Disclosure. I have no disclosures to mention

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1 Disclosure I have no disclosures to mention Liberalization of the Dietary Restrictions for PKU Patients: Large Neutral Amino Acids (LNAA) Supplementation in PKU Mice Dr. Eddy A. van der Zee, University of Groningen, The Netherlands PKU preclinical research PKU clinical research

2 Project group Wild-type C57Bl/6 Mouse E.A. van der Zee, PhD, Groningen, Netherlands D. van Vliet, BSc, Groningen, Netherlands V.M. Bruinenberg, MSc, Groningen, Netherlands P. Mazzola, MSc, Groningen, Netherlands F.J. van Spronsen, MD, PhD, Groningen, Netherlands K. Moseley, MSc, RD, Los Angeles, USA M.R. Heiner-Fokkema, PhD, Groningen, Netherlands PKU (Pah -enuh2 ) C57Bl/6 Mouse

3 PKU cause

4 Normal amino acid metabolism normal intelligence normal development food proteins fats carbohydrates proteins amino acids (AA) water protein synthesis amino acid processing neurotransmitter synthesis non-brain tissues protein synthesis

5 Amino acid metabolism in PKU severe mental retardation abnormal development food proteins fats carbohydrates water Phe PAH Tyr elevated plasma Phe >2500 µmol/l N µmol/l

6 Diagnosing PKU 1974 Newborn screening for PKU

7 Current PKU treatment protein restriction (Phe-restriction) no mental retardation almost normal development amino acid mixture Phe PAH Tyr plasma Phe reduction aim: µmol/l

8 Problems in current treatment Cognitive outcome not normal mildly reduced IQ neuropsychological abnormalities increased risk psychiatrial symptoms? Burden of life-long diet non-compliance Need for new treatment strategies

9 Phe transport from blood to brain

10 Drawing of the LAT1 transporter

11 BBB transport of amino acids blood Leu Met Phe Tyr Trp Ile LNAA-t brain Leu Met Phe Ile Tyr Trp Leu Met Phe Ile Trp Met Ile

12 Disturbed LNAA transport in PKU blood Leu Met Phe Tyr Trp Ile LNAA-t Leu Met Phe Ile Tyr Trp brain neurotoxicity??? Leu Met Phe Ile Trp Met Ile

13 Cerebral protein synthesis in PKU R 2 = 0.40 p < 0.01 Hoeksma et al. High plasma phenylalanine decreases cerebral protein synthesis. Mol. Gen. Metab. 2009

14 Our hypothesis plasma Phe elevated LNAA CREB pathway activation reduced CPS BRAIN DAMAGE

15 Synapses, the place where neurons meet

16 Synaptic changes in response to Phe Phe reduces the formation of synapses Without Phe With Phe Synaptic changes in PKU mouse model - reduced density of dendritic spines - widened synaptic cleft - decreased thickness of PSD

17 PKU mouse model Genetics point mutation PAH gene Biochemistry markedly elevated plasma Phe reduced brain LNAA concentrations reduced CPS Biology C57Bl/6 PKU and C57Bl/6 WT mice impaired learning and memory Molecular neurobiology CREB levels reduced synaptic problems

18 Aim of the study LNAA diet treatments: Liberalization of the Dietary Restrictions for PKU Patients: LNAA Supplementation in PKU Mice Large neutral amino acids in the treatment of PKU: from theory to practice. van Spronsen FJ, de Groot MJ, Hoeksma M, Reijngoud DJ, van Rijn M. J Inherit Metab Dis Dec;33(6): PKU-00 PKU-01 PKU-02 PKU-03 PKU-04 PKU-05 PKU-06 high-protein Tyr + Trp (Tyrosine + Tryptophan) LNAA + Thr (Threonine) normal chow (= BASAL DIET) Thr (Threonine) LNAA Thr Leu + Ile (Leucine + Isoleucine) High-protein diet = isocaloric and isonitrogenic with LNAA diets

19 What we will measure 1. Blood and brain Phe levels 2. Brain non-phe LNAA concentrations 3. Brain tyrosine and tryptophan concentrations 4. Brain proteins important for synaptic plasticity 5. Cerebral protein synthesis indirectly via CREB/pCREB 6. Neurotransmitters and their rate limiting enzymes Westernblot analyses Immunocytochemical analyses

20 Where are we with the experiments? PKU-00 PKU-01 PKU-02 PKU-03 PKU-04 PKU-05 PKU-06 high-protein Tyr + Trp LNAA + Thr normal chow Thr LNAA Thr Leu + Ile Mice on diet grow similarly as mice on normal chow Start LNAA treatment for 6 weeks normal chow high-protein LNAA - Thr LNAA + Thr Threonine Tyr + Trp Leu + Ile WT normal chow male female male female male female male female male female male female male female male female Needed Included Completed

21 Later this year... The final outcomes of the LNAA diet treatments: We anticipate and hope that it will lead to the development of a new LNAA treatment strategy for PKU patients

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