D. Leupold

Atypical phenylketonuria with defective biopterin metabolism. Monotherapy with tetrahydrobiopterin or sepiapterin, screening und study of biosynthesis in man

Administration of a single dose of tetrahydrobiopterin dihydrochloride, 10–20 mg/kg orally, to a patient with dihydrobiopterin deficiency led to disappearance of clinical symptoms for 4 days, normalization of urinary phenylalanine and serotonin and decrease of elevated neopterin for 2–3 days. A dose-dependent stimulation of serotonin production was observed. A similar effect was note with even lower doses of L-sepiapterin. The patient is now under monotherapy with tetrahydrobiopterin·2 HCl, 2.5 mg/kg daily. Other patients with this disease may not respond as well.Results of screening for tetrahydrobiopterin deficiency in 228 cases with hyperphenylalaninemia, including 140 newborns, are reported.There is evidence that biopterin biosynthesis in human kidney and liver proceeds via a dioxo compound and L-sepiapterin.

Atypical phenylketonuria with “dihydrobiopterin synthetase” deficiency: Absence of phosphate-eliminating enzyme activity demonstrated in liver

An assay for the phosphate-eliminating enzyme (PEE) activity in liver was developed which required only 5–10 mg tissue. PEE catalyses the elimination of inorganic triphosphate from dihydroneopterin triphosphate, which is the second and irreversible step in the biosynthesis of tetrahydrobiopterin (BH4). In the presence of substrate, magnesium, NADPH, and a sepiapterin reductase fraction from human liver, PEE catalysed the formation of BH4 which was measured by HPLC and electrochemical detection. In adult human liver, a PEE activity of 1.02±0.134 μU/mg protein (mean ±1 SD; n=5) was observed. In liver needle biopsy material from five patients with defective biopterin biosynthesis, no PEE activity was found (less than 2% and 6% of the control values, respectively). The presence of an endogenous inhibitor was excluded. In a patient who died without definite diagnosis and in a patient with β-thalassaemia liver PEE activity was increased. Sepiapterin reductase activity was present in all cases. Results indicate that in “dihydrobiopterin synthetase” deficiency, the most frequent of the rare BH4-deficient variants of hyperphenylalaninaemia, the molecular defect consists in a defect of PEE.

Molecular analysis and long-term follow-up of patients with different forms of 6-pyruvoyl-tetrahydropterin synthase deficiency

Abstract The outcome of 6-pyruvoyl-tetrahydropterin synthase (PTPS) deficiency, the most common form of tetrahydrobiopterin (BH4) deficiency, depends on factors such as severity of the disease, type of mutation, time of diagnosis, and mode of treatment. We investigated five patients from four different families, four of them presenting with the severe form of PTPS deficiency and one with the mild peripheral form. In this study, missense (L26F, T67M, P87L, V124L, D136G, D136V) and nonsense (R15–16ins) mutations were detected by reverse transcriptase polymerase chain reaction and sequence analysis. Two patients with the severe form were compound heterozygotes (T67M/P87L and D136G/R15–16ins), two siblings were homozygous for the D136V mutation, and in the patient with the mild form, heterozygous L26F/V124L mutations were present. Two patients are on combined therapy with l-dopa/carbidopa/5-hydroxytryptophan plus BH4, the siblings are on monotherapy with BH4, and the patient with the mild form is now off treatment, presenting with normal plasma phenylalanine levels. Conclusion Long-term follow-up shows that the outcome of 6-pyruvoyl-tetrahydrop- terin synthase deficiency benefits from treatment started in the first months of life and that the phenotype may change with age. Additionally, depending on the type of mutations, prenatal damage to the fetus may multiply the clinical abnormalities and thus worsen the prognosis of the disease. In patients initially diagnosed with the mild peripheral form of the disease, therapy with tetrahydrobiopterin should be stopped after some time to test whether hyperphenylalaninaemia was only a transient condition.

3-Hydroxy-3-methylglutaryl-CoA lyase deficiency in an infant with macrocephaly and mild metabolic acidosis

A girl, now three years old, is reported, in whom at the age of 5 months the diagnosis of 3-HMG-CoA lyase deficiency was established. The characteristic excretion pattern consisted of 3-HMG, 3-CH3-glutaconic acid, 3-CH3-glutaric acid and 3-HIVA. Activity of 3-HMG-CoA lyase in leucocytes was very low. She had compensated metabolic acidosis and mild hypoglycemia.Therapy with a leucine restricted diet decreased excretion of metabolites moderaterely but did not influence the tendency to metabolic acidosis.Clinically the infant presented with macrocephaly. At the age of 3 years she is severely retarded. CAT scan revealed the picture of progressive demyelination of the white matter.

3-Methyl-3-butenoic acid: an artefact in the urinary metabolic pattern of patients with 3-hydroxy-3-methylglutaryl-coa lyase deficiency

At present six patients with 3-hydroxy-3-methylglutaryl-CoA lyase deficiency, an inborn error of leucine metabolism have been reported [l-4 and C. Bachmann, personal communication]. Characteristic urinary organic acids in this disease are: 3-hydroxy-3-methylglutaric acid, 3-methylglutaconic acid, 3-methylglutaric acid and 3-hydroxyisovaleric acid. Endogenous accumulation of 3-methylcrotonyl-CoA leads to the formation of 3-methylcrotonylglycine [ 51. However, urinary 3-methylcrotonic acid may also be formed artificially by decarboxylation of 3-methylglutaconic acid [ 61. During the investigation of the decarboxylation of 3-methylglutaconic acid another compound was detected, probably an isomer of 3-methylcrotonic acid. In this article the identification of this compound as 3-methyl-3-butenoic acid is described. Evidence is presented that 3-methyl-3-butenoic acid is not an endogenous metabolite.

Direct identification of propionylcarnitine in propionic acidaemia: Biochemical and clinical results of oral carnitine supplementation

Urinary short-chain acylcarnitine in a patient with propionic acidaemia and low levels of free carnitine was found to consist mainly of propionylcarnitine. The compound was isolated by sequential paper and thin layer chromatography and identified by ammonia desorption chemical ionization mass spectrometry. Treatment of the patient with oral carnitine supplements led to a near-normalization of the plasma free carnitine concentrations and an increase in his muscle tone. The propionylcarnitine excretion rose and there was a simultaneous decrease in the methylcitrate output. Carnitine treatment did not prevent the occurrence of an episode of metabolic decompensation.

Prenatal Diagnosis of Atypical Phenylketonuria

Atypical phenylketonuria (PKU) is a group of very rare and severe diseases caused by tetrahydrobiopterin (BH4) deficiency (Niederwieser and Curtius, 1987). So far three inborn errors of metabolism are known to cause BH4 deficiency. defects in: GTP cyclohydrolase I (GTPCH); 6-pyruvoyl tetrahydropterin synthase (PPH4S); and dihydropteridine reductase (DHPR) (Blau, 1988). Recently a new form of atypical PKU with unusual 7-iso-biopterin excretion in the urine of patients was described (Curtius et al., 1988). Prenatal diagnosis of BH4 deficiency can be achieved mainly by measurement of pterin metabolites in amniotic fluid and of enzyme activities in cultured fluid cells and fetal blood (Blau et al., 1987).

Contribution of odd-numbered fatty acid oxidation to propionate production in neonates with methylmalonic an propionic acidaemias

Newborns with disturbed propionate metabolism (propionic and methylmalonic acidaemias) accumulate during fetal life large amounts of odd-numbered longchain fatty acids in adipose tissue (8% to 10% of total fatty acids vs 1% in normal controls) and other body lipids. During periods of acute catabolism, such as in the early days of life, mobilization and oxidation of odd-chain fatty acids from adipose tissue yield extensive amounts of toxic propionyl-CoA in the mitochondria and thus might contribute to the severe illness of the patients. Our data suggest that not only catabolism of protein but also that of adipose tissue has to be avoided in order to alleviate the neonatal illness of patients with propionic and methylmalonic acidaemias. It seems prudent to avoid prolonged fasting during the 1st year of life.

Evaluation of prenatal treatment in newborns with cobalamin-responsive methylmalonic acidaemia

Odd-numbered fatty acids with 15 an 17 carbon atoms (OLCFAs) are formed from propionyl-CoA, accumulate in subjects with methylmalonic (MMA) and propionic (PA) acidaemias and are incorporated in large quantities into lipids even during fetal life. In fetuses with cobalamin-responsive MMA, treatment with large doses of vitamin B L2 given to the mother can increase the flux through the defective methylmalonyl-CoA mutase step, resulting in decrease of OLCFAs in lipids at birth (Wendel et al 1991). We report our experience using OLCFAs in different lipids for evaluation of prenatal treatment in three fetuses with MMA. Adipose tissue was obtained by needle aspiration. The work-up of lipids and analysis of fatty acids were as described (Wendel et al 1991). The sum of the OLCFAs is expressed as a percentage of the total C14-C22 fatty acids in the sample. In pregnancy I (for details see van der Meer et al 1990) with a fetus affected with cobalamin-responsive MMA, treatment was started with vitamin B12 (22.5mg/day) orally at the 27th week. Methylmalonate excretion in maternal urine ranged from 7.0 to 10.3 mmol/mol creatinine. At birth no detectable quantities of methylmalonate were found in cord blood, and OLCFAs measured in cord plasma lipids were within the normal range. In pregnancy II with a fetus affected with cobalamin-responsive MMA, prenatal treatment was started with 4mg/week of hydroxycobalamin IM in the 21st week and changed to oral application of 20 mg/day in the 29th week. Methylmalonate excretion in maternal urine fluctuated between 2.3 and 4.0mmol/mol creatinine. OLCFAs measured in cord red cell lipids were slightly elevated as compared with normal controls (1.4% vs 1.0+0.2%); however, they were much more elevated in adipose tissue (5.6% vs 1.0+0.2%). In pregnancy III the fetus had MMA that was assumed to be cobalamin-responsive. Treatment started with 2mg/week of hydroxycobalamin IM in the 23rd week. Methylmalonate excretion in maternal urine was not effected by treatment and fluctuated between 7 and 22 mmol/mol creatinine. At birth cord blood contained 1.2mmol/L methylmalonate. The OLCFAs in cord plasma lipids (phopholipids 7.1% vs 0 .7-1 .1%; triglycerides 10.6 vs 0 .9-2 .9%, and sterol esters (10.6% vs 1.2-3.3%) and erythrocyte lipids (7.0% vs 1.0 + 0.29%) were highly elevated. In fetuses with cobalamin-responsive MMA, prenatal treatment with vitamin BI2 is highly effective in keeping the amounts of OLCFAs in the (near) normal range, in cord plasma and erythrocyte lipids but not in adipose tissue, in which OLCFAs are high as compared with healthy controls. However, they are clearly lower than in adipose tissue

Clinical features and diagnostic approach in type I tyrosinaemia in an infant with cytomegaly virus infection and bacterial sepsis

A severely ill 2-month-old female infant was admitted with meningitis and septicaemia caused byStreptococcus pneumoniae. The patient, who also had an acute cytomegalovirus (CMV) infection, revealed the typical clinical and biochemical characteristics of type I tyrosinaemia (TIT). Clinical evidence of severe hepatocellular damage was shown, but urinary succinylacetone was not detected. The diagnosis of TIT was finally confirmed by decreased activity of fumarylacetoacetase (FAA) in skin fibroblasts from the patient and both parents. Following dietary treatment and after overcoming the bacterial and viral infection, the patients liver function improved.