Phenylketonuria: reduced tyrosine brain influx relates to reduced cerebral protein synthesis.
ABSTRACT: BACKGROUND: In phenylketonuria (PKU), elevated blood phenylalanine (Phe) concentrations are considered to impair transport of large neutral amino acids (LNAAs) from blood to brain. This impairment is believed to underlie cognitive deficits in PKU via different mechanisms, including reduced cerebral protein synthesis. In this study, we investigated the hypothesis that impaired LNAA influx relates to reduced cerebral protein synthesis. METHODS: Using positron emission tomography, L-[1-11C]-tyrosine (11C-Tyr) brain influx and incorporation into cerebral protein were studied in 16 PKU patients (median age 24, range 16 - 47 years), most of whom were early and continuously treated. Data were analyzed by regression analyses, using either 11C-Tyr brain influx or 11C-Tyr cerebral protein incorporation as outcome variable. Predictor variables were baseline plasma Phe concentration, Phe tolerance, age, and 11C-Tyr brain efflux. For the modelling of cerebral protein incorporation, 11C-Tyr brain influx was added as a predictor variable. RESULTS: 11C-Tyr brain influx was inversely associated with plasma Phe concentrations (median 512, range 233 - 1362 ?mol/L; delta adjusted R2=0.571, p=0.013). In addition, 11C-Tyr brain influx was positively associated with 11C-Tyr brain efflux (delta adjusted R2=0.098, p=0.041). Cerebral protein incorporation was positively associated with 11C-Tyr brain influx (adjusted R2=0.567, p<0.001). All additional associations between predictor and outcome variables were statistically nonsignificant. CONCLUSIONS: Our data favour the hypothesis that an elevated concentration of Phe in blood reduces cerebral protein synthesis by impairing LNAA transport from blood to brain. Considering the importance of cerebral protein synthesis for adequate brain development and functioning, our results support the notion that PKU treatment be continued in adulthood. Future studies investigating the effects of impaired LNAA transport on cerebral protein synthesis in more detail are indicated.
Project description:The standard treatment for phenylketonuria (PKU) is a lifelong low-phenylalanine (Phe) diet, supplemented with Phe-free protein substitutes; however, adult patients often show poor adherence to therapy. Alternative treatment options include the use of large neutral amino acids (LNAA). The aim of this study was to determine the Phe, tyrosine (Tyr), and Phe/Tyr ratio in a cohort of sub-optimally controlled adult patients with classical PKU treated with a new LNAA formulation. Twelve patients received a Phe-restricted diet plus a slow-release LNAA product taken three times per day, at a dose of 1 g/kg body weight (mean 0.8 ± 0.24 g/kg/day), over a 12-month period. The product is in a microgranulated formulation, which incorporates all amino acids and uses sodium alginate as a hydrophilic carrier to prolong its release. This LNAA formulation provides up to 80% of the total protein requirement, with the rest of the protein supplied by natural food. Patients had fortnightly measurements of Phe and Tyr levels over a 12-month period after the introduction of LNAA. All patients completed the 12-month treatment period. Overall, adherence to the new LNAA tablets was very good compared with a previous amino acid mixture, for which taste was a major complaint by patients. Phe levels remained unchanged (p = 0.0522), and Tyr levels increased (p = 0.0195). Consequently, the Phe/Tyr ratio decreased significantly (p < 0.05) in the majority of patients treated. In conclusion, LNAA treatment increases Tyr levels in sub-optimally controlled adult PKU patients, while offering the potential to improve their adherence to treatment.
Project description:Monoamine neurotransmitter deficiency has been implicated in the etiology of neuropsychiatric symptoms associated with chronic hyperphenylalaninemia in phenylketonuria (PKU). Two proposed explanations for neurotransmitter deficiency in PKU include first, that chronically elevated blood L-phenylalanine (Phe) inhibits the transport of L-tyrosine (Tyr) and L-tryptophan (Trp), the substrates for dopamine and serotonin synthesis respectively, into brain. In the second hypothesis, elevated Phe competitively inhibits brain tyrosine hydroxylase (TH) and tryptophan hydroxylase (TPH) activities, the rate limiting steps in dopamine and serotonin synthesis. Dietary supplementation with large neutral amino acids (LNAA) including Tyr and Trp has been recommended for individuals with chronically elevated blood Phe in an attempt to restore amino acid and monoamine homeostasis in brain. As a potential alternative treatment approach, we demonstrate that pharmacologic inhibition of Tyr degradation through oral administration of nitisinone (NTBC) yielded sustained increases in blood and brain Tyr, decreased blood and brain Phe, and consequently increased dopamine synthesis in a murine model of PKU. Our results suggest that Phe-mediated inhibition of TH activity is the likely mechanism of impaired dopamine synthesis in PKU. Pharmacologic inhibition of Tyr degradation may be a promising adjunct therapy for CNS monoamine neurotransmitter deficiency in hyperphenylalaninemic individuals with PKU.
Project description:The gold standard treatment for phenylketonuria (PKU) is a lifelong low-phenylalanine (Phe) diet supplemented with Phe-free protein substitutes. Adherence to therapy becomes difficult after childhood. Supplementing with large neutral amino acids (LNAAs) has been proposed as an alternative medication to Phe-free protein substitutes (i.e., amino acid mixtures). The aim of this study was to evaluate adherence to therapy and quality of life (QoL) in a cohort of sub-optimally controlled adult PKU patients treated with a new LNAA formulation. Twelve patients were enrolled in a 12-month-trial of slow-release LNAAs (1g/kg/day) plus a Phe-restricted diet. Medication adherence was measured with the Morisky Green Levine Medication Adherence Scale; the QoL was measured using the phenylketonuria-quality of life (PKU-QoL) questionnaire. Phe, tyrosine (Tyr) levels, and Phe/Tyr ratios were measured fortnightly. Before treatment, 3/12 patients self-reported a 'medium' adherence to medication and 9/12 reported a low adherence; 60% of patients reported a full adherence over the past four weeks. After 12 months of LNAA treatment, all patients self-reported a high adherence to medication, with 96% reporting a full adherence. Phe levels remained unchanged, while Tyr levels increased in most patients. The Phy/Tyr ratio decreased. All patients had a significant improvement in the QoL. LNAAs may give patients a further opportunity to improve medication adherence and, consequently, their QoL.
Project description:BACKGROUND:Phenylketonuria (PKU) is due to a defective hepatic enzyme, phenylalanine (Phe) hydroxylase. Transport of the precursor amino acids from blood into the brain for serotonin and dopamine synthesis is reported to be inhibited by high blood Phe concentrations. Deficiencies of serotonin and dopamine are involved in neurocognitive dysfunction in PKU. OBJECTIVE:(1) To evaluate the effects of sapropterin (BH4) and concurrent use of large neutral amino acids (LNAA) on the peripheral biomarkers, melatonin and dopamine with the hypothesis they reflect brain serotonin and dopamine metabolism. (2) To evaluate synergistic effects with BH4 and LNAA. (3) To determine the effects of blood Phe concentrations on the peripheral biomarkers concentrations. METHODS:Nine adults with PKU completed our study consisting of four 4-week phases: (1) LNAA supplementation, (2) Washout, (3) BH4 therapy, and (4) LNAA with BH4 therapy. An overnight protocol measured plasma amino acids, serum melatonin, and 6-sulfatoxymelatonin and dopamine in first void urine after each phase. RESULTS:(1) Three out of nine subjects responded to BH4. A significant increase of serum melatonin levels was observed in BH4 responders with decreased blood Phe concentration. No significant change in melatonin, dopamine or Phe levels was observed with BH4 in the subjects as a whole. (2) Synergistic effects with BH4 and LNAA were observed in serum melatonin in BH4 responders. (3) The relationship between serum melatonin and Phe showed a significant negative slope (p = 0.0005) with a trend toward differing slopes among individual subjects (p = 0.066). There was also a negative association overall between blood Phe and urine 6-sulfatoxymelatonin and dopamine (P = 0.040 and 0.047). CONCLUSION:Blood Phe concentrations affected peripheral monoamine neurotransmitter biomarker concentrations differently in each individual with PKU. Melatonin levels increased with BH4 therapy only when blood Phe decreased. Monitoring peripheral neurotransmitter metabolites may assist in optimizing individualized treatment in PKU.
Project description:Many phenylketonuria (PKU) patients cannot adhere to the severe dietary restrictions as advised by the European PKU guidelines, which can be accompanied by aggravated neuropsychological impairments that, at least in part, have been attributed to brain monoaminergic neurotransmitter deficiencies. Supplementation of large neutral amino acids (LNAA) to an unrestricted diet has previously been shown to effectively improve brain monoamines in PKU mice of various ages. To determine the additive value of LNAA supplementation to a liberalized phenylalanine-restricted diet, brain and plasma monoamine and amino acid concentrations in 10 to 16-month-old adult C57Bl/6 PKU mice on a less severe phenylalanine-restricted diet with LNAA supplementation were compared to those on a non-supplemented severe or less severe phenylalanine-restricted diet. LNAA supplementation to a less severe phenylalanine-restricted diet was found to improve both brain monoamine and phenylalanine concentrations. Compared to a severe phenylalanine-restricted diet, it was equally effective to restore brain norepinephrine and serotonin even though being less effective to reduce brain phenylalanine concentrations. These results in adult PKU mice support the idea that LNAA supplementation may enhance the effect of a less severe phenylalanine-restricted diet and suggest that cerebral outcome of PKU patients treated with a less severe phenylalanine-restricted diet may be helped by additional LNAA treatment.
Project description:In phenylketonuria, elevated plasma phenylalanine concentrations may disturb blood-to-brain large neutral amino acid (LNAA) transport and cerebral protein synthesis (CPS). We investigated the associations between these processes, using data obtained by positron emission tomography with l-[1-(11)C]-tyrosine ((11)C-Tyr) as a tracer. Blood-to-brain transport of non-Phe LNAAs was modeled by the rate constant for (11)C-Tyr transport from arterial plasma to brain tissue (K1), while CPS was modeled by the rate constant for (11)C-Tyr incorporation into cerebral protein (k3). Brain phenylalanine concentrations were measured by magnetic resonance spectroscopy in three volumes of interest (VOIs): supraventricular brain tissue (VOI 1), ventricular brain tissue (VOI 2), and fluid-containing ventricular voxels (VOI 3). The associations between k3 and each predictor variable were analyzed by multiple linear regression. The rate constant k3 was inversely associated with brain phenylalanine concentrations in VOIs 2 and 3 (adjusted R(2)=0.826, F=19.936, P=0.021). Since brain phenylalanine concentrations in these VOIs highly correlated with each other, the specific associations of each predictor with k3 could not be determined. The associations between k3 and plasma phenylalanine concentration, K1, and brain phenylalanine concentrations in VOI 1 were nonsignificant. In conclusion, our study shows an inverse association between k3 and increased brain phenylalanine concentrations.
Project description:Phenylketonuria (PKU) is an inherited metabolic disease characterized by abnormally high concentrations of the essential amino acid L-phenylalanine (Phe) in blood plasma caused by reduced activity of phenylalanine hydroxylase (PAH). While numerous studies have shown association between high plasma Phe concentration and intellectual impairment, it is not clear whether increased Phe fluctuations also observed in PKU affect the brain as well. To investigate this, time-resolved in vivo data on Phe and competing large neutral amino acid (LNAA) concentrations in neurons are needed, but cannot be acquired readily with current methods. We have used in silico modeling as an alternative approach to characterize the interactive dynamics of Phe and competing LNAAs (CL) in the neurovascular unit (NVU). Our results suggest that plasma Phe fluctuations can propagate into the NVU cells and change there the concentration of LNAAs, with the highest magnitude of this effect observed at low frequency and high amplitude-to-mean ratio of the plasma Phe concentration fluctuations. Our model further elucidates the effect of therapeutic LNAA supplementation in PKU, showing how abnormal concentrations of Phe and CL in the NVU move thereby toward normal physiologic levels.
Project description:Phenylketonuria (PKU) was the first disorder in which severe neurocognitive dysfunction could be prevented by dietary treatment. However, despite this effect, neuropsychological outcome in PKU still remains suboptimal and the phenylalanine-restricted diet is very demanding. To improve neuropsychological outcome and relieve the dietary restrictions for PKU patients, supplementation of large neutral amino acids (LNAA) is suggested as alternative treatment strategy that might correct all brain biochemical disturbances caused by high blood phenylalanine, and thereby improve neurocognitive functioning.As a proof-of-principle, this study aimed to investigate all hypothesized biochemical treatment objectives of LNAA supplementation (normalizing brain phenylalanine, non-phenylalanine LNAA, and monoaminergic neurotransmitter concentrations) in PKU mice.C57Bl/6 Pah-enu2 (PKU) mice and wild-type mice received a LNAA supplemented diet, an isonitrogenic/isocaloric high-protein control diet, or normal chow. After six weeks of dietary treatment, blood and brain amino acid and monoaminergic neurotransmitter concentrations were assessed.In PKU mice, the investigated LNAA supplementation regimen significantly reduced blood and brain phenylalanine concentrations by 33% and 26%, respectively, compared to normal chow (p<0.01), while alleviating brain deficiencies of some but not all supplemented LNAA. Moreover, LNAA supplementation in PKU mice significantly increased brain serotonin and norepinephrine concentrations from 35% to 71% and from 57% to 86% of wild-type concentrations (p<0.01), respectively, but not brain dopamine concentrations (p = 0.307).This study shows that LNAA supplementation without dietary phenylalanine restriction in PKU mice improves brain biochemistry through all three hypothesized biochemical mechanisms. Thereby, these data provide proof-of-concept for LNAA supplementation as a valuable alternative dietary treatment strategy in PKU. Based on these results, LNAA treatment should be further optimized for clinical application with regard to the composition and dose of the LNAA supplement, taking into account all three working mechanisms of LNAA treatment.
Project description:Transport of large neutral amino acids (LNAA) across the blood brain barrier (BBB) is facilitated by the L-type amino acid transporter, LAT1. Peripheral accumulation of one LNAA (e.g., phenylalanine (phe) in PKU) is predicted to increase uptake of the offending amino acid to the detriment of others, resulting in disruption of brain amino acid homeostasis. We hypothesized that selected non-physiological amino acids (NPAAs) such as DL-norleucine (NL), 2-aminonorbornane (NB; 2-aminobicyclo-(2,1,1)-heptane-2-carboxylic acid), 2-aminoisobutyrate (AIB), and N-methyl-aminoisobutyrate (MAIB), acting as competitive inhibitors of various brain amino acid transporters, could reduce brain phe in Pah (enu2) mice, a relevant murine model of PKU. Oral feeding of 5 % NL, 5 % AIB, 0.5 % NB and 3 % MAIB reduced brain phe by 56 % (p < 0.01), -1 % (p = NS), 27 % (p < 0.05) and 14 % (p < 0.01), respectively, compared to untreated subjects. Significant effects on other LNAAs (tyrosine, methionine, branched chain amino acids) were also observed, however, with MAIB displaying the mildest effects. Of interest, MAIB represents an inhibitor of the system A (alanine) transporter that primarily traffics small amino acids and not LNAAs. Our studies represent the first in vivo use of these NPAAs in Pah (enu2) mice, and provide proof-of-principle for their further preclinical development, with the long-term objective of identifying NPAA combinations and concentrations that selectively restrict brain phe transport while minimally impacting other LNAAs and downstream intermediates.
Project description:Notwithstanding the success of the traditional dietary phenylalanine restriction treatment in phenylketonuria (PKU), the use of large neutral amino acid (LNAA) supplementation rather than phenylalanine restriction has been suggested. This treatment modality deserves attention as it might improve cognitive outcome and quality of life in patients with PKU. Following various theories about the pathogenesis of cognitive dysfunction in PKU, LNAA supplementation may have multiple treatment targets: a specific reduction in brain phenylalanine concentrations, a reduction in blood (and consequently brain) phenylalanine concentrations, an increase in brain neurotransmitter concentrations, and an increase in brain essential amino acid concentrations. These treatment targets imply different treatment regimes. This review summarizes the treatment targets and the treatment regimens of LNAA supplementation and discusses the differences in LNAA intake between the classical dietary phenylalanine-restricted diet and several LNAA treatment forms.