[Screening of tetrahydrobiopterin deficiency among hyperphenylalaninemic patients]. / Repérage des hyperphénylalaninémies par déficit en tétrahydrobioptérine.

Tetrahydrobiopterin deficiency in hyperphenylalaninemic babies has to be rapidly recognized since the disease requires a specific follow-up. Based on specimen collection on filter paper, a simple strategy for the screening of this condition has been used since 1987. Urine pteridine measurement can detect 6-pyruvoyl-tetrahydropterin synthase, GTPcyclohydrolase I and pterin-4a-carbinolamine dehydratase deficiencies and direct enzyme measurement in dried blood sample detects dihydropteridine-reductase deficiency. A total of 1,814 hyperphenylalaninemic patients have been studied and 34 tetrahydrobiopterin deficiencies have been detected. The strategy must commend itself by its convenience and simplicity, and can be use on all babies with hyperphenylalaninemia screened in the neonatal period, whatever their blood phenylalanine level.

2,4-diamino-7-hydroxy-pteridines in biological fluids of patients on high-dose methotrexate.

Concentrations of 2,4-diamino-7-hydroxy-pteridines in plasma and cerebrospinal fluid (CSF) are reported for the 0.5-8 g/m2 dose range of methotrexate in children with acute lymphoblastic leukemia or non-Hodgkin's lymphoma. This experiment has revealed that: (1) there is a wide inter-individual but relatively narrow intra-individual variability of the maximal concentrations of 2,4-diamino-7-hydroxy-pteridines in plasma during consecutive methotrexate cycles; (2) the increase in the level of the metabolites in plasma was related to increments of the methotrexate dose, but not above 5 g/m2: this can be explained by a saturable conversion of methotrexate to 2,4-diamino-7-hydroxy-pteridines; (3) significant correlations were found between simultaneous values of 2,4-diamino-7-hydroxy-pteridines in plasma and CSF; (4) the formation of 2,4-diamino-7-hydroxy-pteridines did not depend on the ages of patients receiving the same dose of methotrexate. The presence of these compounds in plasma and CSF in significant amounts creates the potential for a number of competitive interactions with pteridine-dependent metabolism which may open up new possibilities for understanding the metabolic side-effects of methotrexate therapy.

[Unconjugated pteridines and neuromeningeal infections]. / Ptéridines non conjuguées et infections neuro-méningées.

Concentrations of unconjugated pteridines (neopterin, monapterin, biopterin, pterin) were measured in the cerebrospinal fluid (CSF) of 310 patients, using a high performance liquid chromatography (HPCL) method. Our cohort included 209 controls (C), 15 patients with meningism (M), 22 with viral meningitis (VM), 17 with bacterial meningitis (BM), 9 with herpetic meningoencephalitis (HME), 2 with tuberculous meningoencephalitis (TME) and 36 with peripheral systemic infections (PI). These measurements, expressed as nmol/litre, showed a gradation of neopterin concentrations according to the type of infection: 20.1 + 6.5 in group C; 46.9 +/- 29.9 in group PI; 274.3 +/- 231.7 in group VM; 699.2 +/- 711.2 in group BM, 1,101.9 +/- 1,107.9 in group HME and 1,169 +/- 1,171.9 in group TME. There was no such gradation with biopterin. Comparisons of means showed that total concentrations in the pathology groups were very different from those observed in controls and in the neuromeningeal infections of the PI group. There was no correlation between the number of lymphocytes and the concentrations of neopterin or biopterin in the CSF. It is concluded that the concentration of neopterin in the CSF is a sensitive but little specific marker of infection, independent of CSF cellular reaction. Measuring this concentration makes it possible: 1) to evaluate the status of immune defences; 2) to predict that a meningitis will become chronic, and 3) to detect a possible parenchymal participation in a meningeal infection.

Impairment of cerebral biogenic amine synthesis in a patient receiving high-dose methotrexate.

A transient acute neurologic syndrome occurred in a 15-year-old boy receiving high-dose methotrexate (MTX) for acute lymphoblastic leukemia. Cerebrospinal fluid analysis revealed a decrease of homovanillic acid and 5-hydroxyindoleacetic acid concentrations with a rise of biopterin levels. These findings suggest that MTX can induce a transient alteration of the metabolism of tetrahydrobiopterin leading to the defect of biogenic amine neurotransmitter synthesis.

2,4-diamino-7-hydroxy-pteridines, a new class of catabolites of methotrexate.

Methotrexate remains a commonly used drug in the chemotherapy of various malignancies. The known catabolites are 7-hydroxy-methotrexate, formed in the liver, and diamino-methyl-pteroic acid formed in the gut. We report for the first time evidence that 2,4-diamino-7-hydroxy-pteridine derivatives are present in the biological fluids of patients on high-dose methotrexate protocols. So far, two major derivatives have been identified as 2,4-diamino-6-hydroxymethyl-7-hydroxy-pteridine and 2,4-diamino-6-methyl-7-hydroxy-pteridine. In regard to the actual knowledge of the catabolism of pteridines, these compounds are presumably formed by intestinal bacteria during enterohepatic circulation of the drug. Their slow clearance from the body raises the question of possible interference of these compounds on pteridine-dependent enzymes, which might explain in part some of the toxic effects of methotrexate.

[High-dose methotrexate and hyperphenylalaninemia]. / Méthotrexate à haute dose et hyperphénylalaninémie.

The occurrence of increased levels of blood phenylalanine after therapeutic administration of folate analogues has been occasionally reported and attributed to the inhibition of dihydropteridine reductase, an enzyme maintaining the cofactor of phenylalanine hydroxylase in its active tetrahydrogenated form (tetrahydrobiopterin). To study further this metabolic effect, 46 patients receiving high dose methotrexate (5 to 8 g/m2) infusions have been studied. Significant increase in serum phenylalanine was observed in 95% of methotrexate cycles, occurring at the end of infusion. In contrast to the large inter-individual variations, maximal phenylalanine concentrations were of the same magnitude in each individual, suggesting individual predispositions. The hypothesis of an inhibition of dihydropteridine reductase by methotrexate was supported by the parallel course of serum biopterin and phenylalanine levels, but in some way contradicted by the rapid return to baseline values of both, 24 hours after the end of methotrexate infusion. This transient and often moderate hyperphenylalaniemia is probably harmless except if it reflects a more general inhibition of pteridine-dependent hydroxylases. Especially, such an inhibition of cerebral tyrosine- and tryptophan-hydroxylase activities might be the reason for transient neurological disturbances observed in some patients on high-dose methotrexate treatment.

[Metabolism of non-conjugated pteridines in man]. / Métabolisme des ptéridines non conjuguées chez l'homme.

In the review, we have attempted to show the growing momentum in the area of unconjugated pteridine research. The de novo biosynthetic pathway of tetrahydrobiopterin has been recently clarified. The development of specific assays to measure pteridines in biological fluids has facilitated the acquisition of data on changes in the levels of pteridines during the course of various diseases. Several human inborn errors of its metabolism have been discovered and have become an important resource to elucidate the consequences of tetrahydrobiopterin deficiency. Changes in pteridine metabolism have also been investigated in diseases involving stimulation of cellular mediated immunological processes.

Unconjugated pteridines in bronchoalveolar lavage as indicators of alveolar macrophage activation.

Bronchoalveolar lavage (BAL) has shown great efficacy in clarifying the role of immune processes in many disorders of the lower respiratory tract. Following the in vitro demonstration that neopterin is an indicator of the activation of macrophages, neopterin was measured in the BAL fluid and cells from patients with various pulmonary diseases. In most of the patients, high levels of neopterin were found in the serum, BAL fluid and BAL cells. Because neopterin in BAL fluid results from local production as well as from plasma transudation, neopterin in BAL cells seems to reflect the macrophage stimulation more directly. In addition, the correlation between cellular neopterin and lymphocyte count was found to be more significant than the correlation between cellular neopterin and macrophage count. Neopterin in BAL fluid and cells may be a useful measurement in the investigation and elucidation of pulmonary pathologies involving the cellular immune system.

Neonatal hyperphenylalaninaemia presumably caused by a new variant of biopterin synthetase deficiency.

Systematic investigation of hyperphenylalaninaemic infants for tetrahydrobiopterin deficiency has recently led to the description of new variants of cofactor deficiency. In the present case, the initial observation was of hyperphenylalaninaemia with a significant increase in the neopterin to biopterin ratio in the urine. A tetrahydrobiopterin loading test resulted in a significant decrease of blood phenylalanine levels. Cerebrospinal fluid (CSF) biopterin and neurotransmitter metabolite levels were within the normal range. The in vivo clearance of phenylalanine remained altered despite a high dietary tolerance. At 9 months of age, the patient was clinically well, but minor neurological signs appeared when blood phenylalanine levels increased. These data were similar to those found in the so-called "peripheral form" of tetrahydrobiopterin deficiency. However, an unidentified pteridine-like compound had been found in the urine and CSF since the birth, suggesting the existence of an unknown block in the biosynthetic pathway of biopterin.

[Comparison of 11 electrochemical detectors used in high performance liquid chromatography]. / Comparaison de onze détecteurs électrochimiques utilisés en chromatographie liquide haute performance.

Electrochemical detection is more and more frequently used in clinical chemistry. The performances of eleven detectors (sensitivity stability, limit of detection, figure of merit) have been compared. There is a large variation in results for sensitivity and figure of merit, but all of these detectors seem stable and reproducible.

[Evaluation of 6-years' experience of screening for hyperphenylalaninemia caused by cofactor deficiency]. / Bilan de 6 années de dépistage des hyperphénylalaninémies par déficit en cofacteur.

Since the treatment of tetrahydrobiopterin deficiency differs from that for phenylalanine-hydrolase deficient patients, early differential diagnosis of the various types of hyperphenylalaninemia is essential. Results of screening for tetrahydrobiopterin deficiency in 261 cases with hyperphenylalaninemia, including 152 newborns, are reported. Urinary pteridine analysis appears as a valuable method for the recognition of cofactor deficiency. However, accurate interpretation depends on the quality of urine sampling (especially for DHRP deficiency recognition), age-paired comparison and possible pathological interferences.

Unconjugated pteridines in amniotic fluid during gestation.

Neopterin and biopterin concentrations were measured in amniotic fluid in 226 pregnancies from the 12th week of gestation to term. At mid-gestation, neopterin and biopterin levels were low and remained relatively constant between 12 and 26 weeks of gestation, whereas during the third trimester, a progressive increase was observed. Near term the values were greater than those in maternal serum and the higher neopterin to biopterin ratio suggested that pteridine concentration in amniotic fluid may reflect the maturation of pteridine metabolism in the fetus.

Developmental aspects of pteridine metabolism and relationships with phenylalanine metabolism.

Large variations of pteridine elimination occur in childhood, due to the ontogenic development of the metabolism of tetrahydrobiopterin. The main feature is the slow maturation of biopterin synthesis whereas neopterin synthesis is high at birth; thus a high neopterin to biopterin ratio (4.4 +/- 2.1) occurs in the neonatal period, a ratio which then decreases to adult values (0.5 +/- 0.2). Comparing pteridine elimination of PKU patients with that of controls of the same age, a high excretion of biopterin and, to a lesser extent, of neopterin is found. In normal subjects, following an oral phenylalanine load, biopterin levels in urine and serum also increase, whereas variations of neopterin concentration are small. In rats, phenylalanine also leads to an increase of serum biopterin whereas liver biopterin decreases. This suggests that the main explanation for the biopterin increase in serum and in urine by phenylalanine is a release of the intracellular biopterin by the aminoacid.