Glycogen storage diseases: Diagnosis, treatment and outcome

2.1.1Clinical presentation

Affected individuals usually present in the first year of life with severe fasting hypoglycemia, hepatomegaly, failure to thrive, growth retardation, and developmental delay. Other common findings related to hypoglycemia include sweating, irritability, muscle weakness, drowsiness, and seizures. Symptoms usually become apparent as infants are weaned from frequent feeds. In addition to severe fasting hypoglycemia, biochemical studies reveal hyperlactatemia, hyperuricemia, and hypertriglyceridemia. Children often experience bruising and epistaxis due to impaired platelet function, and normochromic anemia may be present.

Children with GSD type Ia develop a markedly protuberant abdomen due to massive stores of liver glycogen. The spleen, however, remains normal in size and cirrhosis does not develop. Other physical findings include truncal obesity, doll-like facies, short stature, and hypotrophic muscles. With optimal metabolic control, the hepatomegaly improves and growth normalizes. Complications including hepatic adenomas, osteoporosis, focal segmental glomerulosclerosis, and a small fiber neuropathy used to be common in the 2nd and 3rd decades of life, but the frequency of these complications has markedly decreased with improvements in therapy and good metabolic control [9, 10].

Management of hepatic adenomas when they occur remains a source of debate. Although these tumors are mostly benign, malignant transformation to carcinoma may occur in 5–10% of patients. Most adenomas appear during puberty, and they stabilize following adolescence if metabolic control is optimized. Recently, regression of hepatic adenomas has been reported with improvement in patients whose metabolic control improved [11]. Since hepatocellular carcinoma in GSD Ia arises from adenomas, frequent imaging of adenomas with MRI and ultrasounds is commonly used. If changes are appreciated in adulthood (especially in lesions over 5 cm), surgical resection of the liver lesion or transplantation is usually recommended.

Since glucose-6-phosphatase is also in the kidneys, renal complications can also occur. “Silent” glomerular hyperfiltration almost universally develops in the teen years and may be followed by microalbuminuria and later overt proteinuria. Decreased glomerular filtration rate is due to focal segmental glomerulosclerosis and interstitial fibrosis. Other renal abnormalities include dysfunction of the proximal and/or distal tubules. Dysfunction of the proximal tubules leads to Type II renal tubular acidosis, and distal tubular dysfunction is associated with hypercalciuria. Furthermore, metabolically compensated patients show hypocitraturia that worsens with age [12]. Treatment with ACE inhibitors can slow the progression of kidney damage, and improved metabolic control may slow or even reverse renal disease. Unlike other complications in GSD Ia, kidney stone formation is not primarily related to metabolic control. Hypocitraturia develops in most people with GSD Ia during adolescence, and citrate supplementation has been successful at preventing renal calcification.

Patients with large hepatic adenomas may have severe, iron refractory anemia. This anemia has been observed to resolve spontaneously after adenoma resection or liver transplantation. Based upon these findings, it was determined that large adenomas may express inappropriately high levels of hepcidin mRNA [13]. Hepcidin is a peptide hormone that has been implicated as the key regulator of iron by controlling iron absorption across the enterocyte and macrophage recycling of iron. The increased hepcidin expression in the GSD adenomas is thought to interrupt iron availability and cause iron restricted anemia.

2.1.2Genetics

GSD Type Ia has a disease incidence of approximately 1 in 100,000 births and a carrier rate of approximately 1 in 150. The disorder is found in ethnic groups from all over the world, and the disease is more common in people of Ashkenazi Jewish, Mormon, Mexican, and Chinese heritage [14–16]. The disorder is associated with mutations in the G6PC gene on chromosome 17q21 which encodes the glucose-6-phosphatase-α catalytic subunit. GSD Ia has classic autosomal recessiveinheritance.

G6PC spans 12.6 kilobases and is composed of five exons which encode for a hydrophobic, endoplasmic reticulum (ER)-associated glycoprotein [17]. Despite the gene’s compact size, almost 100 different disease-causing mutations have been reported [18, 19]. Sequencing analysis of all coding exons plus flanking intronic sequence is predicted to detect approximately 94% of cases [20, 21].

2.1.3Pathology

While liver biopsies are no longer required for diagnosing this condition, glycogen filled hepatocytes with prominent steatosis are seen in GSD type Ia. Unlike other forms of GSD, however, fibrosis and cirrhosis do not occur. Hepatocellular carcinoma appears to arise from inflammatory adenomas, and chromosomal alterations have been described in the cancerous lesions with proto-oncogene activation leading to dysregulation of insulin-glucagon-growth hormone signaling [22]. In patients with von Gierke disease, the inability to convert glucose-6-phosphate to glucose results in shunting of G6P to the pentose phosphate shunt and the glycolytic pathway. This, in turn, results in increased synthesis of uric acid, fatty acids and triglycerides. The block in gluconeogenesis also leads to a buildup of lactate and pyruvate which can enter the Krebs cycle and lead to alterations in ATP/ADP and NADH/NAD⁺ ratios that may favor tumorigenesis [23, 24].

2.1.4Treatment

Dietary treatment has immensely improved prognosis. The aim of treatment is to prevent hypoglycemia and counter-regulation thereby minimizing the secondary metabolic derangements. Therapy may consist of continuous gastric tube feeds or uncooked cornstarch may be used depending on the age of the child and patient/family preferences.

Cornstarch therapy can be introduced as early as six months of age, as soon as the baby’s pancreatic amylase activity has matured. Due to intestinal immaturity, diarrhea can limit cornstarch use, and children less than 2 years of age usually require feeds every 3 to 3-1/2 hours. Cornstarch feeds can be spaced usually to every 4-5 hours in older children and adults. The dose of cornstarch can be estimated by calculating the basal glucose production rate using the following formula: y = 0.0014x³ – 0.214x²+10.411 – 9.084 where y = mg/kg/min of glucose and x = weight in kg. Doses of cornstarch are typically individualized based upon home and clinic based glucose and lactate monitoring in order to maintain glucose concentrations above 75 mg/dL and lactate less than 2.2 mmol/L. Cornstarch is mixed with water or a sugar-free liquid in a starch:water ratio of 1 : 2. Adding glucose is not recommended since it stimulates insulin production and offsets the advantage of the starch. Of note, a new extended release formulation (Glycosade) was recently introduced for night feeds, and it has allowed older children and adults to have a 7–10 hour period of coverage without sacrificing metabolic control [25].

Intake of galactose, sucrose, and fructose is restricted since these sugars will worsen the hepatomegaly and metabolic derangements. The GSD diet is very prohibitive, and it can be difficult for individuals to get all required nutrients without multivitamin supplementation. Other medications are also commonly used to prevent complications. Allopurinol is prescribed when serum urate concentrations are elevated, and fish oil supplementation or a prescription fibrate may be used to lower triglycerides and reduce the risk of pancreatitis. Treatment with an angiotensin-converting enzyme (ACE) inhibitor is used in patients with proteinuria to reduce intraglomerular capillary pressure and provide renoprotection. Preventive calcium and vitamin D₃ supplementation is also recommended to prevent osteoporosis.

Most patients with GSD Ia are clinically doing well into adulthood, and complications are becoming uncommon as metabolic control has improved. Many successful pregnancies have occurred [26]. At times, intravenous glucose support may be required. Patients with even minor infections or intercurrent illnesses who are vomiting and/or unable to eat should be taken to the hospital. Surgery should be undertaken with caution due to a bleeding tendency and risk of intraoperative lactic acidosis. Orthotopic liver transplantation has been performed for some individuals with unresectable adenomas or hepatocellular carcinoma. Liver transplantation, however, is deemed a treatment of last resort since renal failure has been a common complication due to the impact of immunosuppression on abnormal kidneys [27].

2.2.1Clinical presentation

Early in life, patients with GSD Ib may be clinically and metabolically identical to those with GSD Ia. With aging, however, most patients develop neutropenia and inflammatory bowel disease. The neutropenia is the hallmark feature of GSD Ib, but the age of onset and clinical course are variable. It may be present at birth or not appear until late in childhood as cyclic or permanent neutropenia.

Inflammatory bowel disease resembling Crohn’s disease is a major complication. This nearly universal complication usually appears between 5–12 years of age, but cases as young as 13 months have been reported. Unlike inflammatory bowel disease in the general population, GSD enterocolitis is most commonly located in the small intestine [28]. Diarrhea and abdominal pain may be late manifestations of the co-morbidity, and it often presents as growth failure, severe anemia, or perioral infections. A normal colonoscopy does not rule out the condition, and a capsule endoscopy sometimes is required to establish its presence.

2.2.2Genetics

Type Ib accounts for only 10–15% of type I GSD. While rare in the general population (1 in 1,000,000 individuals), high risk populations include people of Native American, Iranian Jewish, and Italian heritage. The SLC37A4 gene is located on 11q23.3 and the gene has nine exons [29–31]. Mutations in exon 8 are found in 50% of the documented cases, but full gene sequencing may be required to establish the diagnosis since over 90 mutations have been reported [19, 32].

2.2.3Pathology

The histologic appearance of a GSD Ib liver is identical to that of GSD Ia. Establishing the diagnosis of GSD Ib is therefore a challenge since enzymatic testing cannot be relied upon. While almost all glycogenolytic enzymes are found in the cytoplasm, glucose-6-phosphatase is localized to the inner luminal wall of the endoplasmic reticulum. This means that glucose-6-phosphate must cross the membrane of the endoplasmic reticulum in order to act as substrate for glucose-6-phosphatase. This transport protein for glucose-6-phosphate is defective in GSD Ib. Measurement of glucose-6-phosphate translocase activity is difficult to measure, however, and requires fresh unfrozen liver tissue. While liver sample with intact hepatocytes and microsomes will show deficient glucose-6-phosphatase activity because the translocase cannot deliver the G6P substrate to the ER lumen, microsomes disrupted by solubilization or damage from freezing will show normal glucose-6-phosphatase enzyme activity because the substrate is now readily accessible. Due to the difficulty of the biochemical assay, most clinical diagnostic laboratories do not offer such testing and diagnosis by molecular genetic testing is recommended [21].

2.2.4Treatment

Treatment guidelines for patients with GSD Ib are similar to those for GSD Ia with the addition of therapy for the neutropenia and GSD enterocolitis. Recombinant human granulocyte-colony-stimulating factor (GCSF), a cytokine that induces proliferation and differentiation of bone marrow precursor cells into mature neutrophils, should be used to treat neutropenia if infections, severe mouth ulcers, or chronic diarrhea are occurring. The GSD Ib population has been prone to untoward effects (massive splenomegaly, splenic sequestration, splenic rupture, and portal hypertension) with GCSF therapy. Therefore, a starting dose of 2.5 micrograms/kg/day is recommended, and the lowest dose that controls infections should be used [33]. Supplementation with high dose vitamin E appears to boost the neutrophil count and improve function in GSD Ib, and supplementation may allow lower GCSF doses to be used [34]. Non-absorbable salicylates (Pentasa, Asacol, and Lialda) are the first line therapies for GSD enterocolitis. Steroids and immunomodulators must be used with caution due to the metabolic consequences and associated immune dysfunction [34].

3.5.1Pathology

The biochemical hallmark of the disease is the accumulation of pathologic vacuoles containing glycogen or intermediary metabolites, mainly in skeletal and myocardial muscle with no evidence of enzyme deficiency.

3.5.2Genetics

Danon disease is quite rare and good estimates of the incidence are not available. The disorder is X-linked dominant in nature and is due to LAMP-2 (lysosome-associated membrane protein-2) deficiency. Although biochemical analysis is possible in male patients, diagnosis in females requires DNA mutation analysis [65]. Over fifty different mutations in the LAMP-2 gene have been identified [19, 66].

4.1.1Clinical presentation

GSD type III is a phenotypically heterogeneous disorder with a wide clinical spectrum. While patients with GSD type IIIb mainly present with hepatic findings, affected individuals with type IIIa have both liver and muscle involvement. For both IIIa and IIIb, liver disease predominates in infancy and early childhood including hepatomegaly, hypoglycemia, hyperlipidemia, and growth retardation. For those with type IIIa GSD, there is additional variable skeletal and/or cardiac myopathy which often, though not always, may be detected via elevated serum CK concentrations once children are ambulating. Mild hypotonia and delayed motor development are usually the only manifestation during early childhood. By late childhood and adolescence, decreased stamina and pain with exertion can be noted. Muscle wasting is slowly progressive in adulthood and may be severe by the 3rd or 4th decade of life [70]. Although ventricular hypertrophy is a frequent finding, symptomatic cardiomyopathy leading to death is relatively rare. Unlike muscle disease which is a progressive process, the hypertrophic cardiomyopathy is reversible and appears to be due to excessive storage of glycogen. With a diet restricting intake of simple sugars, the hypertrophic cardiomyopathy can resolve and cardiac function normalize [71, 72].

Childhood hepatic symptoms tend to become milder with age. Complications aside from the myopathy are rare. A minority of patients (4–10%) develop hepatic adenomas [73]. Cirrhosis can also develop in patients with GSD III, and rare cases of hepatocellular carcinoma have been reported [73, 74]. Unlike in GSD Ia, hepatocellular carcinoma can develop anywhere in the liver, and it is not the result of malignant transformation of a hepatic adenoma [23]. Although all individuals with GSD type III show liver involvement, in rare instances the hepatic symptoms are mild and the diagnosis is not made until adulthood when individuals show signs of neuromuscular disease. Other clinical findings include abnormal nerve conduction studies and osteoporosis. Successful pregnancies have been reported.

4.1.2Genetics

GSD Types IIIa and IIIb are autosomal recessive allelic disorders caused by mutations in the AGL gene on the short arm of chromosome 1 [75]. The incidence of GSD III is estimated to be 1 in 100,000 live births, but high risk populations have been identified. Unusually high rates of GSD IIIa occur in the First Nation of Canada and on the Faroe Islands where there are carrier frequencies of 1 : 18 and 1 : 22, respectively [76, 77]. GSD IIIa is also more common on the Indian subcontinent (India, Pakistan, Afghanistan). Type IIIb accounts for 15% of all GSD III, and it is unusually frequent in Jews of Northern Africa (carrier frequency of 1 : 35) [66, 78]. To date, at least 140 different pathogenic AGL mutations have been reported [19].

4.1.3Pathology

The encoded enzyme, glycogen debranching enzyme (GDE), together with glycogen phosphorylase, is responsible for the complete degradation of glycogen. GDE has a presumed glycogen binding site at the carboxy terminal end, as well as two separate sites responsible for independent catalytic activities. These activities include 4-α-glucanotransferase activity (1,4-α-D-glucan:1,4-α-D-glucan 4-α-D glycosyltransferase activity) responsible for the transfer of three glucose units to the outer end of an adjacent chain, and an amylo-1,6-glucosidase activity responsible for hydrolysis of branch point glucose residues.

The variable phenotype seen in GSD type III is partly explained by differences in tissue-specific expression. When the enzyme is deficient in both liver and muscle, GSD type IIIa results; in contrast, when AGL is deficient only in the liver and enzyme activity is retained in muscle, then GSD type IIIb results. Rare cases have also been reported where only one of two GDE catalytic activities is lost [79–81]. When there is loss of only glucosidase activity, the patient is classified as having GSD Type IIIc, and when there is only loss of transferase activity, the patient is classified as having GSD type IIId.

4.1.4Treatment

While glycogenolysis is impaired in GSD III, gluconeogenesis is intact allowing lactate, amino acids, and glycerol (from fatty acid oxidation) to be used to maintain blood glucose concentrations. Protein is used as the primary source of energy in GSD type III since it also can be used directly by the muscles and has been associated with improvement in the myopathy. A diet with 3-4 grams/kg of protein is recommended through dietary protein and supplements [69]. This is supplemented with cornstarch therapy which is dosed to maintain blood glucose concentrations above 70 mg/dL and beta-OH-butyrate concentrations less than 0.3 mmol/L. The frequency of cornstarch doses varies with age. In infancy, frequent cornstarch administration may be required with therapy similar to that used in GSD type I. With older children and adults, cornstarch frequently is only required 2-3 times per day, and sometimes it is only administered prior to bedtime. For patients with moderate to severe hypertrophic cardiomyopathy, a high-protein nocturnal enteral therapy may be beneficial. Intake of simple sugars is limited to 5 grams per meal to minimize postprandial hyperinsulinemia and avoid over-storage of glycogen.

4.2.1Clinical presentation

GSD type IV shows significant variability in terms of age of onset and extent of organ and tissue involvement [82–85]. In its common classic form, patients have failure to thrive and hepatosplenomegaly. Portal hypertension and ascites develop, and progressive cirrhosis often occurs in early childhood. Without a liver transplant, death usually occurs by five years of age. Unlike the other liver forms of GSD, hypoglycemia is a late manifestation of GSD IV.

Neuromuscular forms of GSD type IV are quite variable and may be classified into several different phenotypes; interestingly, they represent the most severe and the most mild forms of GSD type IV. The most severe and relatively rare form of GSD type IV presents perinatally as fetal akinesia deformation sequence with arthrogryposis, hydrops, polyhydramnios, and pulmonary hypoplasia. In this form of the disease, death occurs at an early age due to cardiac or pulmonary insufficiency. Other severe forms of neuromuscular GSD type IV present congenitally or in early infancy with hypotonia and skeletal muscle atrophy. Prognosis varies for these forms of the disease, usually depending on the extent of cardiac and hepatic involvement. Finally, in its milder forms, GSD type IV may present in late childhood, adolescence, or even adulthood as myopathy or adult polyglucosan body disease (APBD) with central and peripheral nervous system dysfunction [85]. APBD is an allelic variant of GSD Type IV characterized by adult-onset progressive neurogenic bladder, gait difficulties due to spasticity and weakness, distal lower extremity sensory loss, and mild cognitive difficulties (OMIM 263570) [86].

4.2.2Genetics

GSD type IV is the result of a deficiency of glycogen branching enzyme which is encoded by the GBE1 gene located on chromosome 3p12.3. This gene is the only gene known to be associated with GSD type IV. Deficiency or absence of the encoded enzyme leads to excessive deposition of abnormally-structured, amylopectin-like glycogen in affected tissues. Because the accumulated glycogen lacks multiple branch points, it has poor solubility and causes irreversible tissue and organ damage.

Residual enzyme activity may confound the results of enzyme analysis; therefore, mutation analysis is often recommended to confirm the diagnosis. Thus far, at least thirty-nine different mutations have been reported across the entire length of the gene, including nonsense, missense, splice site changes, micro insertions and deletions, and several gross deletions spanning multiple exons [19]. At least ten different mutations have been reported to involve exon 12, and therefore this exon is considered a mutation “hotspot” [85, 87].

At present, there does not appear to be a strong genotype-phenotype correlation, and patients with the same mutation may show a wide range of clinical severity. In general, patients with two missense mutations have a milder form of disease than individuals with two null mutations. There is one GBE1 exon 7 missense variant that is predicted to result in the amino acid substitution p.Tyr329Ser. This founder mutation, which decreases branching enzyme activity to approximately 50% of normal, has been documented to cause Adult Polyglucosan Body Disease and is commonly found in the homozygous state in individuals with Ashkenazi Jewish ancestry [88, 89]. It should be noted that there is at least one case report of an individual with APBD who was a manifesting heterozygote with 48% branching enzyme activity [90].

4.2.3Pathology

While the hepatic scarring is the most severe of the glycogenoses, hepatic transaminase elevation is variable. Hepatic dysfunction occurs as the disease progresses. Creatine kinase levels range from normal to very elevated, and electromyography may show diffuse fibrillations.

GSD type IV is characterized by amylopectinosis. Histologic examination of liver tissue reveals periodic acid-Schiff (PAS)-positive, diastase-resistant intracytoplasmic inclusions consistent with abnormal glycogen. Characteristic findings in hematoxylin and eosin stained liver tissue include distorted hepatic architecture with diffuse interstitial fibrosis and wide fibrous septa surrounding micronodular areas of parenchyma. Hepatocytes are generally two to three times normal size with basophilic cytoplasmic inclusions. Electron microscopy of affected tissue reveals normal glycogen particles plus abnormal fibrillary aggregates typical of amylopectin (polyglucosan bodies). Muscle fibers from affected patients demonstrate severe depletion of myofibrils, and there may be amyloplasia with total fatty replacement of skeletal muscle. Polyglucosan bodies are invariably seen which are resistant to diastase digestion. In contrast to classical GSD type IV, the pathologic hallmark of adult polyglucosan body disease is the widespread accumulation of round, intracellular polyglucosan bodies throughout the nervous system, which are confined to neuronal and astrocytic processes [89].

4.2.4Treatment

Treatment of GSD IV is typically supportive. A high protein diet may have some benefit, but it has not prevented progression of the liver disease. Cornstarch is beneficial if hypoglycemia is occurring, but it similarly does not change the natural history of the disease. Due to the poor prognosis, liver transplantation remains the primary treatment for the child with early-onset, classic hepatic presentation. Individuals with adult-onset APBD may require antispasmodic bladder medications or bladder catheterization. Gait assist devices may also help to minimize the risk of falls [86].

5.1.1Clinical presentation

The clinical severity of McArdle disease is highly variable. Virtually all people with GSD V describe lifelong exercise intolerance, but the diagnosis is not usually made until the second to third decade of life when cramping becomes more prominent. Patients present with exercise-induced fatigue, painful muscle cramps, myalgia, and myoglobinuria. Approximately 90% of patients have elevated serum creatine kinase levels, even at rest. Diagnosis can be made by demonstration of failure of venous lactate to rise with an ischemic forearm test or following exercise. Electromyography does not demonstrate specific abnormalities.

In 2003, Vissing and Haller published a diagnostic test for McArdle disease based on moderate cycle exercise [92]. The authors noted that in contrast to patients with other metabolic myopathies, McArdle disease patients show decreased heart rate 7 to 15 minutes into moderate, constant-workload aerobic activity. This “second wind” phenomenon is a defining feature of McArdle disease and is thought to be due to the body’s ability to utilize alternative energy sources, including fatty acids, hepatic glycogen, and protein.

5.1.2Genetics

McArdle disease results from a deficiency of muscle-expressed glycogen phosphorylase, or myophosphorylase [93, 94]. Myophosphorylase is encoded by the PYGM gene on the long arm of chromosome 11 [95, 96]. The gene is comprised of 20 exons, and over 140 mutations have been described [19, 97–99]. Two mutations, Arg50Stop (R50X) in exon 1 (which also has been commonly reported in the research literature as R49X) and Gly205Ser in exon 5 are common mutations in patients with European heritage [91, 100]. Although no strict genotype-phenotype correlations have been made, there have been reports of more severe phenotypes in patients homozygous for both R50X mutations in PYGM and Q12X mutations in the AMPD1 gene encoding muscle adenylate deaminase [101, 102]. Cases of muscle symptoms in heterozygous carriers have been reported [103].

5.1.3Pathology

Enzyme studies on muscle biopsy will reveal absence of myophosphorylase in muscle fibers. Microscopy may also reveal acid-Schiff stained glycogen.

5.1.4Treatment

Because the metabolic block in McArdle disease impairs glycogen breakdown but glucose utilization remains intact, patients with GSD type V benefit from glucose or sucrose loading before exercise [104, 105]. Intense exercise should be avoided as it can lead to rhabdomyolysis with concomitant myoglobinuria and renal failure. Statin usage is contraindicated, and it should be noted that even heterozygous carriers may show adverse side effects to these medications [106]. Oral vitamin B₆ has been reported to impart greater resistance to fatigue, and a high protein diet may also help [107].

5.2.1Clinical presentation

Patients present in infancy or early childhood with varying degrees of growth retardation and prominent hepatomegaly secondary to excessive liver glycogen. Ketotic hypoglycemia or just hyperketosis occur with prolonged fasting or strenuous exercise [110]. Because gluconeogenesis is preserved, hypoglycemia tends to be mild. Hypotonia may lead to delayed motor development even though there is no intrinsic muscle involvement. Mild hyperlipidemia is common, and liver function tests may reveal elevated serum transaminases. Unlike other types of GSD, lactic acid and uric acid concentrations are normal. While patients with GSD VI have a milder course with few complications, treatment improves growth, stamina, and normalizes the biochemical abnormalities. Rarely, liver fibrosis develops in GSD VI, and a cardiomyopathy can occur from over storage of carbohydrate [111]. Most adults are asymptomatic, but adult females may experience hypoglycemia during pregnancy or with alcohol consumption.

5.2.2Genetics

Liver glycogen phosphorylase catalyzes the phosphorylysis of an α-1,4-glycosidic bond in glycogen to yield glucose-1-phosphate that helps meet the body’s energy needs. The enzyme exists as a homodimer of the PYGL protein and requires pyridoxal phosphate (PLP) as a cofactor. The enzyme switches between an active conformation (GPa) and an inactive conformation (GPb), with activation dependent upon phosphorylation of a serine located at amino acid position 14. Such phosphorylation occurs in response to the hormones glucagon and epinephrine.

GSD type VI is inherited in an autosomal recessive fashion. The PYGL gene on chromosome 14 spans over 39,000 base pairs, consists of 20 coding exons, and encodes a protein that is 846 amino acids in length [112]. Thus far, thirty disease-causing mutations have been reported [19, 113–115]. The vast majority of pathogenic variants are missense mutations [116]. No affected individuals have been described with two null alleles, suggesting that complete absence of liver glycogen phosphorylase activity may be incompatible with life. The estimated disease incidence ranges from 1 in 65,000 to 1 in 85,000 births, but many people with this condition are undiagnosed. In the Mennonite community, however, there is a founder mutation (c.1620+1G>A, at the intron 13 splice donor site), and the prevalence is approximately 0.1% (1 in 1,000 individuals) [112].

5.2.3Pathology

Although hepatic glycogen phosphorylase enzyme is expressed in several cell types and its activity can be assayed using erythrocytes, leukocytes, or hepatocytes, such testing is neither highly sensitive nor specific. False negative results are common because enzyme activity is significantly reduced in Hers disease but is never completely absent. False positive results also are not rare, because reduced liver phosphorylase activity may be due to mutations in the PYGL gene or mutations in several other genes (including PHKA2, PHKB, and PHKG2) that encode phosphorylase b kinase, the activating enzyme for hepatic glycogen phosphorylase. As a result, mutation detection is now the preferred method to differentiate liver phosphorylase deficiency from the much more common deficiency of the phsophorylase b kinase activating enzyme. Liver biopsies demonstrate glycogen filled hepatocytes with or without fibrosis, but DNA analysis or enzymatic testing is needed to differentiate GSD VI from the other forms of GSD.

5.2.4Treatment

Affected individuals should avoid prolonged fasting, and eat frequent small meals. Uncooked cornstarch (1–4 times per day) and protein supplementation may help stabilize blood glucose levels and prevent complications such as short stature, delayed puberty, and osteoporosis. Protein supplementation typically is lower than in GSD III (2–2.5 g/kg/day) with a goal of normalizing prealbumin concentrations. Rarely, cirrhosis and hepatocellular carcinoma can occur in Hers disease [111, 117]. Patients should avoid excessive amounts of simple sugars. In addition, growth hormone therapy should not be used to treat short stature since it will lead to increased ketone production. To assess metabolic control, blood glucose levels and blood ketones should be routinely monitored. Height and weight measurements should also be assessed regularly since growth is normal when treatment is optimized.

5.3.1Clinical presentation

GSD type IX has the most heterogeneous clinical picture of all of the glycogen storage diseases. Most patients are diagnosed after hepatomegaly is incidentally found, and it is the most common identifiable cause of ketotic hypoglycemia in males [118]. While most patients are relatively mild, a severe variant exists that mimics type I GSD in infancy with severe fasting hypoglycemia. Patients with GSD type IX may present with hepatic and/or muscle disease, and many patients suffer from a combination of the two. There is at least one form of GSD type IX which is strictly muscle-specific, and affected patients may present with muscle pain and weakness, exercise intolerance, and myoglobinuria. Another form of GSD type IX strictly presents as hepatic disease that typically begins in the first few months of life, and affected individuals may have ketotic hypoglycemia, hepatomegaly due to elevated glycogen content, liver disease, growth retardation, hypotonia, abnormal lipid profile, and increased lactate and uric acid. In its mildest hepatic form, patients may have a phenotype similar to GSD Type VI and symptoms may gradually subside with age. In patients with hepatic transaminase elevation, liver complications can develop including fibrosis, cirrhosis, adenomas, and hepatocellular carcinoma [111].

5.3.2Genetics

Glycogen storage disease type IX is a genetically heterogeneous disorder. The phosphorylase kinase (Phk) enzyme is a hexadecameric structure comprised of four copies each of four different polypeptides, including alpha (α), beta (β), gamma (γ), and delta (δ) subunits [119]. To add to the molecular complexity, various tissue-specific isoforms exist for each subunit; these isoforms may be due to expression from separate genes or from alternative splicing of a single gene. α-associated GSD Type IX may result from mutations in one of two X-linked genes: PHKA1 or PHKA2. PHKA1 is located on the long arm of chromosome X at Xq13 while PHKA2 is located on the short arm of the X chromosome at Xp22.13 [120]. PHKA1 expression is confined to muscle, and therefore PHKA1 mutations are associated with exercise intolerance, muscle pain, weakness, and myoglobinuria [121–123]. To date, there are only seven reported mutations in the PHKA1 gene [19]. In contrast, PHKA2 gene expression is confined to liver and blood cells, and patients with PHKA2 mutations strictly have a hepatic presentation with ketotic hypoglycemia, hepatomegaly, chronic liver disease, retarded growth and motor development, and elevated lipids [124–128].

In contrast to the α-subunit, there is only one gene known to encode the β-subunit of the Phk enzyme. This gene, PHKB, is located on the long arm of chromosome 16 at 16q12.1 [120]. Alternative splicing of several exons gives rise to tissue-specific transcripts, and PHKB mutations have been associated with phosphorylase kinase enzyme deficiency in both liver and muscle [129–133]. Most mutations identified in PHKB have been severe null mutations expected to lead to premature protein truncation or mRNA decay; nevertheless, patients generally have mild symptoms including hypoglycemia after prolonged fasting, hepatomegaly, and mild hypotonia [19]. Cirrhosis and other major complications have not been reported in patients with PHKB mutations to date. It is important to note that PHKB mutations have not been found in patients with only muscle disease [134].

The PHKG2 gene on chromosome 16 encodes the liver- and testis-specific form of the γ-subunit [135]. This subunit contains the phosphorylase kinase enzyme’s active site responsible for the phosphorylation and activation of glycogen phosphorylase [136]. PHKG2-linked disease is associated with a more severe phenotype, which may include fasting hypoglycemia, impaired glucagon response, muscle weakness and fatigue, hepatomegaly, liver fibrosis and cirrhosis [133, 137–139].

5.3.3Pathology

Since hepatomegaly is often the presenting symptom, the diagnosis of GSD IX is still commonly made by liver biopsy. As with the other forms of GSD, glycogen filled hepatocytes with prominent steatosis is seen, but fibrosis is usually present in GSD IX. Clinical diagnostic laboratories in the United States require samples from affected tissues (i.e., liver and/or muscle) for enzyme studies. However, non-invasive analysis of phosphorylase kinase enzyme has been reported using blood cells [133]. Although biochemical testing may be used to provide a GSD type IX diagnosis, enzyme analysis cannot determine which gene is causing disease. Mutation analysis is therefore recommended in individuals suspected to have GSD type IX. Mutations in PHKA2 cause GSD IX in over 80% of males with the disease. Sequencing of PHKB should be considered first in females.

5.3.4Treatment

Although GSD type IX was once considered a benign condition, it is now clear that patients may experience more long-term complications. Patients who have elevated hepatic transaminases and post-prandial hyperlactatemia are particularly at risk for development of cirrhosis, and aggressive management is imperative [140]. In all patients with GSD IX, treatment improves growth, stamina, and normalizes biochemical tests.

Affected individuals should restrict intake of simple sugars and eat frequent small meals. A high protein diet is used (2-3 g/kg), and uncooked cornstarch supplementation is used to maintain glucose concentrations and avoid ketosis. While most patients with GSD type IX can make it through the night with cornstarch and protein, overnight feeds are sometimes needed in patients with mutations in PHKA2 and PHKG2. For these patients, the extended release cornstarch preparation (Glycosade) can be considered [141]. Growth hormone therapy should not be used to treat short stature since it will lead to increased ketone production. Liver ultrasound examinations are recommended beginning in childhood since patients are at risk for developing hepatic adenomata and cirrhosis.