Supplementary material
Supplement to: Staufner C et al, Adenosine kinase deficiency: expanding the clinical spectrum and evaluating therapeutic options, Journal of Inherited Metabolic Disease 2015.
I. Case reports
II. Supplementary tables and figure
Table S1. Anthropometrical data at birth and at last assessment
Table S2. Biochemical response to low methionine diet in patient 7
Figure S1. Impact of low methionine diet on liver function and methionine levels
I. Case reports
Case report family I (patient 1 and 2)
Family I is a consanguineous Turkish family with four children; patient 1 is the oldest and patient 2 the youngest child, the other two children are healthy.
Patient 1 is a girl delivered at 36+4 weeks of gestation by caesarean section due to pathological CTG. Birth weight was normal for gestational age. Cyanosis was noted at 4 hrs post-delivery with hypoglycemia (blood glucose not detectable) and hypothermia (35.1 °C). Glucose infusion of up to 16mg/kg/min was needed to stabilize blood glucose levels and insulin was high during hypoglycemia (blood glucose 1.67 mmol/L, insulin 30.9 mU/L), so hyperinsulinism was diagnosed. From the second day of life she received phototherapy due to hyperbilirubinemia. There was mild liver dysfunction with an increase of transaminases and INR but no cholestasis. Hyperinsulinism was responsive to diazoxide. Because of mild hypothyroidism, supplementation with L-thyroxine was started. Methionine concentrations were mildly and intermittently elevated (max. 107 µmol/L), whereas concentrations of AdoMet and AdoHcy were markedly and consistently elevated in plasma (AdoMet 355 nmol/L; AdoHcy 224 nmol/L) and CSF (AdoMet 384 nmol/L; AdoHcy 218 nmol/L). Recurrent mild to moderate hyperammonemia (80-140 µmol/L) was observed but could not be explained. Echocardiography revealed a perimembranous ventricular septum defect (VSD), persistent ductus arteriosus (PDA) and persistent foramen ovale (PFO). Because of hemodynamic decompensation, cardiosurgical correction of the defects became necessary at the age of 4 months. By that time patient 1 showed hepatomegaly, severe muscular hypotonia without head control, severe global developmental delay and frontal bossing. Epilepsy developed at the age of 5 months. Liver function had nearly normalized but treatment with diazoxid was needed to control hyperinsulinism. At the age of 7 months, liver biopsy showed periportal and intralobular fibrosis with low grade parenchymal fat and no signs of cholestasis. At the age of 15 months, patient 1 died due to a pulmonary infection.
Patient 2 (male) was born at term as the 4th child of family I with normal birth weight. On his first day of life, he presented an episode of cyanosis and bradycardia and hypoglycemia of 0.56 mmol/L was detected. Infusion of glucose 10 mg/kg/min was needed to stabilize blood sugar levels and hyperinsulinism was detected in the further workup (blood glucose 1.17 mmol/L; insulin 4.3 mU/L), responsive to diazoxide. The patient showed muscular hypotonia and hepatomegaly. From day 3 on, cholestatic hyperbilirubinaemia, increased transaminases and severe liver dysfunction presented (see table 2 for maximum values). Hypermethioninemia was detected (methionine 176 µmol/L at day 6; 400 µmol/L at day 34). A low methionine diet was initiated, but was discontinued after 5 days because of lacking clinical and biochemical response. Notably, methionine concentrations spontaneously normalized in the course of the disease. Echocardiography revealed a persistent arterial duct (PDA) and an atrial septum defect (ASD), which both resolved spontaneously. Liver biopsy at the age of 1 month showed severe portal, septal and perisinoidal fibrosis with transition to cirrhotic rearrangement with pronounced ductal reaction and incipient sings of chronic cholestasis, and hardly viewable extra hepatic bile ducts. Liver function recovered under symptomatic treatment, but there was neurological impairment with muscular hypotonia and global retardation. Clinical seizures were first observed at the age of 4 months, but EEG has already been highly pathologic during the newborn period. At the age of 2.5 years, frontal bossing was noted. Treatment with diazoxide was stopped at the age of 2.7 years, but single hypoglycemic episodes were still noted in the further course.
At time of report, the patient is 7 years old. He is a happy and cooperative boy, but there is severe global developmental retardation; he has a hypotonic-dystonic movement disorder with changing muscle tone. He has no speech but communicates with sounds and gesticulation. His developmental age is 2-4 months (according to DENVER testing). He is wheel chair bound and fed through a percutaneous gastrostomy. He is nearly blind because of retinal dystrophy. He is free of seizures on therapy with lamotrigin and phenobarbital. Liver function is normal. Orchidopexia was performed due to maldescensus testis on both sides. Methionine levels are normal but plasma AdoMet and AdoHcy and urine adenosine are increased.
After ADK deficiency had been published, genetic testing of the ADK gene was performed and revealed a homozygous missense mutation (c.905 T>C, p.Phe302Ser). The same mutations could be confirmed in DNA from fibroblasts of patient 1.
Case report family II (patient 3)
Family II is a consanguineous Turkish family with 7 children and is related to family I, as the father of patient 1 and 2 is brother of patient 3. Patient 3 is a girl born at term who presented on her first day of life with severe hypoglycemia (0.39 mmol/L). Blood glucose stabilized quickly after glucose infusion. On her 2nd day of life she developed hyperbilirubinaemia (total bilirubin 345.3 µmol/L, direct bilirubin 6.8 µmol/L) that remained on the same level till the 8th day of life although intensive phototherapy was applied. Transaminases and gamma-GT were mildly increased (ASAT 85 U/L, gamma-GT 441 U/L), and coagulopathy (INR max. 2) developed within the first week of life. During the neonatal period, truncal muscular hypotonia became evident; eye fixation was inconsistent and social contact not appropriate for age. Partial tube feeding was required. Echocardiography revealed persistent ductus arteriosus, persistant foramen ovale and neonatal coarctation of the aorta, which resolved spontaneously during follow-up. Newborn screening from dried blood spot (DBS) revealed hypermethioninaemia (138 µmol/L, N < 91; day 4 of life), but on day 11 methionine had become normal in DBS (70 µmol/L). Neither amino acid analysis in plasma nor further metabolic work-up were investigated at this stage. Due to cholestatic hyperbilirubinaemia (total bilirubin 230.8 µmol/L, direct bilirubin 70.1 µmol/L) and increased ASAT, ALAT and gamma-GT at the age of 2 months (ASAT 240 U/L and GGT 219 U/L) liver biopsy was performed, showing predominantly lobular hepatitis and a reduced number of bile ducts. Frontal bossing was noticed at 3 months of age.
In the further course recurrent asymptomatic hypoglycemia (min 1.22 mmol/L) was noted. In one hypoglyacemic episode ketone bodies were low but insulin was not measured at this time. In an earlier hypoglycemic episode insulin was not detectable. At the age of 5 months, there was a continuous cholestatic icterus and hepatomegaly with increased transaminases but near to normal coagulation (total bilirubin 365.8 µmol/L, direct bilirubin 265.0 µmol/L, ASAT 235 U/L, INR 1.2). Metabolic work up was then initiated and revealed massive hypermethioninaemia (methionine 809 µmol/L) with clearly elevated AdoMet (2299 nmol/l) and moderately increased AdoHcy (187 nmol/L) in plasma. Betaine (trimethylglycine) (995 µmol/L; N 18-41), dimethylglycine (54.6 µmol/L; N 1.8-3.7) and sarcosine/ monomethylglycine (6 µmol/L; N not detectable) were also increased in plasma.
At the age of 7 months liver function had further deteriorated (see maximum values in table 2) and hepatorenal syndrome with metabolic acidosis and ascites developed. Low methionine diet was started as experimental therapy with a mean daily intake of 15 mg methionine per kg of body weight. As neither laboratory results nor the clinical situation improved within the first week, the parents decided to take her daughter home but to continue the low methionine diet in an overall setting which was believed to be palliative. In the further months, liver function stabilized and improved until about one year later, liver function had normalized nearly completely (see figure S1). Also concentrations of methionine, AdoHcy, betaine and DMG had normalized whereas concentrations of AdoMet and sarcosine were still above the normal range but had decreased (AdoMet 164 nmol/L, sarcosine 4 µmol/L).
At the age of 1.9 years the low methionine diet was stopped but successfully reintroduced one month later, as liver impairment and hypermethioninaemia redeveloped. However, the patient showed severe neurological impairment with pronounced psychomotor retardation. Epilepsy developed at the age of 4.8 years. The low methionine diet was stopped again at the age of 4.9 years and liver function remained stable since then, but there were two episodes with slight increase of transaminases, total bilirubin and methionine with spontaneous normalization within a few days; one of these episodes was triggered by a viral infection. At time of report the patient is 11.75 years old. She has moderate to severe muscular hypotonia and dystonia. She had learned to crawl and was able to walk a few steps without support but has lost these abilities during the last year. She has no speech but communicates on a low level with sounds and gesticulation. Her developmental age is 9-11 months (according to DENVER testing). Apart from frontal bossing and abnormal dentition, slender hands and feet are noticeable. Anticonvulsive treatment has become increasingly challenging and the patient has daily seizures despite a combination therapy with topiramat, levetiracetam and lamotrigin.
Patient 3 shares the same homozygous missense mutation as her cousins patient 1 and 2 (c.905 T>C, p.Phe302Ser).
Case report family III (patient 4 and 5)
Family III is a consanguineous Arabic family from Kuwait with 3 children, two of them affected (patient 4 and 5).
Patient 4 was born at term as 1st child of family III with a birth weight of 2300g. She had phototherapy for 2 days to treat hyperbilirubinemia and was discharged home in stable condition. She was re-admitted to hospital at the 12th day of life with severe hyperbilirubinaemia (total bilirubin 427.4 µmol/L, direct bilirubin 128.2 µmol/L), raised transaminases and muscular hypotonia. The patient had recurrent hypoglycemia since infancy, but insulin levels were not checked during hypoglycemia episodes. Due to elevated levels of tyrosine (and methionine), tyrosinaemia type I was suspected although urine succinylacetone was negative and feeding with a Phe-Meth-Tyr free formula was initiated. Repeated blood amino acid examinations showed no significant abnormalities under diet. At the age of 1.2 years liver biopsy had revealed cholestasis with microvesicular fatty degeneration and periportal fibrosis. Ultrasound showed cholelithiasis. Dietary treatment was stopped at that time, after tyrosinaemia type I had been considered very unlikely. Psychomotor development was retarded, but she learned to sit at the age of 3 years and to stand at the age of 5 years. From 3 years of age she was able to speak single words. Epilepsy was diagnosed at the age of 8 years. At the age of 14 years, ADK deficiency was suspected because of an isolated elevation of methionine with a moderate increase of AdoMet and AdoHcy in plasma and elevated adenosine concentration in urine (see table 2 for laboratory values). Molecular genetics revealed a homozygous deletion in exon 3 of the ADK gene (p.30-47del). The patient can walk independently, gait is broad-based and she speaks single words. There is an increased muscular tone of the lower extremities. She is of short stature, shows frontal bossing and has sparse hair. Furthermore, downward slanted palpebral fissures, hypertelorism, a broad nose, low-set ears, and severely distorted teeth are noticeable. Seizures are well controlled with lamotrigin monotherapy. After prolonged fasting she develops episodes of hypoglycemia.
Patient 5 was born at term as the 3rd child of family III with a birth weight of 2900g. Due to neonatal hyperbilirubinaemia, she received phototherapy for 10 days. There was muscular hypotonia from birth on. Seizures started at 6 months of age. Recurrent hypoglycemia was first noted when she was 2 years old. It is not reported whether insulin levels were measured. Psychomotor development has been severely retarded. At the age of 5 years, she started to crawl and was able to speak very few single words. ADK deficiency was suspected at the age of 8.3 years, when elevated concentrations of methionine, AdoMet and AdoHcy (all in plasma) and adenosine (urine) were measured (see table 2 for laboratory values). At that time patient 5 was short of stature, showed frontal bossing, hypertelorism and downward slanted palpebral fissures. She can pull herself up, but there is no standing or walking. There is an increased muscular tonus and coarse tremor of the extremities and she shows dystonic movements. She has lost her skill of speaking a few words, uses no speech anymore and has no understanding of words. Electrophysiology shows signs of a mild demyelination polyneuropathy. At the age of 9 years, she had a gall bladder removal because of symptomatic cholelithiasis. She is free of seizures with lamotrigin and levetiracetam. She shares the same homozygous deletion as her older sister.
Case report family IV (patient 6 and his three sisters, published by Labrune et al. in 1990)
Family IV is a consanguineous Moroccan family with 9 children. The eldest, born in 1978, died at the age of 9 months due to an unknown cause. Three girls of this family were previously reported to have suffered from a syndrome of hypermethioninaemia, liver dysfunction, recurrent hypoglycemia, neurological impairment and facial dysmorphism including a prominent forehead, sparse hair and abnormal teeth. All had become symptomatic on their first day of life. Plasma concentrations of AdoMet and AdoHcy were not measured, but in urine, concentrations of AdoMet were high. A beneficial effect had been observed when a low methionine diet was applied. The cases have been published by Labrune et al. in 1990 as “familial hypermethioninemia partially responsive to dietary restriction” (Labrune, et al. 1990). One of the siblings died at the age of 19 years due to respiratory distress in the course of an influenza infection, the others are alive and 34 and 28 years old at time of report. Both have a severe neurological phenotype with pronounced mental disability: one presenting very severe behavioral disturbances (such as coprophagia), severe epilepsy and schizencephaly on brain MRI (frontal right). She has an asymptomatic cholelithiasis. The other patient is bedbound. Due to the genetically confirmed diagnosis of their brother these siblings can be classified as ADK deficiency.