By Joachim Pietz, M.D.
Dietary treatment of phenylketonuria (PKU) places stringent demands on patients. Some patients are unable to meet these demands early in life and subsequently exhibit poor outcomes. High levels of cerebral phenylalanine (Phe) can also cause acute but most likely reversible brain dysfunction in older and normally developed patients with PKU (Krause, Epstein, Averbook, et al., 1986; Krause, Halminski, McDonald, et al., 1985; Leuzzi, Fois, Carducci, et al., 1997). Consequently, attempts have been made to find alternative treatments for young noncompliant patients and for adolescents and adults unable to adhere to a strict diet. Three approaches have been used: (1) to overcome suspected depletion of cerebral amine neurotransmitters, patients with PKU off diet have been treated with tyrosine (Tyr), L-dopa, and tryptophan (Trp), both individually and in combination; (2) the branched-chain amino acids valine, isoleucine, and leucine (VIL), and a mixture of all large neutral amino acids (LNAA), have been administered to reduce Phe influx into the brain; and (3) ammonia lyase capsules have been developed to reduce uptake of Phe from the gastrointestinal tract.
Use of neurotransmitter precursors
In vitro and animal studies, as well as investigation of specimens (plasma, urine, and cerebrospinal fluid [CSF]) from patients with PKU, have shown that increased Phe interferes with amine neurotransmitter synthesis.Because an increase in Phe has led to decreases in plasma L-dopa in older patients with PKU, it was speculated that Phe-related brain dysfunction may be caused by disturbed dopamine synthesis (Krause, Epstein, Averbrook, et al., 1986; Krause, Halminski, McDonald, et al., 1985).Dopamine depletion and the resulting dysfunction of the prefrontal cortex then became a focus of research.Lou (1985) first reported that supplementation of a free diet with high doses of both Tyr and Trp caused normalization of amine neurotransmitter metabolites in CSF and improvement in performance of a reaction-time task in two patients with PKU.Tyr alone was used in two studies and also seemed to improve cognitive function (Lou, Lykkelund, Gerdes, et al., 1987; Lykkelund, Nielson, Lou, et al., 1988).Trp supplementation alone normalized serotonin metabolites in CSF, but cognitive function was not investigated (Lykkelund, Nielson, Lou, et al., 1988).
These results were said to confirm the theoretical framework of disturbed neurotransmitter synthesis and its effects on brain function in PKU.However, two placebo-controlled studies failed to confirm the results (Pietz, Landwehr, Kutscha, et al., 1995; Smith, Hanley, Clarke, et al., 1998).Although both of the latter studies demonstrated impairment of brain function in adult patients with PKU off a strict diet, no change was observed during high-dose Tyr treatment.In another study, L-dopa was used and failed to demonstrate beneficial effects on VEP (visually evoked potentials) changes (Ullrich, Weglage, Oberwittler, et al., 1994).
Several points must be made about these various studies.First, the two studies with positive results apparently included the same patients and therefore do not confirm each other.Second, the samples were heterogeneous, with approximately 50 percent neurologically impaired and late-treated patients.Third, improvement in brain function was confirmed for only one parameter, the 90 percentile values from a reaction-time task.The clinical relevance of this parameter remains unclear.
Other explanations for the discrepant results of the studies should be mentioned.First, low concentrations of neurotransmitters in plasma, urine, and CSF may be misleading indicators of intracellular concentrations in the central nervous system (CNS) neuron.Second, low neurotransmitters may be clinically insignificant and show only a relative depletion that may be counterbalanced by autoregulative processes—for example, up-regulation of receptor density or activity.Third, it may be that blockage of the rate-limiting enzyme Tyr hydroxylase (Tyr to L-dopa) is the relevant mechanism, and elevation of substrate Tyr could therefore have no effect.This hypothesis, however, is contradicted by confirmation of increased CSF neurotransmitters during Tyr treatment.An early attempt at high-dose Tyr supplementation in a young child failed to prevent brain damage (Batshaw, Valle, Bessman, 1981).It therefore is not likely that dopamine depletion plays a significant role in the etiology of permanent brain damage in untreated PKU.
In summary, the clinical relevance of impaired neurotransmitter synthesis in PKU is not clear.Although treatment with Tyr and Trp has been shown to influence neurotransmitter synthesis, clinically relevant improvements of brain function following Tyr, Trp, or L-dopa treatment have not been confirmed, and high-dose Tyr, Trp, and L-dopa treatment cannot be recommended.
Attempts to block Phe influx through the blood-brain barrier
LNAAs, including Phe, compete for transport across the blood-brain barrier via the L-type amino acid carrier.Net uptake through the blood-brain barrier is determined by their ratio in plasma and differing affinity to the carrier system.Carrier saturation and inhibition of blood-brain barrier transport can be expected at the levels of supraphysiological plasma Phe usually found in PKU and may even be present at concentrations in the range of 200-500 mol/L (Pardridge, 1998).The direct effects of elevated brain Phe and depleted LNAAs are probably major causes of disturbed brain development and function in PKU.Competition for the carrier therefore might be put to use to lower the influx of Phe by increasing plasma concentrations of other LNAAs.
Branched-chain amino acids: valine, leucine, isoleucin (VIL)
Oral VIL treatment was designed to inhibit influx of Phe into the brain.Although plasma Phe remained unchanged during VIL intake, a decrease of approximately 20 percent in concentration in CSF confirmed the hypothesis (Berry, Bofinger, Hunt, et al., 1982), which was corroborated by concomitant, moderate improvements in neuropsychological performance (Berry, Brunner, Hunt, et al., 1990).CSF Tyr concentrations were lowered during VIL treatment, and administration of VIL did not produce any relevant side effects.Leuzzi and colleagues (1997) recently reported beneficial effects of VIL treatment on neuropsychological performance during high Phe intake, with no MRI-visible changes in white matter.It remains unclear, however, why these three branched-chain amino acids were chosen and other LNAAs were neglected.The rate of protein synthesis depends on a supply of all essential amino acids, and an increase in only three (i.e., VIL) may accentuate disturbances of neurotransmitter or protein synthesis.
In summary, VIL treatment was able to lower CSF concentrations of Phe, and effects on brain function were moderate, but VIL therapy cannot be recommended on the basis of the results of published studies.
Administration of LNAAs
Since earlier studies showed that cerebral Phe can be measured noninvasively by proton MR spectroscopy ( 1H-MRS), 1H-MRS was used in a recent study which investigated Phe transport through the blood-brain barrier in patients with PKU while blood concentrations of Phe and LNAAs (valine, methionine, isoleucine, leucine, Tyr, histidine, and Trp) were manupulated during two series of amino acid loading experiments (Pietz, Kreis, Rupp, et al., 1999).Brain activity was monitored by EEG spectral analysis.Baseline plasma Phe was approximately 1,000 µmol/L, and brain Phe was approximately 250 µmol/L in both series.Without LNAA supplementation, brain Phe increased to approximately 400 µmol/L after oral Phe loading, and EEG spectral analysis revealed acute disturbances in brain activity.With concurrent LNAA supplementation, Phe influx from blood into brain tissue was completely blocked, and there was no slowing of brain activity.These results could become significant for treatment of patients with PKU.Further studies are warranted to find out whether brain Phe concentration can be lowered during steady state and brain function improved by long-term LNAA supplementation.
Use of ammonia lyase
Phenylalanine ammonia lyase (EC 184.108.40.206) can now be produced through a recombinant approach that degrades Phe to a harmless metabolite (Sarkissian, Shao, Blain, et al., 1999).Attempts have been made to use this enzyme as a substitute for phenylalanine hydroxylase (PAH).Tests in animal models of PKU have shown that ammonia lyase lowered blood levels of Phe when the enzyme was injected or administered orally in order to degrade Phe in the gastrointestinal tract.In the only report of a test using humans (Ambrus, Anthone, Horvath, et al., 1987), ammonia lyase was used to lower the Phe blood level in one patient with PKU by means of a hemodialysis-like procedure.Because of its high invasiveness, this procedure cannot be recommended as a routine treatment for PKU.
- Ambrus CM, Anthone S, Horvath C, Kalghatgi K, Lele AS, Eapen G, et al. Extracorporeal enzyme reactors for depletion of phenylalanine in phenylketonuria.Ann Intern Med 1987;106:531-7.
- Batshaw ML, Valle D, Bessman SP.Unsuccessful treatment of phenylketonuria with tyrosine. J Pediatr 1981;99:159-60.
- Berry HK, Brunner RL, Hunt MM, White PP.Valine, isoleucine, and leucin: a new treatment for phenylketonuria.Am J Dis Child 1990;336:539-43.
- Berry HK, Bofinger MK, Hunt MM, Phillips PJ, Guilfoile MB.Reduction of cerebrospinal fluid phenylalanine after oral administration of valine, isoleucine, and leucine.Pediatr Res 1982;16:751-5.
- Krause W, Epstein C, Averbook A, Dembure P, Elsas L.Phenylalanine alters the mean power frequency of electroencephalograms and plasma L-dopa in treated patients with phenylketonuria.Pediatr Res 1986;20:1112-6.
- Krause W, Halminski M, McDonald L, Dembure P, Salvo R, Freides D, et al.Biochemical and neuropsychological effects of elevated plasma phenylalanine in patients with treated phenylketonuria.A model for the study of phenylalanine and brain function in man.J Clin Invest 1985;75:40-8.
- Leuzzi V, Fois D, Carducci C, Antonozzi I, Trasimeni G.Neuropsychological and neuroradiological (MRI) variations during phenylalanine load: protective effect of valine, leucine, and isoleucine supplementation.J Child Neurol 1997;12:338-40.
- Lou H.Large doses of tryptophan and tyrosine as potential therapeutic alternative to dietary phenylalanine restriction in phenylketonuria.Lancet 1985;2:150-1.
- Lou HC, Lykkelund C, Gerdes AM, Udesen H, Bruhn P.Increased vigilance and dopamine synthesis by large doses of tyrosine or phenylalanine restriction in phenylketonuria.Acta Paediatr Scand 1987;76:560-5.
- Lykkelund C, Nielsen JB, Lou HC, Rasmussen V, Gerdes AM, Christensen E, et al.Increased neurotransmitter biosynthesis in phenylketonuria induced by phenylalanine restriction or by supplementation of unrestricted diet with large amounts of tyrosine.Eur J Pediatr 1988;148:238-45.
- Pardridge WM.Blood-brain barrier carrier-mediated transport and brain metabolism of amino acids. Neurochem Res 1998;23:635-44.
- Pietz J, Kreis R, Rupp A, Mayatepek E, Rating D, Boesch C, et al.Large neutral amino acids block phenylalanine transport into brain tissue in patients with phenylketonuria.J Clin Invest 1999;103:1169-78.
- Pietz J, Landwehr R, Kutscha A, Schmidt H, Sonneville LMJ, Trefz FK.Effect of high-dose tyrosine supplementation in adults with phenylketonuria.J Pediatr 1995;127:1-8.
- Sarkissian CN, Shao Z, Blain F, Peevers R, Su H, Heft R, et al.A different approach to treatment of phenylketonuria: phenylalanine degradation with recombinant phenylalanine ammonia lyase.Proc Natl Acad Sci USA 1999;96:1811-3.
- Smith ML, Hanley WB, Clarke JTR, Klim P, Schoonheyt W, Austin V, et al.Randomized controlled trial of tyrosine supplementation on neuropsychological performance in phenylketonuria.Arch Dis Child 1998;78:116-21.
- Ullrich K, Weglage J, Oberwittler C, Pietsch M, Fünders B, van Eckardstein H, et al.Effect of L-dopa on pattern visual evoked potentials (P-100) and neuropsychological tests in untreated adult patients with phenylketonuria.J Inherit Metab Dis 1994;17:349-52.