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Screening for late-onset Pompe disease in Finland
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ScienceDirect Neuromuscular Disorders 24 (2014) 982–985 www.elsevier.com/locate/nmd
Screening for late-onset Pompe disease in Finland Johanna Palmio a,⇑, Mari Auranen b,c, Sari Kiuru-Enari b, Mervi Lo¨fberg b, Olaf Bodamer d, Bjarne Udd a,e,f a Neuromuscular Research Center, Tampere University Hospital and University of Tampere, Tampere, Finland Department of Neurology, Unit for Neuromuscular Diseases, Helsinki University Central Hospital, Helsinki, Finland c Research Programs Unit, Molecular Neurology, University of Helsinki, 00029 Helsinki, Finland d Department of Human Genetics, Miller School of Medicine, University of Miami, Miami, USA e Folkha¨lsan Institute of Genetics and Department of Medical Genetics, Haartman Institute, University of Helsinki, Helsinki, Finland f Neurology Department, Vaasa Central Hospital, Vaasa, Finland b
Received 28 March 2014; received in revised form 15 May 2014; accepted 20 June 2014
Abstract Pompe disease (glycogen storage disease type II) is caused by autosomal recessive mutations in GAA gene. The estimated frequency of late-onset Pompe disease is around 1:60,000. However, only two infantile and one late-onset Pompe patients have been reported in Finland with a population of 5 million. We screened for late-onset Pompe disease in a cohort of undetermined myopathy patients with proximal muscle weakness and/or elevated serum creatine kinase values. Acid a-glucosidase (GAA) activity in dried blood spots was measured and clinical data collected in 108 patients. Four patients had low normal GAA activity; all the others had activities well within the normal range. Re-analyses of these patients did not reveal new Pompe patients. Our findings suggest that Pompe disease is extremely rare in Finland. Finland is an example of an isolated population with enrichment of certain mutations for genetic disorders and low occurrence of some autosomal recessive diseases. Ó 2014 Elsevier B.V. All rights reserved.
Keywords: Pompe disease; Glycogen storage disease type II; Late-onset; Screening
1. Introduction Pompe disease, also known as glycogen storage disease type II (GSD II) or acid maltase deficiency (OMIM 232300) is a lysosomal storage disorder characterized by an absence or deficiency of acid a-glucosidase (GAA). The disease is caused by autosomal recessive mutations in GAA gene [1]. The gene mutations lead to a varying degree of GAA deficiency; total or nearly total loss of GAA activity results in classic infantile form of the disease, whereas late-onset disease patients may have GAA activity ranging from 2% to 40% [2]. Deficiency of ⇑ Corresponding author. Tel.: +358 3 3116111; fax: +358 3 35516164.
E-mail address: johanna.palmio@uta.fi (J. Palmio). http://dx.doi.org/10.1016/j.nmd.2014.06.438 0960-8966/Ó 2014 Elsevier B.V. All rights reserved.
the GAA enzyme results in glycogen accumulation in lysosomes causing tissue damage, especially in skeletal, respiratory, and cardiac muscles [3]. Infantile-onset Pompe is severe rapidly progressive, and if not treated, fatal disease which is characterized by hypotonia, feeding difficulties and cardiorespiratory insufficiency. Late-onset disease presents with a more heterogeneous phenotype including overlap of signs and symptoms with other neuromuscular diseases [4]. Enzyme replacement therapy (ERT) with alglucosidase alfa can stabilize or improve symptoms in Pompe patients. Thus, accurate and early diagnosis also in late-onset disease is needed [5]. The measurement of GAA activity in dried blood spots (DBS) has been successfully used to screen
J. Palmio et al. / Neuromuscular Disorders 24 (2014) 982–985
for Pompe disease, and is suggested to be the optimal initial diagnostic test procedure [6,7]. Based on the data from the Dutch and USA population the estimated frequency of late-onset Pompe disease is around 1:60,000, and of all types 1:40,000 [8,9]. To date in Finland, only two infantile Pompe patients were previously reported [10], and one molecularly diagnosed late-onset patient is currently on enzyme replacement therapy [11]. The low number of diagnosed patients prompted us for a screening project to know whether or not late-onset Pompe patients have remained undiagnosed in Finland. We screened for late-onset Pompe disease in a cohort of myopathy patients in Finland using GAA activity measurement in DBS [12,13]. Based on these findings Pompe disease seems to be a very rare disease entity in the Finnish population in contrast to many central European populations. 2. Materials and methods We screened 108 Finnish patients with undetermined proximal myopathy, with or without respiratory symptoms for late-onset Pompe disease. All patients were of Finnish origin. The patients had been examined by neurologists at our nationwide neuromuscular diagnostic center in Tampere and at the neuromuscular outpatient clinic in Helsinki, Finland. All patients had also been examined by electrophysiological nerve conduction velocity (NCV) studies and electromyography (EMG), by muscle biopsy, creatine kinase (CK) measurements, and in most of the patients by muscle imaging, computed tomography (CT) or MRI. The inclusion criteria for selected patients were proximal myopathy of unknown etiology with or without distal weakness based on findings such as muscle weakness, exercise intolerance and myalgia, elevated serum CK levels, abnormal muscle histology, or myopathic and/or myotonic electromyography. Pulmonary function tests and/or polysomnography had been performed in patients with respiratory symptoms or signs. Blood samples were collected after informed consent and the study was performed in accordance with the Helsinki declaration. 2.1. Blood samples A single 3 mm spot was punched from the dried blood on the filter card into a 96 well plate containing extraction buffer and incubated at 37 °C for 1h Following extraction the reminder of the spot was removed and the assay cocktail was added. The assay cocktail contains a tandem mass spectrometry (MS–MS) specific GAA substrate and internal standard. The solution is incubated overnight and subjected to a series of liquid–liquid extraction and one final solid phase extraction step. The final solution is dried and
983
reconstituted in an ethylacetate/methanol mixture prior to injection into the MS_MS (ABI 3200). The MS_MS is run in positive ion mode using multiple –reaction monitoring mode for the analysis of GAA internal standard and product using the following trasitions: m/z 503 > 403 (IS) and m/z 498 > 398 (product). For details see our recent publication [14]. Normal reference value for GAA activity was P3 lmol/l/ h. Three patients were retested with GAA activity measurement in DBS at the other laboratory (Lysosomal Laboratory Eppendorf in Hamburg, Germany), and by conventional biochemistry in whole leukocytes at a public service laboratory (University Hospital of Kuopio, Finland). 2.2. Gene sequencing Genomic DNA was extracted from venous blood according to standard procedures in two patients. GAA exons 10, 11, 12 covering the two known Finnish mutations were screened. Primer sequences for the exons studied were designed to include the entire exon and exon–intron borders and are available upon request. 3. Results Included in the study were 55 male and 53 female patients with a mean age of 50.8 years (range 18–80 y). The age at symptom onset varied from four to 75 years of age (mean 40.2 y). Fourteen patients had disease onset in childhood or adolescence (range 4–18 y). The majority of patients had disease onset in adulthood (n = 67, range 19–50 y) and the rest had late-onset disease (n = 24, range 51–75 y). Most patients had a diagnosis of limbgirdle muscular dystrophy (LGMD), or proximal muscle weakness. There were also 34 patients whose main symptoms were myalgia, muscle weakness and/or fatigue especially related to exercise and elevated CK values. The rest of the patients had miscellaneous symptoms and diagnoses (Table 1). Twenty-three patients had respiratory involvement. Of these, eight patients had respiratory insufficiency and restriction with decreased forced vital capacity (FVC) ranging from 21% to 50%. Eight patients had symptoms of dyspnea and in two of them FVC was slightly reduced (70%). Three had a diagnosis of asthma, four of sleep apnea/nocturnal hypoxia and one was treated with a non-invasive ventilator. EMG findings were consistent with myopathy in most patients but remained non-diagnostic in 34 patients. Six patients had myotonia on EMG. CK levels were normal in 40% of the patients; the other patients had a variable degree of elevated CK levels, reaching up to 30 times the upper limit of normal, and there were five cases of rhabdomyolysis. Muscle biopsy had been performed in all but one patient with non-diagnostic histology in 22 biopsies. Histopathological findings were mostly
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J. Palmio et al. / Neuromuscular Disorders 24 (2014) 982–985
Table 1 Clinical characteristics of the patients. Age at disease onset
Number of patients
4–18 years 19–50 years 51–75 years Not available Diagnosis LGMD or proximal weakness Myalgia, exercise intolerance Proximal and distal weakness Rhabdomyolysis Axial myopathy Rigid spine Not available Respiratory symptoms Insufficiency or restriction(FVC 40–50%) Dyspnea Sleep apnea/nocturnal hypoxia Asthma
14 67 24 3 56 34 6 5 2 1 4 8 8 3 3
LGMD, limb-girdle muscular dystrophy; FVC, forced vital capacity.
nonspecific mild to moderate myopathic or dystrophic changes. Neurogenic findings were observed in a few samples. Metabolic myopathy was suspected only in two cases based on vacuolar pathology on muscle biopsies. 3.1. Enzyme activity findings The GAA enzyme activity was normal in all patients, ranging from 3.0 to 39.5 lmol/l/h (mean 9.8 lmol/l/h). Enzyme activity was low (1.8 lmol/l/h) in the first sample of one patient, but the control sample yielded a normal result (3.5 lmol/l/h). This patient, as well as three other patients, who had normal low levels of activity (3.0– 3.9 lmol/l/h), were retested with GAA activity measurement in DBS (Lysosomal Laboratory Eppendorf in Hamburg, Germany), and by conventional biochemistry in whole leukocytes. The confirmatory tests were otherwise normal but two of these patients had low activity levels in the second DBS but normal activity levels in leukocytes. GAA exons 10, 11 and 12 were screened in them: no mutations in GAA gene were identified. 4. Discussion Diagnosis of adult onset Pompe disease can be very difficult, especially because characteristic muscle histopathology findings of lysosomal glycogenosis and autophagic vacuoles may be totally absent. Based on estimated incidences, there should be at least one new patient diagnosed per year in Finland [11]. Since only one late-onset patient has been identified during the last 10 years, the question was raised whether the low occurrence was real or not. However, in our screening project including 108 undetermined myopathy patients no new cases of Pompe disease were discovered. In a similar screening study of 103 unclassified neuromuscular disease
patients in Denmark, three Pompe patients were found [15]. Our result indicates that the frequency of Pompe disease in Finland is lower than in other European countries. The estimated incidence of Pompe disease varies between different clinical subtypes and different ethnic groups. The combined incidence of all types of Pompe disease ranges from 1:14,000 in African Americans to 1:100,000 in individuals of European descent [2]. Based on the calculations of three most common GAA mutations carrier frequencies in the Netherlands, it has been estimated that the predicted frequency of the disease should be 1:40,000, whereas it is 138,000 for infantile Pompe and 1:57,000 for adult disease [8]. Roughly the same estimates for a population of European descent were reported in a study from the United States [9]. In Finland, two cases of infantile or childhood onset Pompe disease have previously been reported [10], and currently there is only one patient diagnosed with late-onset Pompe [11]. To our knowledge, no other Pompe patients have been identified during these last four decades in Finland. Although we cannot exclude that exceptional cases of late onset Pompe disease may have remained undiagnosed, the present study clearly indicates that the frequency of the disease in Finland is much lower than in other European populations. The reason for the low frequency is most likely due to the unusual population history in Finland. The three most common mutations in the Dutch population (IVS1–13 T > G, 525delT and Ex18del) account for a major part of the total disease alleles in that population [16]. The molecularly diagnosed Finnish patient is compound heterozygous for two different mutations (Y575X and P545L) suggesting that the most common GAA mutations in central Europe are not present in the Finnish population to the same degree. The exons 10, 11 and 12 were sequenced in two of our patients with low enzyme activity in retesting. The mutations identified earlier in Finland are in exons 11 and 12, although, other possible mutations in GAA gene cannot be excluded. The next generation sequencing of known myopathy genes, including GAA, is currently ongoing in these patients. The lack of major common European mutations may well explain the lower occurrence in Finland. Finland is known as an example of a genetically isolated population [17]. The size of the original founder population was very small and population growth encountered several bottle-necks during the last 2000 years. Genetic drift and founder effects with enrichment of certain mutations have two types of consequences: about 30 autosomal recessive disorders and two dominant diseases are much more common in Finland than in any other population. On the other hand, some elsewhere frequent autosomal recessive diseases are either non-existent or very rare in Finland [18], such as Friedreich ataxia, cystic fibrosis, phenylketonuria, galactosaemia or maple syrup urine disease. Pompe disease is apparently another example of disorders with lower frequency in Finland.
J. Palmio et al. / Neuromuscular Disorders 24 (2014) 982–985
In conclusion, GAA activity measurement with DBS is a simple and reliable test which clinicians should readily apply in undiagnosed myopathy patients also in Finland despite the low frequency of the disease. References [1] Kishnani PS, Steiner RD, Bali D, et al. Pompe disease diagnosis and management guideline. Genet Med 2006;8:267–88. [2] Leslie N, Tinkle BT. Glycogen storage disease type II (Pompe Disease) In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong CT, Stephens K, editors. GeneReviewse [Internet]. Seattle (WA): Seattle, University of Washington; 1993–2013. 2007 Aug 31 [updated 2013 May 09]. [3] Kishnani PS, Howell RR. Pompe disease in infants and children. J Pediatr 2004;144:35–43. [4] Byrne BJ, Kishnani PS, Case LE, et al. Pompe disease: design, methodology, and early findings from the Pompe Registry. Mol Genet Metab 2011;103:1–11. [5] Cupler EJ, Berger KI, Leshner RT, et al. AANEM consensus committee on late-onset Pompe disease: AANEM consensus committee on late-onset Pompe disease. consensus treatment recommendations for late-onset Pompe disease. Muscle Nerve 2012;45:319–33. [6] Zhang H, Kallwass H, Young SP, et al. Comparison of maltose and acarbose as inhibitors of maltase-glucoamylase activity in assaying acid alpha-glucosidase activity in dried blood spots for the diagnosis of infantile Pompe disease. Genet Med 2006;8:302–6. [7] Mechtler TP, Metz TF, Mu¨ller HG, et al. Short-incubation mass spectrometry assay for lysosomal storage disorders in newborn and high-risk population screening. J Chromatogr B Analyt Technol Biomed Life Sci 2012;908:9–17.
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[8] Ausems MG, Verbiest J, Hermans MP, et al. Frequency of glycogen storage disease type II in The Netherlands: implications for diagnosis and genetic counseling. Eur J Hum Genet 1999;7:713–6. [9] Martiniuk F, Chen A, Mack A, et al. Carrier frequency for glycogen storage disease type II in New York and estimates of affected individuals born with the disease. Am J Med Genet 1998;79:69–72. [10] Vanto T, Salmi TT, Kalimo H, et al. Pompe’s disease or glycogen storage disease. Duodecim 1982;98:709–16 [Finnish]. [11] Korpela MP, Paetau A, Lo¨fberg MI, Timonen MH, Lamminen AE, Kiuru-Enari SM. A novel mutation of the GAA gene in a Finnish late-onset Pompe disease patient: clinical phenotype and follow-up with enzyme replacement therapy. Muscle Nerve 2009;40:143–8. [12] Chamoles NA, Niizawa G, Blanco M, Gaggioli D, Casentini C. Glycogen storage disease type II: enzymatic screening in dried blood spots on filter paper. Clin Chim Acta 2004;347:97–102. [13] Winchester B, Bali D, Bodamer OA, et al. Pompe disease diagnostic working group methods for a prompt and reliable laboratory diagnosis of Pompe disease: report from an international consensus meeting. Mol Genet Metab 2008;93:275–81. [14] Bodamer OA, Dajnoki A. Diagnosing lysosomal storage disorders: Pompe disease. Curr Protoc Hum Genet 2012 Oct; Chapter 17, Unit 17. [15] Preisler N, Lukacs Z, Vinge L, et al. Late-onset Pompe disease is prevalent in unclassified limb-girdle muscular dystrophies. Mol Genet Metab 2013;110:287–9. [16] Kroos MA, Van der Kraan M, Van Diggelen OP, et al. Glycogen storage disease type II: frequency of three common mutant alleles and their associated clinical phenotypes studied in 121 patients. J Med Genet 1995;32:836–7. [17] de la Chapelle A. Disease gene mapping in isolated human populations: the example of Finland. J Med Genet 1993;30:857–65. [18] Norio R, Nevanlinna HR, Perheentupa J. Hereditary diseases in Finland; rare flora in rare soil. Ann Clin Res 1973;5:109–41.
ScienceDirect Neuromuscular Disorders 24 (2014) 982–985 www.elsevier.com/locate/nmd
Screening for late-onset Pompe disease in Finland Johanna Palmio a,⇑, Mari Auranen b,c, Sari Kiuru-Enari b, Mervi Lo¨fberg b, Olaf Bodamer d, Bjarne Udd a,e,f a Neuromuscular Research Center, Tampere University Hospital and University of Tampere, Tampere, Finland Department of Neurology, Unit for Neuromuscular Diseases, Helsinki University Central Hospital, Helsinki, Finland c Research Programs Unit, Molecular Neurology, University of Helsinki, 00029 Helsinki, Finland d Department of Human Genetics, Miller School of Medicine, University of Miami, Miami, USA e Folkha¨lsan Institute of Genetics and Department of Medical Genetics, Haartman Institute, University of Helsinki, Helsinki, Finland f Neurology Department, Vaasa Central Hospital, Vaasa, Finland b
Received 28 March 2014; received in revised form 15 May 2014; accepted 20 June 2014
Abstract Pompe disease (glycogen storage disease type II) is caused by autosomal recessive mutations in GAA gene. The estimated frequency of late-onset Pompe disease is around 1:60,000. However, only two infantile and one late-onset Pompe patients have been reported in Finland with a population of 5 million. We screened for late-onset Pompe disease in a cohort of undetermined myopathy patients with proximal muscle weakness and/or elevated serum creatine kinase values. Acid a-glucosidase (GAA) activity in dried blood spots was measured and clinical data collected in 108 patients. Four patients had low normal GAA activity; all the others had activities well within the normal range. Re-analyses of these patients did not reveal new Pompe patients. Our findings suggest that Pompe disease is extremely rare in Finland. Finland is an example of an isolated population with enrichment of certain mutations for genetic disorders and low occurrence of some autosomal recessive diseases. Ó 2014 Elsevier B.V. All rights reserved.
Keywords: Pompe disease; Glycogen storage disease type II; Late-onset; Screening
1. Introduction Pompe disease, also known as glycogen storage disease type II (GSD II) or acid maltase deficiency (OMIM 232300) is a lysosomal storage disorder characterized by an absence or deficiency of acid a-glucosidase (GAA). The disease is caused by autosomal recessive mutations in GAA gene [1]. The gene mutations lead to a varying degree of GAA deficiency; total or nearly total loss of GAA activity results in classic infantile form of the disease, whereas late-onset disease patients may have GAA activity ranging from 2% to 40% [2]. Deficiency of ⇑ Corresponding author. Tel.: +358 3 3116111; fax: +358 3 35516164.
E-mail address: johanna.palmio@uta.fi (J. Palmio). http://dx.doi.org/10.1016/j.nmd.2014.06.438 0960-8966/Ó 2014 Elsevier B.V. All rights reserved.
the GAA enzyme results in glycogen accumulation in lysosomes causing tissue damage, especially in skeletal, respiratory, and cardiac muscles [3]. Infantile-onset Pompe is severe rapidly progressive, and if not treated, fatal disease which is characterized by hypotonia, feeding difficulties and cardiorespiratory insufficiency. Late-onset disease presents with a more heterogeneous phenotype including overlap of signs and symptoms with other neuromuscular diseases [4]. Enzyme replacement therapy (ERT) with alglucosidase alfa can stabilize or improve symptoms in Pompe patients. Thus, accurate and early diagnosis also in late-onset disease is needed [5]. The measurement of GAA activity in dried blood spots (DBS) has been successfully used to screen
J. Palmio et al. / Neuromuscular Disorders 24 (2014) 982–985
for Pompe disease, and is suggested to be the optimal initial diagnostic test procedure [6,7]. Based on the data from the Dutch and USA population the estimated frequency of late-onset Pompe disease is around 1:60,000, and of all types 1:40,000 [8,9]. To date in Finland, only two infantile Pompe patients were previously reported [10], and one molecularly diagnosed late-onset patient is currently on enzyme replacement therapy [11]. The low number of diagnosed patients prompted us for a screening project to know whether or not late-onset Pompe patients have remained undiagnosed in Finland. We screened for late-onset Pompe disease in a cohort of myopathy patients in Finland using GAA activity measurement in DBS [12,13]. Based on these findings Pompe disease seems to be a very rare disease entity in the Finnish population in contrast to many central European populations. 2. Materials and methods We screened 108 Finnish patients with undetermined proximal myopathy, with or without respiratory symptoms for late-onset Pompe disease. All patients were of Finnish origin. The patients had been examined by neurologists at our nationwide neuromuscular diagnostic center in Tampere and at the neuromuscular outpatient clinic in Helsinki, Finland. All patients had also been examined by electrophysiological nerve conduction velocity (NCV) studies and electromyography (EMG), by muscle biopsy, creatine kinase (CK) measurements, and in most of the patients by muscle imaging, computed tomography (CT) or MRI. The inclusion criteria for selected patients were proximal myopathy of unknown etiology with or without distal weakness based on findings such as muscle weakness, exercise intolerance and myalgia, elevated serum CK levels, abnormal muscle histology, or myopathic and/or myotonic electromyography. Pulmonary function tests and/or polysomnography had been performed in patients with respiratory symptoms or signs. Blood samples were collected after informed consent and the study was performed in accordance with the Helsinki declaration. 2.1. Blood samples A single 3 mm spot was punched from the dried blood on the filter card into a 96 well plate containing extraction buffer and incubated at 37 °C for 1h Following extraction the reminder of the spot was removed and the assay cocktail was added. The assay cocktail contains a tandem mass spectrometry (MS–MS) specific GAA substrate and internal standard. The solution is incubated overnight and subjected to a series of liquid–liquid extraction and one final solid phase extraction step. The final solution is dried and
983
reconstituted in an ethylacetate/methanol mixture prior to injection into the MS_MS (ABI 3200). The MS_MS is run in positive ion mode using multiple –reaction monitoring mode for the analysis of GAA internal standard and product using the following trasitions: m/z 503 > 403 (IS) and m/z 498 > 398 (product). For details see our recent publication [14]. Normal reference value for GAA activity was P3 lmol/l/ h. Three patients were retested with GAA activity measurement in DBS at the other laboratory (Lysosomal Laboratory Eppendorf in Hamburg, Germany), and by conventional biochemistry in whole leukocytes at a public service laboratory (University Hospital of Kuopio, Finland). 2.2. Gene sequencing Genomic DNA was extracted from venous blood according to standard procedures in two patients. GAA exons 10, 11, 12 covering the two known Finnish mutations were screened. Primer sequences for the exons studied were designed to include the entire exon and exon–intron borders and are available upon request. 3. Results Included in the study were 55 male and 53 female patients with a mean age of 50.8 years (range 18–80 y). The age at symptom onset varied from four to 75 years of age (mean 40.2 y). Fourteen patients had disease onset in childhood or adolescence (range 4–18 y). The majority of patients had disease onset in adulthood (n = 67, range 19–50 y) and the rest had late-onset disease (n = 24, range 51–75 y). Most patients had a diagnosis of limbgirdle muscular dystrophy (LGMD), or proximal muscle weakness. There were also 34 patients whose main symptoms were myalgia, muscle weakness and/or fatigue especially related to exercise and elevated CK values. The rest of the patients had miscellaneous symptoms and diagnoses (Table 1). Twenty-three patients had respiratory involvement. Of these, eight patients had respiratory insufficiency and restriction with decreased forced vital capacity (FVC) ranging from 21% to 50%. Eight patients had symptoms of dyspnea and in two of them FVC was slightly reduced (70%). Three had a diagnosis of asthma, four of sleep apnea/nocturnal hypoxia and one was treated with a non-invasive ventilator. EMG findings were consistent with myopathy in most patients but remained non-diagnostic in 34 patients. Six patients had myotonia on EMG. CK levels were normal in 40% of the patients; the other patients had a variable degree of elevated CK levels, reaching up to 30 times the upper limit of normal, and there were five cases of rhabdomyolysis. Muscle biopsy had been performed in all but one patient with non-diagnostic histology in 22 biopsies. Histopathological findings were mostly
984
J. Palmio et al. / Neuromuscular Disorders 24 (2014) 982–985
Table 1 Clinical characteristics of the patients. Age at disease onset
Number of patients
4–18 years 19–50 years 51–75 years Not available Diagnosis LGMD or proximal weakness Myalgia, exercise intolerance Proximal and distal weakness Rhabdomyolysis Axial myopathy Rigid spine Not available Respiratory symptoms Insufficiency or restriction(FVC 40–50%) Dyspnea Sleep apnea/nocturnal hypoxia Asthma
14 67 24 3 56 34 6 5 2 1 4 8 8 3 3
LGMD, limb-girdle muscular dystrophy; FVC, forced vital capacity.
nonspecific mild to moderate myopathic or dystrophic changes. Neurogenic findings were observed in a few samples. Metabolic myopathy was suspected only in two cases based on vacuolar pathology on muscle biopsies. 3.1. Enzyme activity findings The GAA enzyme activity was normal in all patients, ranging from 3.0 to 39.5 lmol/l/h (mean 9.8 lmol/l/h). Enzyme activity was low (1.8 lmol/l/h) in the first sample of one patient, but the control sample yielded a normal result (3.5 lmol/l/h). This patient, as well as three other patients, who had normal low levels of activity (3.0– 3.9 lmol/l/h), were retested with GAA activity measurement in DBS (Lysosomal Laboratory Eppendorf in Hamburg, Germany), and by conventional biochemistry in whole leukocytes. The confirmatory tests were otherwise normal but two of these patients had low activity levels in the second DBS but normal activity levels in leukocytes. GAA exons 10, 11 and 12 were screened in them: no mutations in GAA gene were identified. 4. Discussion Diagnosis of adult onset Pompe disease can be very difficult, especially because characteristic muscle histopathology findings of lysosomal glycogenosis and autophagic vacuoles may be totally absent. Based on estimated incidences, there should be at least one new patient diagnosed per year in Finland [11]. Since only one late-onset patient has been identified during the last 10 years, the question was raised whether the low occurrence was real or not. However, in our screening project including 108 undetermined myopathy patients no new cases of Pompe disease were discovered. In a similar screening study of 103 unclassified neuromuscular disease
patients in Denmark, three Pompe patients were found [15]. Our result indicates that the frequency of Pompe disease in Finland is lower than in other European countries. The estimated incidence of Pompe disease varies between different clinical subtypes and different ethnic groups. The combined incidence of all types of Pompe disease ranges from 1:14,000 in African Americans to 1:100,000 in individuals of European descent [2]. Based on the calculations of three most common GAA mutations carrier frequencies in the Netherlands, it has been estimated that the predicted frequency of the disease should be 1:40,000, whereas it is 138,000 for infantile Pompe and 1:57,000 for adult disease [8]. Roughly the same estimates for a population of European descent were reported in a study from the United States [9]. In Finland, two cases of infantile or childhood onset Pompe disease have previously been reported [10], and currently there is only one patient diagnosed with late-onset Pompe [11]. To our knowledge, no other Pompe patients have been identified during these last four decades in Finland. Although we cannot exclude that exceptional cases of late onset Pompe disease may have remained undiagnosed, the present study clearly indicates that the frequency of the disease in Finland is much lower than in other European populations. The reason for the low frequency is most likely due to the unusual population history in Finland. The three most common mutations in the Dutch population (IVS1–13 T > G, 525delT and Ex18del) account for a major part of the total disease alleles in that population [16]. The molecularly diagnosed Finnish patient is compound heterozygous for two different mutations (Y575X and P545L) suggesting that the most common GAA mutations in central Europe are not present in the Finnish population to the same degree. The exons 10, 11 and 12 were sequenced in two of our patients with low enzyme activity in retesting. The mutations identified earlier in Finland are in exons 11 and 12, although, other possible mutations in GAA gene cannot be excluded. The next generation sequencing of known myopathy genes, including GAA, is currently ongoing in these patients. The lack of major common European mutations may well explain the lower occurrence in Finland. Finland is known as an example of a genetically isolated population [17]. The size of the original founder population was very small and population growth encountered several bottle-necks during the last 2000 years. Genetic drift and founder effects with enrichment of certain mutations have two types of consequences: about 30 autosomal recessive disorders and two dominant diseases are much more common in Finland than in any other population. On the other hand, some elsewhere frequent autosomal recessive diseases are either non-existent or very rare in Finland [18], such as Friedreich ataxia, cystic fibrosis, phenylketonuria, galactosaemia or maple syrup urine disease. Pompe disease is apparently another example of disorders with lower frequency in Finland.
J. Palmio et al. / Neuromuscular Disorders 24 (2014) 982–985
In conclusion, GAA activity measurement with DBS is a simple and reliable test which clinicians should readily apply in undiagnosed myopathy patients also in Finland despite the low frequency of the disease. References [1] Kishnani PS, Steiner RD, Bali D, et al. Pompe disease diagnosis and management guideline. Genet Med 2006;8:267–88. [2] Leslie N, Tinkle BT. Glycogen storage disease type II (Pompe Disease) In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong CT, Stephens K, editors. GeneReviewse [Internet]. Seattle (WA): Seattle, University of Washington; 1993–2013. 2007 Aug 31 [updated 2013 May 09]. [3] Kishnani PS, Howell RR. Pompe disease in infants and children. J Pediatr 2004;144:35–43. [4] Byrne BJ, Kishnani PS, Case LE, et al. Pompe disease: design, methodology, and early findings from the Pompe Registry. Mol Genet Metab 2011;103:1–11. [5] Cupler EJ, Berger KI, Leshner RT, et al. AANEM consensus committee on late-onset Pompe disease: AANEM consensus committee on late-onset Pompe disease. consensus treatment recommendations for late-onset Pompe disease. Muscle Nerve 2012;45:319–33. [6] Zhang H, Kallwass H, Young SP, et al. Comparison of maltose and acarbose as inhibitors of maltase-glucoamylase activity in assaying acid alpha-glucosidase activity in dried blood spots for the diagnosis of infantile Pompe disease. Genet Med 2006;8:302–6. [7] Mechtler TP, Metz TF, Mu¨ller HG, et al. Short-incubation mass spectrometry assay for lysosomal storage disorders in newborn and high-risk population screening. J Chromatogr B Analyt Technol Biomed Life Sci 2012;908:9–17.
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