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Non-physiological amino acid (NPAA) therapy targeting brain phenylalanine reduction: pilot studies in PAH ENU2 mice

  • Original Article
  • Published:
Journal of Inherited Metabolic Disease

Abstract

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.

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Abbreviations

3-MT:

3-methoxytyramine

5-HIAA:

5-hydroxyindoleacetic acid

5-HT:

Serotonin

5-HTP:

5-hydroxytryptophan

AIB:

Aminoisobutyric acid

BBB:

Blood brain barrier

DA:

Dopamine

DOPAC:

3,4-dihydroxyphenylacetic acid

HVA:

Homovanillic acid

Ile; I:

Isoleucine

L-DOPA:

L-dihydroxyphenylalanine

LAT1:

L-type amino acid transporter

Leu; L:

Leucine

LNAA:

Large neutral amino acid

NL:

DL-norleucine

MAIB:

Methyl-aminoisobutyric acid

Met; M:

Methionine

NB:

2-aminobicyclo(2,2,1)heptane-2-carboxylic acid (2-aminonorbornane)

NPAA:

Non-physiological amino acid

PAH:

Phenylalanine hydroxylase

Phe; F:

Phenylalanine

PKU:

Phenylketonuria

SAH:

S-adenosylhomocysteine

SAMe:

S-adenosylmethionine

Trp; W:

Tryptophan

Tyr; Y:

Tyrosine

Val; V:

Valine

References

  • Aaslestad HG, Larson AD (1964) Bacterial metabolism of 2-methylalanine. J Bacteriol 88:1296–1303

    PubMed  CAS  Google Scholar 

  • Arning E, Bottiglieri T, Sun Q et al (2009) Metabolic profiling in phenylalanine hydroxylase-deficient (Pah-/-) mouse brain reveals decreased amino acid neurotransmitters and preferential alterations of the serotoninergic system. Mol Genet Metab 98:21

    Google Scholar 

  • Babu E, Kanai Y, Chairoungdua A et al (2003) Identification of a novel system L amino acid transporter structurally distinct from heterodimeric amino acid transporters. J Biol Chem 278:43838–43845

    Article  PubMed  CAS  Google Scholar 

  • Bodoy S, Martín L, Zorzano A, Palacín M, Estévez R, Bertran J (2005) Identification of LAT4, a novel amino acid transporter with system L activity. J Biol Chem 280:12002–12011

    Article  PubMed  CAS  Google Scholar 

  • Bröer S, Brookes N (2001) Transfer of glutamine between astrocytes and neurons. J Neurochem 77:705–719

    Article  PubMed  Google Scholar 

  • Choi TB, Pardridge WM (1986) Phenylalanine transport at the human blood–brain barrier. Studies with isolated human brain capillaries. J Biol Chem 261:6536–6541

    PubMed  CAS  Google Scholar 

  • Christensen HN, Streicher JA, Elbinger RL (1948) Effects of feeding individual amino acids upon the distribution of other amino acids between cells and extracellular fluid. J Biol Chem 172:515–524

    PubMed  CAS  Google Scholar 

  • Christensen HN (1990) Role of amino acid transport and countertransport in nutrition and metabolism. Physiol Rev 70:43–77

    PubMed  CAS  Google Scholar 

  • Chrostowski MK, McGonnigal BG, Stabila JP, Padbury JF (2009) LAT-1 expression in pre- and post-implantation embryos and placenta. Placenta 30:270–276

    Article  PubMed  CAS  Google Scholar 

  • Crawhall JC, Purkiss P (1973) Transport of methionine and proline by rat liver slices and the effect of certain hormones. Biochem J 136:15–24

    PubMed  CAS  Google Scholar 

  • Der Garabedian PA, Vermeersch JJ (1987) Candida L-norleucine, leucine:2-oxoglutarate aminotransferase. Purification and properties. Eur J Biochem 167:141–147

    Article  Google Scholar 

  • Dotremont H, Francois B, Diels M, Gillis P (1995) Nutritional value of essential amino acids in the treatment of adults with phenylketonuria. J Inherit Metab Dis 18:127–130

    Article  PubMed  CAS  Google Scholar 

  • Ennis SR, Ren XD, Betz AL (1994) Transport of alpha-aminoisobutyric acid across the blood–brain barrier studied with in situ perfusion of rat brain. Brain Res 643:100–107

    Article  PubMed  CAS  Google Scholar 

  • Enns GM, Koch R, Brumm V, Blakely E, Suter R, Jurecki E (2010) Suboptimal outcomes in patients with PKU treated early with diet alone: revisiting the evidence. Mol Genet Metab 101:99–109

    Article  PubMed  CAS  Google Scholar 

  • Geldenhuys WJ, Van der Schyf CJ (2011) Role of serotonin in Alzheimer’s disease: a new therapeutic target. CNS Drugs 25:765–781

    Article  PubMed  CAS  Google Scholar 

  • Han SJ, Choi S-E, Yi S-A et al (2012) β-Cell-protective effect of 2-aminobicyclo-(2,1,1)-heptane-2-carboxylic acid as a glutamate dehydrogenase activator in db/db mice. J Endocrinol 212:307–315

    Google Scholar 

  • Kanai Y, Segawa H, Miyamoto K, Uchino H, Takeda E, Endou H (1998) Expression cloning and characterization of a transporter for large neutral amino acids activated by the heavy chain of 4F2 antigen (CD98). J Biol Chem 273:23629–23632

    Article  PubMed  CAS  Google Scholar 

  • Knudsen GM, Hasselbalch S, Toft PB, Christensen E, Paulson OB, Lou H (1995) Blood–brain barrier transport of amino acids in healthy controls and in patients with phenylketonuria. J Inherit Metab Dis 18:653–664

    Article  PubMed  CAS  Google Scholar 

  • Kurian MA, Gissen P, Smith M, Heales S Jr, Clayton PT (2011) The monoamine neurotransmitter disorders: an expanding range of neurological syndromes. Lancet Neurol 10:721–733

    Article  PubMed  CAS  Google Scholar 

  • Lepley PR, Mukkada AJ (1983) Characteristics of an uptake system for alpha-aminoisobutyric acid in Leishmania tropica promastigotes. J Protozool 30:41–46

    PubMed  CAS  Google Scholar 

  • Lin J, Raoof DA, Thomas DG et al (2004) L-type amino acid transporter-1 overexpression and melphalan sensitivity in Barrett’s adenocarcinoma. Neoplasia 6:74–84

    Article  PubMed  CAS  Google Scholar 

  • Matalon R, Michals-Matalon K, Bhatia G et al (2006) Large neutral amino acids in the treatment of phenylketonuria (PKU). J Inherit Metab Dis 29:732–738

    Article  PubMed  CAS  Google Scholar 

  • McKean CM, Boggs DE, Peterson NA (1968) The influence of high phenylalanine and tyrosine on the concentrations of essential amino acids in brain. J Neurochem 15:235–241

    Article  PubMed  CAS  Google Scholar 

  • Michals-Matalon K, Bhatia G, Guttler F, Tyring SK, Matalon R (2007) Response of phenylketonuria to tetrahydrobiopterin. J Nutr 137:1564S–1567S

    PubMed  CAS  Google Scholar 

  • Ogburn KD, Bottiglieri T, Wang Z, Figueiredo-Pereira ME (2006) Prostaglandin J2 reduces catechol-O-methyltransferase activity and enhances dopamine toxicity in neuronal cells. Neurobiol Dis 22:294–301

    Article  PubMed  CAS  Google Scholar 

  • Pietz J, Kreis R, Rupp A, Mayatepek E, Rating D, Boesch C, Bremer HJ (1999) Large neutral amino acids block phenylalanine transport into brain tissue in patients with phenylketonuria. J Clin Invest 103:1169–1178

    Article  PubMed  CAS  Google Scholar 

  • Pisoni RL, Flickinger KS, Thoene JG, Christensen HN (1987) Characterization of carrier-mediated transport systems for small neutral amino acids in human fibroblast lysosomes. J Biol Chem 262:6010–6017

    PubMed  CAS  Google Scholar 

  • Segawa H, Fukasawa Y, Miyamoto K, Takeda E, Endou H, Kanai Y (1999) Identification and functional characterization of a Na + -independent neutral amino acid transporter with broad substrate selectivity. J Biol Chem 274:19745–19751

    Article  PubMed  CAS  Google Scholar 

  • Shennan DB, McNeillie SA (1994) Characteristics of alpha-aminoisobutyric acid transport by lactating rat mammary gland. J Dairy Res 61:9–19

    Article  PubMed  Google Scholar 

  • Skvorak KJ, Hager EJ, Arning E et al (2009) Hepatocyte transplantation (HTx) corrects selected neurometabolic abnormalities in murine intermediate maple syrup urine disease (iMSUD). Biochim Biophys Acta 1792:1004–1010

    Article  PubMed  CAS  Google Scholar 

  • Smith QR, Momma S, Aoyagi M, Rapoport SI (1987) Kinetics of neutral amino acid transport across the blood–brain barrier. J Neurochem 49:1651–1658

    Article  PubMed  CAS  Google Scholar 

  • Tews JK, Harper AE (1986) Tissue amino acids in rats fed norleucine, norvaline, homoarginine or other amino acid analogues. J Nutr 116:1464–1472

    PubMed  CAS  Google Scholar 

  • Tews JK, Repa JJ, Harper AE (1990) Norleucine: a branched-chain amino acid analog affecting feeding behavior of rats. Pharmacol Biochem Behav 35:911–921

    Article  PubMed  CAS  Google Scholar 

  • Tews JK, Repa JJ, Harper AE (1991) Branched-chain and other amino acids in tissues of rats fed leucine-limiting amino acid diets containing norleucine. J Nutr 121:364–378

    PubMed  CAS  Google Scholar 

  • Tovar A, Tews JK, Torres N, Harper AE (1988) Some characteristics of threonine transport across the blood–brain barrier of the rat. J Neurochem 51:1285–1293

    Article  PubMed  CAS  Google Scholar 

  • van Spronsen FJ, Enns GM (2010) Future treatment strategies in phenylketonuria. Mol Genet Metab 99:S90–S95

    Article  PubMed  Google Scholar 

  • van Spronsen FJ, de Groot MJ, Hoeksma M, Reijngoud DJ, van Rijn M (2010) Large neutral amino acids in the treatment of PKU: from theory to practice. J Inherit Metab Dis 33:671–676

    Article  PubMed  Google Scholar 

  • Wadhwani KC, Smith QR, Rapoport SI (1990) Facilitated transport of L-phenylalanine across blood-nerve barrier of rat peripheral nerve. Am J Physiol 258:R1436–R1444

    PubMed  CAS  Google Scholar 

  • Zagreda L, Goodman J, Druin DP, McDonald D, Diamond A (1999) Cognitive deficits in a genetic mouse model of the most common biochemical cause of human mental retardation. J Neurosci 19:6175–6182

    PubMed  CAS  Google Scholar 

  • Zinnanti WJ, Lazovic J, Griffin K et al (2009) Dual mechanism of brain injury and novel treatment strategy in maple syrup urine disease. Brain 132:903–918

    Article  PubMed  Google Scholar 

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Acknowledgements

The authors are indebted to Dr. Cary Harding, Oregon Health Sciences University, for providing the Pah enu2 murine model, and to Dr. Kristen J. Skvorak for supplying wild-type subjects for characterization of brain amino acids and monoamines. The guidance and advice of Drs. Viktor Kozich, Harvey Mudd, and William Zinnanti are gratefully acknowledged. This work was supported by a grant from the National PKU Alliance (www.npkua.org), which is gratefully acknowledged.

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Authors and Affiliations

  1. Section of Clinical Pharmacology, College of Pharmacy, Washington State University, 313 Wegner Hall, PO Box 646510, Pullman, WA, 99164-6510, USA

    Kara R. Vogel & K. Michael Gibson

  2. Institute of Metabolic Disease, Baylor Research Institute, Baylor University Medical Center, Dallas, TX, USA

    Erland Arning, Brandi L. Wasek & Teodoro Bottiglieri

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  1. Kara R. Vogel
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  2. Erland Arning
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  3. Brandi L. Wasek
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  4. Teodoro Bottiglieri
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  5. K. Michael Gibson
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Correspondence to K. Michael Gibson.

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Communicated by: Nenad Blau

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Vogel, K.R., Arning, E., Wasek, B.L. et al. Non-physiological amino acid (NPAA) therapy targeting brain phenylalanine reduction: pilot studies in PAH ENU2 mice. J Inherit Metab Dis 36, 513–523 (2013). https://doi.org/10.1007/s10545-012-9524-8

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  • DOI: https://doi.org/10.1007/s10545-012-9524-8

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