Phosphorylase Kinase Deficiency Research Articles

Splicing abnormalities caused by a novel mutation in the PHKA2 gene in children with glycogen storage disease type IX

Objective: Glycogen storage disease type IX (GSD-IX) is a rare primary glucose metabolism abnormality caused by phosphorylase kinase deficiency and a series of pathogenic gene mutations. The clinical characteristics, gene analysis, and functional verification of a mutation in a child with hepatomegaly are summarized here to clarify the pathogenic cause of the disease. Methods: The clinical data of a child with GSD-IX was collected. Peripheral blood from the child and his parents was collected for genomic DNA extraction. The patient's gene diagnosis was performed by second-generation sequencing. The suspected mutations were verified by Sanger sequencing and bioinformatics analysis. The suspected splicing mutations were verified in vivo by RT-PCR and first-generation sequencing. Results: Hepatomegaly, transaminitis, and hypertriglyceridemia were present in children. Liver biopsy pathological examination results indicated glycogen storage disease. Gene sequencing revealed that the child had a c.285 + 2_285 + 5delTAGG hemizygous mutation in the PHKA2 gene. Sanger sequencing verification showed that the mother of the child was heterozygous and the father of the child was of the wild type. Software such as HSF3.1 and ESEfinder predicted that the gene mutation affected splicing. RT-PCR of peripheral blood from children and his mother confirmed that the mutation had caused the skipping of exon 3 during the constitutive splicing of the PHKA2 gene. Conclusion: The hemizygous mutation in the PHKA2 gene (c.285 + 2_285 + 5delTAGG) is the pathogenic cause of the patient's disease. The detection of the novel mutation site enriches the mutation spectrum of the PHKA2 gene and serves as a basis for the family's genetic counseling.

243.1: Pediatric Liver Transplantation Indications and Outcomes in Glycogen Storage Disease: A Single Center Experience

Introduction: Glycogen storage diseases (GSDs) are inherited metabolic disorders result from impaired glycogen storage, glycogen or glucose breakdown. Liver transplantation (LT) is considered as a treatment option for patients with glycogen storage diseases present with hepatic malignancy, liver failure or metabolic decompensation. GSD type Ia is associated with mutations in the G6PC gene due to deficiency of glucose-6-phosphotase, while glycogen storage disease type 9 results from phosphorylase kinase deficiency, an enzyme necessary for glycogen breakdown. Herein, we present our experience and transplantation outcomes in 7 pediatric patients with GSD who underwent LT. Methods: We retrospectively analyzed data from patients diagnosed with GSD who underwent LT between 1993-2021 years. Results: LT was performed in 6 patients with GSD type 1a and one patient with GSD type 9. Four patients had a family history of consanguinity (3 patients 1st degree, 1 patient 2nd degree). Mean age of diagnosis was 7.28 (1-24) months. Mean age of LT was 128.5 (13-216) months. Transplantation indications were hepatic adenoma (4/7), poor metabolic control (4/7) and cirrhosis (1/7). Extra-hepatic findings included proteinuria (4/7), Focal segmental glomerulosclerosis (1/7), proximal tubulopathy (4/7), nephrolithiasis (2/7), hyperlipidemia (7/7), delayed puberty (3/7), hypertension (2/7), short stature (6/7), epilepsy (2/7) and osteoporosis (4/7). One patient with GSD1a underwent cataract surgery. Three patients with GSD1a incidentally had the same G6PC gene mutations (c.247C>T). Three patients had normal cognitive functions and development (WISC-R Test). Two patients had mild where one patient had moderate mental retardation. One patient had %30 developmental delay (AGTE Test). Six patients underwent LT from living-related donors while one patient had cadaveric donor. Patients who underwent LT from live donors received left lateral segment. Explant liver pathology revealed variable numbers (1-12) of hepatic adenoma in 4 patients. Hepatocellular carcinoma (HCC) was detected in one patient. Serum lactate, uric acid and triglyceride levels normalized after LT. Tubulopathy resolved in one patient while epilepsy continued in two patients. Height SD scores were improved in 4 patients while remained unchanged in one patient after 5-year follow up. Acute graft rejection was detected in 4 patients. Mean follow up time was 80 (3-183) months. All patients are still alive and routinely followed up at our department. Conclusion: LT improved the quality of life in our GSD patients with metabolic decompensation and severe disease manifestations. Biochemical parameters (lactate, uric acid and triglyceride levels) normalized after transplantation. Our patients displayed improved height growth. LT should be considered in GSD patients with poor metabolic control despite dietary management, hepatic malignancy or progressive liver failure.

EP208: GSD IX natural history and novel liver disease severity score: Multicenter international collaboration uncovers longitudinal trends in liver disease severity

GSD IX is a glycogen storage disease which results from a deficiency of the enzyme phosphorylase kinase. While PhK is expressed in both muscle and liver, the muscle and liver isoforms are encoded by different genes, with the liver isozyme serving as the focus of this project. The clinical spectrum of hepatic GSD IX varies widely with genetic subtype (PHKA2, PHKB, PHKG2). Hepatic GSD IX presents early in life with mild to moderate hepatomegaly, growth retardation, fasting hyperketotic hypoglycemia, transaminitis, and elevated plasma lipids.

Generation of human induced pluripotent stem cell line, KRIBBi003-A, from urinary cells of a patient with glycogen storage disease type IXa

Glycogen storage disease type IXa (GSD IXa) is a rare genetic disorder characterized by phosphorylase kinase (PhK) deficiency, which leads to excessive glycogen accumulation in the liver. Urinary cells (UCs) were isolated from a GSD IXa patient and reprogrammed into induced pluripotent stem cells (iPSCs) using Sendai virus. The established iPSC line, KRIBBi003-A, exhibited pluripotency marker expression and a normal karyotype. The differentiation capacity of the cell line was confirmed by the differentiation of the three germ layers in vitro. The established iPSC line is a potential useful resource for disease modeling of GSD IXa.

The Phenotypic and Genetic Spectrum of Glycogen Storage Disease Type VI.

Glycogen storage disease type VI (GSD VI) is an autosomal recessive disorder of glycogen metabolism due to mutations in the glycogen phosphorylase gene (PYGL), resulting in a deficiency of hepatic glycogen phosphorylase. We performed a systematic literature review in order to collect information on the clinical phenotypes and genotypes of all published GSD VI patients and to compare the data to those for GSD IX, a biochemically and clinically very similar disorder caused by a deficiency of phosphorylase kinase. A total of 63 genetically confirmed cases of GSD VI with clinical information were identified (median age: 5.3 years). The age at presentation ranged from 5 weeks to 38 years, with a median of 1.8 years. The main presenting symptoms were hepatomegaly and poor growth, while the most common laboratory findings at initial presentation comprised elevated activity of liver transaminases, hypertriglyceridemia, fasting hypoglycemia and postprandial hyperlactatemia. Liver biopsies (n = 37) showed an increased glycogen content in 89.2%, liver fibrosis in 32.4% and early liver cirrhosis in 10.8% of cases, respectively. No patient received a liver transplant, and one successful pregnancy was reported. Our review demonstrates that GSD VI is a disorder with broad clinical heterogeneity and a small number of patients with a severe phenotype and liver cirrhosis. Neither clinical nor laboratory findings allow for a differentiation between GSD VI and GSD IX. Early biochemical markers of disease severity or clear genotype phenotype correlations are missing. Given the overall benign and unspecific phenotype and the need for enzymatic or genetic analyses for confirmation of the diagnosis, GSD VI is likely underdiagnosed. With new treatment approaches in sight, early, pre-symptomatic diagnosis, especially with respect to hepatic cirrhosis, will become even more important.

Male inheritance of X-linked liver glycogenosis from an undiagnosed maternal grandfather in a Chinese pedigree: a report of two cases

Abstract Hepatic phosphorylase kinase (PhK) plays an important role in glycogen metabolism by activating phosphorylase. Patients with PhK deficiency may get glycogen storage disease (GSD) type-IXa, an X-linked liver glycogenosis disease. To inform genetic counseling in a family with two affected GSD brothers, we performed a genetic analysis. The GSD in the older brother was confirmed by histological examination of a liver biopsy, which showed glycogen accumulation in liver cells. A liver biopsy was not available from the younger brother. The two patients and their parents were analyzed by whole exome sequencing. A pathogenic mutation in a gene encoding a regulatory subunit of PhK, PHKA2 located on chromosome Xp22, was identified as c.G3373A (p.E1125K) and confirmed by Sanger sequencing. The proband's maternal grandparents and the brothers and sisters of the proband's maternal grandfather were physically examined and genetically tested by Sanger sequencing. Pedigree analysis showed that the mother was a carrier and that the two patients inherited the mutation from their undiagnosed maternal grandfather. Moreover, among the maternal grandfather and four granduncles, three of them possessed the same mutation and four suffered from fatty liver. This is the first report of this mutation causing X-linked liver glycogenosis in a Chinese family and shows that GSD IXa is a mild form of glycogenosis in terms of clinical symptoms, indicating that GSD may be undiagnosed or underestimated. Nevertheless, to provide appropriate intervention and genetic counseling, early identification of the genetic cause is imperative. This study was approved by the Ethics Committee of First Affiliated Hospital, Hunan University of Chinese Medicine (approval No. HN-LL-ZFKY-2018-001-01) on January 12, 2018.

Molecular diagnosis of glycogen storage disease type IX using a glycogen storage disease gene panel.

Glycogen storage disease type IX (GSD IX) is caused by a deficiency of hepatic phosphorylase kinase. The aim of this study was to clarify the clinical features, long term outcomes, and genetic analysis of GSD IX in Korea. A GSD gene panel was created and hybridization capture-based next-generation sequencing was performed. We investigated clinical laboratory data, results of molecular genetic analysis, liver biopsy findings, and long-term outcomes. Ten children were diagnosed with GSD IX at Seoul National University Children's Hospital. Hypoglycemia, hyperlactacidemia, hypertriglyceridemia, hyperuricemia, liver fibrosis on liver biopsy, and short stature was found in 30%, 56%, 100%, 60%, 80% and 50% of the children, respectively. Seven PHKA2 variants were identified in eight children with GSD IXa-one nonsense (c.2268dupT; p.(Asp757Ter)), two splicing (c.918+1G>A, c.718-2A>G), one frameshift (c.405_419delinsTCCTGGCC; p.(Asp136ProfsTer11)), and three missense variants (c.3628G>A; p.(Gly1210Arg), c.1245G>T and c.2746C>T; p.(Arg916Trp)). Two variants of PHKG2 were identified in two children with GSD IXc-one frameshift (c.783delC; p.(Ser262AlafsTer6)) and one missense (c.661G>A; p.(Val221Met)). Elevated liver enzymes and hypertriglyceridemia in children with GSD IXa tended to improve with age. For the first time, we report hepatocellular carcinoma in a patient with GSD IXc. The GSD gene panel is a useful diagnostic tool to confirm GSD IX. The clinical phenotype of GSD IXc is severe and monitoring for the development of hepatocellular carcinoma should be implemented.

A novel PHKA2 mutation in a Chinese child with glycogen storage disease type IXa: a case report and literature review

BackgroundPHKA2 gene mutations can cause liver phosphorylase kinase (PhK) deficiency, resulting in glycogen storage disease type IXa (GSD IXa). Elevated liver transaminase levels and liver enlargement are the most frequent phenotypes of GSD IXa. However, whether the phenotypes are applicable to Chinese patients remains unclear.Case reportA boy aged 2 years and 8 months with a history of episodic fatigue and weakness since he was 2 years old was referred to our endocrinology clinic. Apart from symptomatic ketotic hypoglycemic episodes (palpitation, hand shaking, sweating, etc.), no abnormalities of liver transaminase levels or liver size were found. To identify the aetiology of his clinically diagnosed hypoglycaemia, the proband and his parents were screened for PHKA2 gene mutations by next-generation sequencing. A heterozygous mutation (c.2972C > G, p.G991A) in PHKA2 was found in the proband and his mother. Twenty-one Chinese cases with GSD IXa have been reported in the literature to date, and elevated liver transaminase levels (95%) and liver enlargement (91%) are the most frequent phenotypes of GSD IXa in Chinese patients. Hypoglycaemia may be one of the early onset symptoms in infants with GSD IXa.ConclusionsThis study enriches our knowledge of the PHKA2 gene mutation spectrum and provides further information about the phenotypic characteristics of Chinese GSD IXa patients.

Поиск и классификация генетических вариантов в генах фосфорилазкиназного комплекса у группы пациентов с подозрением на наследственные нарушения метаболизма гликогена

Glycogen storage diseases (GSD) are a heterogeneous group of inherited defects of metabolism of glycogen. GSD du to phosphorylase kinase (PhK) deficiency named GSD IX is the most common of them. It belongs to liver GSDs and occurs in several subtypes that differ in mode of inheritance and tissue-specificity. PhK consists of several subunits coding by different genes. The genes PHKA2, PHKB and PHKG2 make the main contribution to GSD IX etiology. The aim of this study is analysis of genetic specify and genotype-phenotype correlations in patients with mutations in genes of PhK complex. 36 children with liver pathology from 30 unrelated families were studied. Using massively parallel sequencing technology 47 target genes with mutations resulted in liver pathology were analyzed in patients. In 28 patients 18 different hemizygous mutant alleles of PHKA2 were found. In 4 patients 5 homozygous or compound-heterozygous mutations of PHKG2 were detected. 14 PHKA2 variants and 3 PHKG2 variantss are novel. Heterozygous mutations of PHKB and PHKG2 were found in 3 and 1 pts respectively. Clinical significance of some variants in PHKA2, PHKB and PHKG2 is unclear. Most of detected mutations are missense-mutations. There are a few unclear cases with unreliable missense variants and variants in heterozygous state combined with ambiguous clinical features. The analysis of determined novel mutations and its possible effects on PhK complex shows that the finding of a rare variant is not ample for the diagnosis. Diagnosis confirmation must be based on complex study of all available data of frequencies, homology, gene structure and enzyme activity and combine analysis of genetic, clinical and laboratory features and signs.

Two Cases of X-linked Liver Glycogenosis in Hunan Province: Transmission from the Undiagnosed Maternal Grandfather

X-linked liver glycogenosis (XLG), also known as glycogen storage disease (GSD) type-IXa, is characterized by hepatomegaly, abnormal liver functions and growth retardation. This disease is a result of a deficiency of hepatic phosphorylase kinase (PHK), which plays an important role in glycogen metabolism by activating phosphorylase. We aim at identifying the genetic cause of the GSD running in a family with two affected boys, and hence develop proper management and genetic counseling. The boys presented with typical GSD signs and symptoms. The histology of the liver biopsy from the old brother showed glycogen accumulation in hepatocytes and confirmed his condition of GSD. The younger brother did not have a biopsy. Whole exome sequencing was used to analyze the genetic structure of the proband patients and their parents. Sanger sequencing was used to validate and confirm the identified mutation. The results showed that there was a known pathogenic mutation (p.E1125K) in PHKA2 gene located on chromosome Xp22, which encodes the regulatory subunit of PHK. Pedigree analysis revealed that the mother was a carrier, and the disease of both brothers was transmitted through their undiagnosed maternal grandfather. This is the first report of XLG caused by this mutation in China, and it indicated that GSD may be undiagnosed or underestimated since GSD IXa is one of the mild form of glycogenosis in terms of clinical symptoms. However, it is necessary to identify the genetic cause in order to perform effective intervention and genetic counseling.

PO-0384 The Practical Method To Diagnosis Of Fourteen Cases Of Glycogen Storage Diseases In Our Laboratory

Glycogen storage diseases (GSD) are a group of inherited disorders of metabolism that result in storage of excess glycogen. Several well defined defects in one of the enzymes involved in the synthesis or degradation of glycogen have been described. There are over 15 types and they are classified based on the enzyme deficiency and the affected tissue (liver, muscle or both). In this study, we wish to report the biochemical investigations adopted in main infantile GSD diagnosed in our laboratory. Four steps diagnostic procedure have been assumed, taking into account several frequent clinical observations leading to further targeted biochemical parameters: Assessment of the metabolic disorders with standard tests (fast blood glucose, uric acid, triglycerides, total cholesterol, ASAT, ALAT, CK, lactic acid). Quantitative determination of glycogen in leucocytes (or erythrocytes) after extraction, precipitation and treatment with an throne reagent. Oral galactose test with blood lactate and glucose estimations, in combination with a glucagon tolerance test to screen the main types of liver glycogenosis. Lysosomal acid a-glucosidase activity when GSD type II (Pompe disease) is suspected. Since 1995 and on the basis of this screening procedure and clinical features, 14 cases of GSD have been categorised: · 6 forbes’s disease (GSD III, debranching-enzyme deficiency) · 3 von Gierke’s disease (GSD I, glucose-6-phosphatase deficiency) · 2 pompe’s disease (GSD II, maltase acid deficiency) · 1 Andersen’s disease (GSD IV, branching-enzyme deficiency) · 1 Hers’s disease (GSD VI, hepatic phosphorylase deficiency) · 1 GSD IX (phosphorylase kinase deficiency) Our laboratory diagnostic approach include simple screening tests easy to implement in clinical chemistry laboratories. Thus, Pompe disease diagnosis is easily done in our laboratory. The measurement of tissue enzyme activities (liver and muscle) of the other enzymes is limited to some specialised laboratories. The molecular diagnosis offers a good alternative for GSD type 0, I and III but requires better financial means.

Variability of disease spectrum in children with liver phosphorylase kinase deficiency caused by mutations in the PHKG2 gene

Liver phosphorylase b kinase (PhK) deficiency (glycogen storage disease type IX), one of the most common causes of glycogen storage disease, is caused by mutations in the PHKA2, PHKB, and PHKG2 genes. Presenting symptoms include hepatomegaly, ketotic hypoglycemia, and growth delay. Clinical severity varies widely. Autosomal recessive mutations in the PHKG2 gene, which cause about 10–15% of cases, have been associated with severe symptoms including increased risk of liver cirrhosis in childhood. We have summarized the molecular, biochemical, and clinical findings in five patients, age 5–16years, diagnosed with liver PhK deficiency caused by PHKG2 gene mutations. We have identified five novel and two previously reported mutations in the PHKG2 gene in these five patients. Clinical severity was variable among these patients. Histopathological studies were performed for four of the patients on liver biopsy samples, all of which showed signs of fibrosis but not cirrhosis. One of the patients (aged 9years) developed a liver adenoma which later resolved. All patients are currently doing well. Their clinical symptoms have improved with age and treatment. These cases add to the current knowledge of clinical variability in patients with PHKG2 mutations. Long term studies, involving follow-up of these patients into adulthood, are needed.

Novel PHKG2 mutation causing GSD IX with prominent liver disease: report of three cases and review of literature

Glycogen storage disease type IX (GSD IX) is a common form of glycogenosis due to mutations in PHKA1, PHKA2, or PHKB and PHKG2 genes resulting in the deficiency of phosphorylase kinase. The first two genes are X-linked while the latter two follow an autosomal recessive inheritance. The majority of cases of GSD IX are attributed to defects in PHKA2 which usually cause a mild disease. We report three patients with PHKG2-related GSD IX presenting with significant hepatic involvement, fibrosis, and cirrhosis. Interestingly, the homozygosity mapping resolved a dilemma about an erroneously normal phosphorylase kinase activity in patient 1. The novel mutation found in all the three patients (p.G220E) affects the catalytic subunit of the phosphorylase kinase. Increasing evidence shows that patients with PHKG2 mutations have a severe hepatic phenotype within the heterogeneous GSD IX disorder. Therefore, defect in PHKG2 should be considered in patients with suspected glycogenosis associated with significant liver fibrosis and cirrhosis.

Muscle phosphorylase kinase deficiency

To examine metabolism during exercise in 2 patients with muscle phosphorylase kinase (PHK) deficiency and to further define the phenotype of this rare glycogen storage disease (GSD). Patient 1 (39 years old) had mild exercise-induced forearm pain, and EMG showed a myopathic pattern. Patient 2 (69 years old) had raised levels of creatine kinase (CK) for more than 6 months after statin treatment. Both patients had increased glycogen levels in muscle and PHK activity <11% of normal. Two novel pathogenic nonsense mutations were found in the PHKA1 gene. The metabolic response to anaerobic forearm exercise and aerobic cycle exercise was studied in the patients and 5 healthy subjects. Ischemic exercise showed a normal 5-fold increase in plasma lactate (peak 5.7 and 6.9 mmol/L) but an exaggerated 5-fold increase in ammonia (peak 197 and 171 μmol/L; control peak range 60-113 μmol/L). An incremental exercise test to exhaustion revealed a blunted lactate response (5.4 and 4.8 mmol/L) vs that for control subjects (9.6 mmol/L; range 7.1-14.3 mmol/L). Fat and carbohydrate oxidation rates at 70% of peak oxygen consumption were normal. None of the patients developed a second wind phenomenon or improved their work capacity with an IV glucose infusion. Our findings demonstrate that muscle PHK deficiency may present as an almost asymptomatic condition, despite a mild impairment of muscle glycogenolysis, raised CK levels, and glycogen accumulation in muscle. The relative preservation of glycogenolysis is probably explained by an alternative activation of myophosphorylase by AMP and P(i) at high exercise intensities.

3D mapping of glycogenosis-causing mutations in the large regulatory alpha subunit of phosphorylase kinase

Mutations in the liver isoform of the Phosphorylase Kinase (PhK) α subunit (PHKA2 gene) cause X-linked liver glycogenosis (XLG), the most frequent type of PhK deficiency (glycogen-storage disease type IX). XLG patients can be divided in two subgroups, with similar clinical features but different activity of PhK (decreased in liver and blood cells for XLG-I and low in liver but normal or enhanced in blood cells for XLG-II). Here, we show that the PHKA2 missense mutations and small in-frame deletions/insertions are concentrated into two domains of the protein, which were recently described. In the N-terminal glucoamylase domain, mutations (principally leading to XLG-II) are clustered within the predicted glycoside-binding site, suggesting that they may have a direct impact on a possible hydrolytic activity of the PhK α subunit, which remains to be demonstrated. In the C-terminal calcineurin B-like domain (domain D), mutations (principally leading to XLG-I) are clustered in a region predicted to interact with the regulatory region of the PhK catalytic subunit and in a region covering this interaction site. Altogether, these results show that PHKA2 missense mutations or small in-frame deletions/insertions may have a direct impact on the PhK α functions and provide a framework for further experimental investigation.

G.P.16.03. A clinical and genetic study of 2 patients with myopathy and phosphorylase kinase deficiency (glycogenosis type VIII)

G.P.16.03. A clinical and genetic study of 2 patients with myopathy and phosphorylase kinase deficiency (glycogenosis type VIII)

Fatal Infantile Cardiac Glycogenosis with Phosphorylase Kinase Deficiency and a Mutation in the γ2-Subunit of AMP-Activated Protein Kinase

A 10-wk-old infant girl with severe hypertrophy of the septal and atrial walls by cardiac ultrasound, developed progressive ventricular wall thickening and died of aspiration pneumonia at 5 mo of age. Postmortem examination revealed ventricular hypertrophy and massive atrial wall thickening due to glycogen accumulation. A skeletal muscle biopsy showed increased free glycogen and decreased activity of phosphorylase b kinase (PHK). The report of a pathogenic mutation (R531Q) in the gene (PRKAG2) encoding the gamma2 subunit of AMP-activated protein kinase (AMPK) in three infants with congenital hypertrophic cardiomyopathy, glycogen storage, and "pseudo PHK deficiency" prompted us to screen this gene in our patient. We found a novel (R384T) heterozygous mutation in PRKAG2, affecting an arginine residue in the N-terminal AMP-binding domain. Like R531Q, this mutation reduces the binding of AMP and ATP to the isolated nucleotide-binding domains, and prevents activation of the heterotrimer by metabolic stress in intact cells. The mutation was not found in DNA from the patient's father, the only available parent, and is likely to have arisen de novo. Our studies confirm that mutations in PRKAG2 can cause fatal infantile cardiomyopathy, often associated with apparent PHK deficiency.

Identification of Alu-mediated, large deletion-spanning introns 19–26 in PHKA2 in a patient with X-linked liver glycogenosis (hepatic phosphorylase kinase deficiency)

X-linked liver glycogenosis (XLG) is one of the most common glycogen storage diseases. We present the first case of a large PHKA2 gene deletion from intron 19 to intron 26 in an XLG patient. An aberrant cDNA with skipping of exons 20–26 was detected. Alu element-mediated unequal homologous recombination between an Alu-Jo in intron 19 and another Alu-Sg in intron 26 appears to be responsible for this deletion.

Fatal Congenital Heart Glycogenosis Caused by a Recurrent Activating R531Q Mutation in the γ2-Subunit of AMP-Activated Protein Kinase ( PRKAG2), Not by Phosphorylase Kinase Deficiency

Fatal congenital nonlysosomal cardiac glycogenosis has been attributed to a subtype of phosphorylase kinase deficiency, but the underlying genes and mutations have not been identified. Analyzing four sporadic, unrelated patients, we found no mutations either in the eight genes encoding phosphorylase kinase subunits or in the two genes encoding the muscle and brain isoforms of glycogen phosphorylase. However, in three of five patients, we identified identical heterozygous R531Q missense mutations of the PRKAG2 gene, which encodes the gamma 2-subunit of AMP-activated protein kinase, a key regulator of energy balance. Biochemical characterization of the recombinant R531Q mutant protein showed >100-fold reduction of binding affinities for the regulatory nucleotides AMP and ATP but an enhanced basal activity and increased phosphorylation of the alpha -subunit. Other PRKAG2 missense mutations were previously identified in patients with autosomal dominant hypertrophic cardiomyopathy with Wolff-Parkinson-White syndrome, characterized by juvenile-to-adult clinical onset, moderate cardiac glycogenosis, disturbed excitation conduction, risk of sudden cardiac death in midlife, and molecular perturbations that are similar to--but less severe than--those observed for the R531Q mutation. Thus, recurrent heterozygous R531Q missense mutations in PRKAG2 give rise to a massive nonlysosomal cardiac glycogenosis of fetal symptomatic onset and rapidly fatal course, constituting a genotypically and clinically distinct variant of hypertrophic cardiomyopathy with Wolff-Parkinson-White syndrome. R531Q and other PRKAG2 mutations enhance the basal activity and alpha -subunit phosphorylation of AMP-activated protein kinase, explaining the dominant nature of PRKAG2 disease mutations. Since not all cases displayed PRKAG2 mutations, fatal congenital nonlysosomal cardiac glycogenosis seems to be genetically heterogeneous. However, the existence of a heart-specific primary phosphorylase kinase deficiency is questionable, because no phosphorylase kinase mutations were found.

Biochemical characteristics and increased tetraglucoside excretion in patients with phosphorylase kinase deficiency

Patients with glycogen storage disease type IXa present with infantile hepatomegaly and a specific growth pattern, and variable biochemical alterations in blood. We studied the clinical and biochemical characteristics including the urinary oligosaccharide excretion of seven unrelated children. The urinary tetraglucoside excretion was increased in four children, three of whom had persistently high cholesterol and triglyceride concentrations. We propose screening for urine tetraglucoside excretion and the measurement of serum cholesterol in patients with growth delay and/or hepatomegaly to assess a possible glycogenosis.