene

Background Peroxisomes are organelles that proliferate continuously and play an indispensable role in human metabolism. Consequently, peroxisomal gene defects can cause multiple, often severe disorders, including the peroxisome biogenesis disorders. Currently, 13 different PEX proteins have been implicated in various stages of peroxisome assembly and protein import. Defects in any of these proteins result in a peroxisome biogenesis disorder. The authors present here a novel genetic defect specifically affecting the division of peroxisomes. Methods The authors have studied biochemical and microscopical peroxisomal parameters in cultured patient fibroblasts, sequenced candidate PEX genes and determined the consequence of the identified PEX11β gene defect on peroxisome biogenesis in patient fibroblasts at different temperatures. Results The patient presented with congenital cataracts, mild intellectual disability, progressive hearing loss, sensory nerve involvement, gastrointestinal problems and recurrent migraine-like episodes. Although microscopical investigations of patient fibroblasts indicated a clear defect in peroxisome division, all biochemical parameters commonly used for diagnosing peroxisomal disorders were normal. After excluding mutations in all PEX genes previously implicated in peroxisome biogenesis disorders, it was found that the defect was caused by a homozygous non-sense mutation in the PEX11β gene. The peroxisome division defect was exacerbated when the patients fibroblasts were cultured at 40°C, which correlated with a marked decrease in the expression of PEX11γ. Conclusions This novel isolated defect in peroxisome division expands the clinical and genetic spectrum of peroxisomal disorders and indicates that peroxisomal defects exist, which cannot be diagnosed by standard laboratory investigations.

Increased de novo Lipogenesis and Delayed Conversion of Large VLDL into Intermediate Density Lipoprotein Particles Contribute to Hyperlipidemia in Glycogen Storage Disease Type 1a

Glycogen storage disease type 1a (GSD-1a) is a metabolic disorder characterized by fasting-induced hypoglycemia, hepatic steatosis, and hyperlipidemia. The mechanisms underlying the lipid abnormalities are largely unknown. To investigate these mechanisms seven GSD-1a patients and four healthy control subjects received an infusion of [1-13C]acetate to quantify cholesterogenesis and lipogenesis. In a subset of patients, [1-13C]valine was given to assess lipoprotein metabolism and [2-13C]glycerol to determine whole body lipolysis. Cholesterogenesis was 274 ± 112 mg/d in controls and 641 ± 201 mg/d in GSD-1a patients (p < 0.01). Plasma triglyceride-palmitate derived from de novo lipogenesis was 7.1 ± 9.4 and 86.3 ± 42.5 μmol/h in controls and patients, respectively (p < 0.01). Production of VLDL did not show a consistent difference between the groups, but conversion of VLDL into intermediate density lipoproteins was relatively retarded in all patients (0.6 ± 0.5 pools/d) compared with controls (4.3 ± 1.8 pools/d). Fractional catabolic rate of intermediate density lipoproteins was lower in patients (0.8 ± 0.6 pools/d) compared with controls (3.1 ± 1.5 pools/d). Whole body lipolysis was similar, i.e., 4.5 ± 1.9 μmol/kg/min in patients and 3.8 ± 1.9 μmol/kg/min in controls. Hyperlipidemia in GSD-1a is associated with strongly increased lipid production and a slower relative conversion of VLDL to LDL.

The Glycogen Storage Diseases and Related Disorders

The liver glycogen storage disorders (GSDs) comprise GSD I, the hepatic presentations of GSD III, GSD IV, GSD VI, the liver forms of GSD IX, and GSD 0. GSD I, III, VI, and IX present similarly with hypoglycemia, marked hepatomegaly, and growth retardation. GSD I is the most severe affecting both glycogen breakdown and gluconeogenesis. In GSD Ib there is additionally a disorder of neutrophil function. Most patients with GSD III have a syndrome that includes hepatopathy, myopathy, and often cardiomyo pathy. GSD VI and GSD IX are the least severe: there is only a mild tendency to fasting hypoglycemia, liver size normalises with age, and patients reach normal adult height. GSD IV manifests in most patients in infancy or childhood as hepatic failure with cirrhosis leading to end-stage liver disease. GSD 0 presents in infancy or early childhood with fasting hypoglycemia and ketosis and, in contrast, with postprandial hyperglycemia and hyperlactatemia. Treatment is primarily dietary and aims to prevent hypoglycemia and suppress secondary metabolic decompensation. This usually requires frequent feeds by day, and in GSD I and in some patients with GSD III, continuous nocturnal gastric feeding.

Renal Function in Glycogen Storage Disease Type I, Natural Course, and Renopreservative Effects of ACE Inhibition

BACKGROUND AND OBJECTIVES Renal failure is a major complication in glycogen storage disease type I (GSD I). We studied the natural course of renal function in GSD I patients. We studied differences between patients in optimal and nonoptimal metabolic control and possible renoprotective effects of angiotensin converting enzyme inhibition. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS Thirty-nine GSD I patients that visited our clinic were studied. GFR and effective renal plasma flow (ERPF) were measured by means of I(125) iothalamate and I(131) hippuran clearance and corrected for body surface area. Microalbuminuria was defined as >2.5 mg albumin/mmol creatinine and proteinuria as >0.2 g protein per liter. Optimal metabolic control was present when blood glucoses were >3.5 mmol/L, urine lactate/creatinine ratios <0.06 mmol/mmol, triglycerides <6.0 mmol/L, and uric acid concentrations <450 micromol/L. RESULTS Quadratic regression analysis showed a biphasic pattern in the course of GFR and ERPF related to age. Microalbuminuria was observed significantly less frequently in the patients with optimal metabolic control compared with the patients with nonoptimal metabolic control. A significant decrease in GFR was observed after starting ACE inhibition. CONCLUSIONS This study describes a biphasic pattern of the natural course of GFR and ERPF in GSD I patients, followed by the development of microalbuminuria and proteinuria. Optimal metabolic control has a renoprotective effect on the development of microalbuminuria and proteinuria in GSD I patients. Treatment with ACE inhibitors significantly decreases the GFR, especially in GSD I patients with glomerular hyperfiltration.

Disturbed hepatic carbohydrate management during high metabolic demand in medium-chain acyl-CoA dehydrogenase (MCAD)-deficient mice

Medium‐chain acyl–coenzyme A (CoA) dehydrogenase (MCAD) catalyzes crucial steps in mitochondrial fatty acid oxidation, a process that is of key relevance for maintenance of energy homeostasis, especially during high metabolic demand. To gain insight into the metabolic consequences of MCAD deficiency under these conditions, we compared hepatic carbohydrate metabolism in vivo in wild‐type and MCAD−/− mice during fasting and during a lipopolysaccharide (LPS)‐induced acute phase response (APR). MCAD−/− mice did not become more hypoglycemic on fasting or during the APR than wild‐type mice did. Nevertheless, microarray analyses revealed increased hepatic peroxisome proliferator‐activated receptor gamma coactivator‐1α (Pgc‐1α) and decreased peroxisome proliferator‐activated receptor alpha (Ppar α) and pyruvate dehydrogenase kinase 4 (Pdk4) expression in MCAD−/− mice in both conditions, suggesting altered control of hepatic glucose metabolism. Quantitative flux measurements revealed that the de novo synthesis of glucose‐6‐phosphate (G6P) was not affected on fasting in MCAD−/− mice. During the APR, however, this flux was significantly decreased (−20%) in MCAD−/− mice compared with wild‐type mice. Remarkably, newly formed G6P was preferentially directed toward glycogen in MCAD−/− mice under both conditions. Together with diminished de novo synthesis of G6P, this led to a decreased hepatic glucose output during the APR in MCAD−/− mice; de novo synthesis of G6P and hepatic glucose output were maintained in wild‐type mice under both conditions. APR‐associated hypoglycemia, which was observed in wild‐type mice as well as MCAD−/− mice, was mainly due to enhanced peripheral glucose uptake. Conclusion: Our data demonstrate that MCAD deficiency in mice leads to specific changes in hepatic carbohydrate management on exposure to metabolic stress. This deficiency, however, does not lead to reduced de novo synthesis of G6P during fasting alone, which may be due to the existence of compensatory mechanisms or limited rate control of MCAD in murine mitochondrial fatty acid oxidation. (HEPATOLOGY 2008.)

The difference between observed and expected prevalence of MCAD deficiency in The Netherlands: a genetic epidemiological study

Medium chain acyl coenzyme A dehydrogenase (MCAD) deficiency is assumed to be the most common inherited disorder of mitochondrial fatty acid oxidation. Few reports mention the difference between the expected and observed prevalence of MCAD deficiency on the basis of the carrier frequency in the population. We performed a population-wide retrospective analysis of all known MCAD-deficient patients in The Netherlands. In this study, the observed prevalence of MCAD deficiency in The Netherlands was 1/27 400 (95% confidence interval (CI) 1/23 000–1/33 900), significantly different from the expected prevalence of 1/12 100 (95% CI 1/8450–1/18 500). The observed prevalence of MCAD deficiency showed a remarkable north–south trend within the country. From the patients in this cohort, it can be observed that underdiagnosis contributes to a larger extent to the difference between the expected and observed prevalences of MCAD deficiency in our country, than reduced penetrance. We determined estimates of the segregation proportion in a cohort of 73 families under the assumption of complete ascertainment (pLM=0.41, 95% CI 0.31–0.51) and single ascertainment (pD=0.28, 95% CI 0.19–0.37). With the expectation–maximization algorithm, a third estimate was obtained (pEM=0.28, 95% CI 0.20–0.37). The agreement between the latter two estimates supports incomplete selection and the segregation proportions were in agreement with normal mendelian autosomal recessive inheritance.

Inhibition of mitochondrial fatty acid oxidation in vivo only slightly suppresses gluconeogenesis but enhances clearance of glucose in mice

Mitochondrial fatty acid oxidation (mFAO) is considered to be essential for driving gluconeogenesis (GNG) during fasting. However, quantitative in vivo data on de novo synthesis of glucose‐6‐phosphate upon acute inhibition of mFAO are lacking. We assessed hepatic glucose metabolism in vivo after acute inhibition of mFAO by 30 mg kg−1 2‐tetradecylglycidic acid (TDGA) in hypoketotic hypoglycemic male C57BL/6J mice by the infusion of [U‐13C]glucose, [2‐13C]glycerol, [1‐2H]galactose, and paracetamol for 6 hours, which was followed by mass isotopomer distribution analysis in blood glucose and urinary paracetamol‐glucuronide. During TDGA treatment, endogenous glucose production was unaffected (127 ± 10 versus 118 ± 7 μmol kg−1 minute−1, control versus TDGA, not significant), but the metabolic clearance rate of glucose was significantly enhanced (15.9 ± 0.9 versus 26.3 ± 1.1 mL kg−1 minute−1, control versus TDGA,P < 0.05). In comparison with control mice, de novo synthesis of glucose‐6‐phosphate (G6P) was slightly decreased in TDGA‐treated mice (108 ± 19 versus 85 ± 6 μmol kg−1 minute−1, control versus TDGA, P < 0.05). Recycling of glucose was decreased upon TDGA treatment (26 ± 14 versus 12 ± 4 μmol kg−1 minute−1, control versus TDGA, P < 0.05). Hepatic messenger RNA (mRNA) levels of genes encoding enzymes involved in de novo G6P synthesis were unaltered, whereas glucose‐6‐phosphate hydrolase mRNA expressions were increased in TDGA‐treated mice. Glucokinase and pyruvate kinase mRNA levels were significantly decreased, whereas pyruvate dehydrogenase kinase isozyme 4 expression was increased 30‐fold; this suggested decreased glycolytic activity. Conclusion: Acute pharmacological inhibition of mFAO using TDGA had no effect on endogenous glucose production and only a marginal effect on de novo G6P synthesis. Hence, fully active mFAO is not essential for maintenance of hepatic GNG in vivo in fasted mice.(HEPATOLOGY 2008.)