Does hyperphenylalaninemia induce brain glucose hypometabolism? Cerebral spinal fluid findings in treated adult phenylketonuric patients.

Despite numerous studies in human patients and animal models for phenylketonuria (PKU; OMIM#261600), the pathophysiology of PKU and the underlying causes of brain dysfunction and cognitive problems in PKU patients are not well understood. In this study, lumbar cerebral spinal fluid (CSF) was obtained immediately after blood sampling from early-treated adult PKU patients who had fasted overnight. Metabolite and amino acid concentrations in the CSF of PKU patients were compared with those of non-PKU controls. The CSF concentrations and CSF/plasma ratios for glucose and lactate were found to be below normal, similar to what has been reported for glucose transporter1 (GLUT1) deficiency patients who exhibit many of the same clinical symptoms as untreated PKU patients. CSF glucose and lactate levels were negatively correlated with CSF phenylalanine (Phe), while CSF glutamine and glutamate levels were positively correlated with CSF Phe levels. Plasma glucose levels were negatively correlated with plasma Phe concentrations in PKU subjects, which partly explains the reduced CSF glucose concentrations. Although brain glucose concentrations are unlikely to be low enough to impair brain glucose utilization, it is possible that the metabolism of Phe in the brain to produce phenyllactate, which can be transported across the blood-brain barrier to the blood, may consume glucose and/or lactate to generate the carbon backbone for glutamate. This glutamate is then converted to glutamine and carries the Phe-derived ammonia from the brain to the blood. While this mechanism remains to be tested, it may explain the correlations of CSF glutamine, glucose, and lactate concentrations with CSF Phe.

Retrospective analysis of 19 patients with 6-Pyruvoyl Tetrahydropterin Synthase Deficiency: Prolactin levels inversely correlate with growth.

BACKGROUND: Pyruvoyl Tetrahydropterin Synthase (PTPS) Deficiency is the most common form of BH4 deficiency resulting in hyperphenylalaninemia. It can have variable clinical severity and there is limited information on the clinical presentation, natural history and effectiveness of newborn screening for this condition. METHODS: Retrospective data (growth and clinical parameters, biochemical and genetic testing results, treatment) were collected from 19 patients with PTPS deficiency in different centers, to evaluate biochemical and clinical outcomes. Descriptive statistics was used for qualitative variables, while linear regression analysis was used to correlate quantitative variables. RESULTS: Patients with PTPS deficiency had an increased incidence of prematurity (4/18) with an average gestational age only mildly reduced (37.8 ± 2.4 weeks) and low birth weight (-1.14 ± 0.97 SD below that predicted for gestational age). With time, weight and height approached normal. VALUES: All patients were identified by newborn screening for an elevated phenylalanine level. However, phenylalanine levels were normal in two whose testing was performed at or before 24 h of age. Sapropterin dihydrochloride treatment normalized phenylalanine levels. Molecular testing identified novel variants in the PTS gene, some of which present in more than one affected family. The neurotransmitter derivatives 5-hydroxyindoleacetic acid (5HIAA) and homovanillic acid (HVA) in the CSF were decreased in most cases except in 2 families with the peripheral form of PTPS deficiency. With time, HVA and 5HIAA became abnormally low in two of these patients requiring therapy. Prolactin (whose secretion is inhibited by dopamine) levels were elevated in several patients with PTPS deficiency and inversely correlated with the z-scores for height (p < 0.01) and weight (p < 0.05). Most patients with PTPS deficiency had delayed development early in life, improving around school age with IQs mostly in the normal range, with a small decline in older individuals. From a neurological standpoint, most patients had normal brain MRI and minor EEG anomalies, although some had persistent neurological symptoms. DISCUSSION: Patients with PTPS deficiency have not only an increased incidence of prematurity, but also decreased birth weight when corrected for gestational age. Hyperphenylalaninemia can be absent in the first day of life. Therapy with sapropterin dihydrochloride normalizes phenylalanine levels and neurotransmitter precursors can improve CSF neurotransmitter metabolites levels. Insufficient dopaminergic stimulation (as seen from elevated prolactin) might result in decreased height in patients with PTPS deficiency. Despite early delays in development, many patients can achieve independence in adult life, with usually normal neuroimaging and EEG.

Long-term clinical outcome of 6-pyruvoyl-tetrahydropterin synthase-deficient patients.

INTRODUCTION: 6-Pyruvoyl-tetrahydropterin synthase deficiency (PTPSd) is a rare autosomal recessive disorder of synthesis of biogenic amines, which is characterized by variable neurological impairment and hyperphenylalaninemia. We aimed to assess the long-term clinical outcome of this disorder and the factors affecting it. METHODS: At total of 28 PTPSd patients (aged 19.9 ±â€¯10.9 years) underwent clinical (neurological and psychiatric) and neuropsychological assessment (BRIEF, VABS-II, and IQ). Based on CSF homovanillic (HVA) and 5-hydroxyindolacetic acid (5-HIAA) and pterin concentrations at diagnosis, patients were classified as having either a severe [SF; low level of CSF, HVA, and 5-HIAA with altered neopterin/biopterin (Neo/Bio)] or mild form (MF; normal HVA and 5-HIAA with altered Neo/Bio) of PTPSd. RESULTS: Approximately 36% of patients had MF PTPSd. At the last examination, 43% of patients had movement disorders (2 MF, 10 SF), 43% of patients had variable degrees of intellectual disability (SF only), 39% met the criteria for a psychiatric disorder (3 MF, 9 SF). Applying a linear regression model, we found that HVA and phenylalanine levels at birth had a significant influence on IQ, BRIEF, and VABS-II variability. Lastly, 5-HIAA further contributed to VABS-II variability. The disease showed a self-limiting clinical course and its treatment, although delayed, is effective in improving the neurological status. CONCLUSIONS: Neurodevelopmental impairment due to PTPSd shows a self-limiting course. A continuous improvement in the neurological condition has been observed in patients receiving treatment, even when delayed. The severity of brain biogenic amine depletion at diagnosis predicts neurological and psychiatric outcomes.

Cerebrospinal fluid biogenic amines depletion and brain atrophy in adult patients with phenylketonuria.

Biogenic amines synthesis in phenylketonuria (PKU) patients with high phenylalanine (Phe) concentration is thought to be impaired due to inhibition of tyrosine and tryptophan hydroxylases and competition with amino acids at the blood-brain barrier. Dopamine and serotonin deficits might explain brain damage and progressive neuropsychiatric impairment in adult PKU patients. Ten early treated adult PKU patients (mean age 38.2 years) and 15 age-matched controls entered the study. Plasma and cerebrospinal fluid (CSF) Phe, 5-hydroxyindoleacetic acid (5-HIAA), 5-hydroxytryptophan (5-HTP), 3,4-dihydroxy-l-phenylalanine (l-DOPA) and homovanillic acid (HVA) were analyzed. Voxel-based morphometry statistical nonparametric mapping was used to test the age-corrected correlation between gray matter atrophy and CSF biogenic amines levels. 5-HIAA and 5-HTP were significantly reduced in PKU patients compared to controls. Significant negative correlations were found between CSF 5-HIAA, HVA, and 5-HTP and Phe levels. A decrease in 5-HIAA and 5-HTP concentrations correlated with precuneus and frontal atrophy, respectively. Lower HVA levels correlated with occipital atrophy. Biogenic amines deficits correlate with specific brain atrophy patterns in adult PKU patients, in line with serotonin and dopamine projections. These findings may support a more rigorous Phe control in adult PKU to prevent neurotransmitter depletion and accelerated brain damage due to aging.

DNAJC12 deficiency: A new strategy in the diagnosis of hyperphenylalaninemias.

Patients with hyperphenylalaninemia (HPA) are detected through newborn screening for phenylketonuria (PKU). HPA is known to be caused by deficiencies of the enzyme phenylalanine hydroxylase (PAH) or its cofactor tetrahydrobiopterin (BH4). Current guidelines for the differential diagnosis of HPA would, however, miss a recently described DNAJC12 deficiency. The co-chaperone DNAJC12 is, together with the 70kDa heat shock protein (HSP70), responsible for the proper folding of PAH. All DNAJC12-deficient patients investigated to date responded to a challenge with BH4 by lowering their blood phenylalanine levels. In addition, the patients presented with low levels of biogenic amine in CSF and responded to supplementation with BH4, L-dopa/carbidopa and 5-hydroxytryptophan. The phenotypic spectrum ranged from mild autistic features or hyperactivity to severe intellectual disability, dystonia and parkinsonism. Late diagnosis result in permanent neurological disability, while early diagnosed and treated patients develop normally. Molecular diagnostics for DNAJC12 variants are thus mandatory in all patients in which deficiencies of PAH and BH4 are genetically excluded.

Reduced nitric oxide metabolites in CSF of patients with tetrahydrobiopterin deficiency.

We investigated CSF concentrations of nitrite and nitrate as indicators of nitric oxide (NO) production in patients with tetrahydrobiopterin (BH4) deficiencies. Patients with 6-pyruvoyl-tetrahydropterin synthase, sepiapterin reductase and dihydropteridine reductase deficiencies exhibited decreased CSF nitrite + nitrate levels compared with healthy control subjects. Reduced levels of nitrite + nitrate were not influenced by oral administration of 2.5-5.0 mg/kg tetrahydrobiopterin. Our data indicate impaired NO synthase function in patients with BH4 deficiency and suggest possible involvement in the neuronal cell dysfunction.

Indications of changed amino acid homeostasis in untreated and treated PKU.

Amino acid patterns were investigated in children with untreated and treated phenylketonuria (PKU) and compared with a hospitalized paediatric control group. Correlation ratios, i.e. the ratio between the number of statistically significant correlations between pairs of amino acids, were calculated and found to be 0.125, 0.331 and 0.35 in the three groups. Link ratios, i.e. the frequency of statistically significant correlations between a particular amino acid and other amino acids, varied between 0.00 and 0.18, 0.06 and 0.56, and 0.18 and 0.59 in the same groups. Link ratios for especially isoleucine, histidine and arginine were decreased in treated PKU compared to the control group. A reverse nonlinear relationship was established between decreased plasma phenylalanine induced by dietary treatment and increased threonine in plasma. These findings indicate that amino acid homeostasis is disturbed in untreated PKU and not fully normalized in treated patients. The clinical relevance of the findings is unknown.

Correlation between cerebrospinal fluid phenylalanine and beta-endorphin in patients with phenylketonuria.

Previous animal and human studies have suggested an analgesic effect of phenylalanine involving endogenous opioid peptides. Phenylalanine was measured by a HPLC method with electrochemical detection and beta-endorphin by a specific radioimmunoassay in 14 lumbar cerebrospinal fluid samples from 13 patients with phenylketonuria. Cerebrospinal fluid beta-endorphin was also determined in 6 age-matched control subjects. We found a trend towards a higher beta-endorphin level in phenylketonuria (median 26.0 pM, range 13.0-37.8) than in the control subjects (20.6 pM, 12.7-28.0), P = 0.13. Cerebrospinal fluid concentrations of phenylalanine and beta-endorphin were significantly correlated (r = 0.68, P = 0.008). The results support the hypothesis that phenylalanine modifies the central endogenous opioid system.

Cerebrospinal fluid beta-endorphin in Rett syndrome.

An hypothesis of increased endorphinergic activity has been proposed to account for the characteristic symptoms of Rett syndrome. Cerebrospinal fluid samples from eight girls with Rett syndrome were analysed for beta-endorphin (beta-EP) immunoactivity and compared with samples from a control group of 15 children with acute leukaemia in remission. Severity of symptoms was not found to be related to beta-EP level. A group of early-treated adolescents with phenylketonuria had beta-EP levels similar to the Rett syndrome patients, but no symptoms resembling theirs. Therefore it is unlikely that increased levels of beta-EP are of primary pathogenetic significance. The conflicting findings of many earlier reports may be a result of differences between control groups.

Content of phenylalanine, tyrosine and their metabolites in CSF in phenylketonuria.

By using ion-exchange chromatography and gas chromatography coupled with mass spectrometry, the content of phenylalanine, tyrosine and their metabolites typical of phenylketonuria (PKU) was determined in the cerebrospinal fluid (CSF) of 8 untreated children with classical PKU and 9 controls. At the same time, plasma and urine were analysed. In PKU the content of phenylalanine is increased on average 23 times in plasma and CSF. The content of phenylalanine and tyrosine in CSF is about 4 times less as compared with plasma. The phenylalanine-to-tyrosine ratio is approximately the same for these fluids both in control and in PKU. This indicates that the transport of phenylalanine and tyrosine through the blood-brain barrier is not disturbed in PKU. Phenylpyruvate and 4-hydroxyphenylpyruvate are either not detected or present in very low concentrations in the CSF of children with PKU; their derivatives, phenyllactate and 4-hydroxyphenyllactate, are present in relatively higher concentrations. This indicates increased metabolic conversion in brain tissues.

On the brain barrier system function and changes of cerebrospinal fluid concentrations of phenylalanine and tyrosine in human phenylketonuria.

In 6 patients with classic phenylketonuria (PKU) the plasma and cerebrospinal fluid (CSF) concentrations of phenylalanine and tyrosine were measured fluorimetrically. The results of the PKU group were compared with data obtained from 17 children without abnormal CSF parameters and free of metabolic or central nervous disorders, in whom a diagnostic lumbal puncture has been performed. The PKU patients showed statistically significant differences in comparison with the controls: plasma and CSF phenylalanine contents were markedly higher (on the average 6.4 and 4.6 times, respectively) in PKU patients. Plasma tyrosine was 1.8 times lower, but CSF tyrosine was about 2.2 times higher in comparison to the controls. In general, the plasma CSF-ratio ( PLR ) of phenylalanine did not change in PKU and could be found in the same range as in the normal controls. In contrast to this, the PLR of tyrosine was found to be significantly lower in PKU patients. The results are discussed with respect to an altered function of the brain barrier systems for the amino acid transport in PKU, and that increased CSF tyrosine contents in PKU may rather reflect disturbances of the intracellular metabolism of the brain cells than changes of the amino acid transport through the brain barrier produced by hyperphenylalaninemia.

Use of tetrahydropterins in the treatment of hyperphenylalaninemia due to defective synthesis of tetrahydrobiopterin: evidence that peripherally administered tetrahydropterins enter the brain.

Substantial amounts of tetrahydrobiopterin and 6-methyltetrahydropterin can be detected in CSF when these pterins are given peripherally to patients with hyperphenylalaninemia due to defective biopterin synthesis. Results of this study suggest that administration of either of these pterins in proper doses may prove to be a treatment not only for the impaired peripheral phenylalanine metabolism, but also for the neurologic disorders that are characteristic of the variant forms of hyperphenylalaninemia due to defective tetrahydrobiopterin synthesis or metabolism.

Impaired permeability of the blood-cerebrospinal fluid barrier in hyperphenylalaninaemia.

In hyperphenylalaninaemic rats the accumulation of 5-hydroxytryptophan (5-HTP) in the cerebrospinal fluid (CSF) could be shown in spite of the fact, that the 5-HTP levels of serum and brain remain constant. In vitro studies of the influx and efflux of 5-HTP and phenylalanine on isolated beef choroid plexus suggested that both aminoacids use the same carrier system. It is concluded that a high concentration of phenylalanine inhibits the re-uptake of 5-HTP by the endothelial cells of the choroid plexus. Additionally, an increased efflux of 5-HTP from choroid plexus leads to the accumulation of 5-HTP in the cerebrospinal fluid.

Comparison of CSF-free amino acid concentrations among various neuropathological conditions.

The level of free amino acids in cerebrospinal fluid from eight patients in comatose states has been determined. The variations of cerebrospinal fluid-free amino acids observed in coma have been compared with those reported by other authors in patients affected by epilepsy, multiple sclerosis, infantile amaurotic idiocy (GM2-gangliosidosis) and phenylketonuria.