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RESEARCH UPDATE IN PHENYLKETONURIA Dr. Maureen Cleary Great Ormond Street Hospital NHS Trust.

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Presentation on theme: "RESEARCH UPDATE IN PHENYLKETONURIA Dr. Maureen Cleary Great Ormond Street Hospital NHS Trust."— Presentation transcript:

1 RESEARCH UPDATE IN PHENYLKETONURIA Dr. Maureen Cleary Great Ormond Street Hospital NHS Trust

2 Blood-brain barrier studies in PKU Large Neutral amino acids Essential fatty acids supplementation Biopterin treatment Ammonia lyase Gene therapy

3 Blood-brain barrier studies

4 Phenylketonuria Monitor metabolic control by blood phe Preferable to measure phe at site of action (brain) rather than point of delivery (blood)

5 1H-Magnetic Resonance Spectroscopy Nucleus is magnetic –Magnetic field causes all the magnetic nuclei to align themselves to the major axis of the field A second magnetic field is applied –Nuclei tilted to a specific angle When field removed they re-align themselves to the major axis of the magnetic field

6 Magnetic Resonance Spectroscopy capable of identifying different molecules Same nuclei eg protons experience different local magnetic fields Give rise to different MR spectra Area under peak proportional to concentration

7 NAA Cr Cho

8 1H-Magnetic Resonance Spectroscopy in PKU Non-invasive assessment of changes in brain metabolism Initial reports measure N-Acetyl-Aspartate, NA Choline, inositol, creatinine


10 1H-Magnetic Resonance Spectroscopy in PKU Normal NAA, choline, creatinine Suggests no demyelination

11 PKU and Magnetic Resonance Spectroscopy (MRS) Rabbit made hyperphe MRS detected ‘phe’ peak Intensity correlated with brain phe on postmortem Correlated poorly with plasma phe

12 MRS and brain phe 1995 –Detection and quantitation methodology of brain metabolites in patients with PKU

13 NAA Cr Cho

14 MRS: normal phe NAA

15 MRS: PKU NAA phe

16 Measurement of phe Present in relatively small quantities –Cf NAA, choline –Need to use ‘difference spectroscopy’ –i.e. subtract spectra from non PKU controls

17 MRS of brain phe studies in PKU 17 PKU (mean age 25.8 yrs) 10 healthy controls (25.3) Early treated 6 off diet, 3 protein restricted, 8 on aa supp (stopped 2 weeks pre-scan) ‘steady state’ »(Rupp et al., 2001)

18 MRS results Control brain phe mean 0.05, sd 0.025 Blood versus brain linear relationship Blood to brain phe: 4:1 Measurement error 0.03mol/kg ww

19 Blood- brain relationship Pietz et al.,(1999)

20 Magnetic Resonance Spectroscopy Weglage et al., 1998 two siblings aged 17 and 30 yrs early treated R408W/R408W IQ’s 90 and 77 oral load phe max brain phe 12-23 hrs post phe load sib I blood 2448: brain 642 (IQ 90) sib II blood 2316: brain 804 (IQ 77)

21 Magnetic Resonance Spectroscopy Weglage et al., 1998 4 untreated adults two IQ unobtainable ages 34 and 28 yrs blood 1320,1211/ brain 650,670 two IQ 100 and 105 ages 33 and 31 yrs blood 1200, 1210/ brain <200, <200

22 Suggests –Intervariability of brain phe –Explains different outcomes Only really explains unusual patients

23 MRS blood:brain Pietz et al., (1999) –linear blood: brain 4:1 Moller et al.,(2000) –saturated at higher phe levels Moats et al. (2000) –?? exponential

24 Blood-brain relationships Pietz et al.,(1999) Moller et al.,(2000) Moats et al., (2000)

25 Large Neutral amino acids

26 LNAA and PKU Large neutral amino acids compete for entry to brain with phenylalanine

27 Large neutral amino acids and PKU Administer large quantities of LNAA and reduce phe entry to the brain

28 Large Neutral AminoAcids and PKU What is the evidence that it should work? –Earlier studies (animals or functional testing) –Later studies (humans) using Magnetic Resonance Spectroscopy

29 LNAA and PKU Cerebral protein synthesis reduced in hyperphe state in rats Improves upon supplementing with LNAA –Binek-Singer & Johnson, 1982

30 LNAA and PKU: effect of supplements on brain amino acids in animals Rats phe hydroxylase inhibited –Phe load –Phe load + LNAA –LNAA group had lower brain phe and similar blood phe Andersen & Avins, 1976

31 LNAA and cerebral function in PKU Valine, isoleucine and leucine supplements –Reduced brain and CSF phe in rats Six patients with pku improved neuropsych performance whilst taking VIL –Berry et al., 1985

32 LNAA, PKU and MRS Later studies using MRS in humans One study Pietz et al., (1999) –Six adults –Loading with oral phe 100mg/kg –Loading with oral phe plus LNAA –EEG testing

33 LNAA, PKU and MRS: Pietz et al. (1999) Rise in brain phe occurred after loading This rise blocked when LNAA taken with phe load EEG spectra abnormalities not seen when LNAA ingested

34 LNAA study Brain phe after oral phe load –mean preload 252 –mean post load 6 hrs 344 –mean post load 12 hrs377 Brain phe after oral phe + LNAA –mean preload226 –mean post load 6 hrs 235 –mean post load 12 hrs210

35 Further considerations Is MRS sufficiently robust tool for intervention studies? What are the relationships between BB phe entry and actual brain tissue phe levels?

36 LNAA, MRS and MOUSE PAH ENU-2 mouse model –0.5g/kg or 1.0 g/kg PreKUnil –Reduction in blood phe and brain phe –Spectroscopy on homogenized mouse brain –BCAT activity increased on LNAA ( only two mice in each group) Matalon et al, (2003)

37 Conclusions MRS can define a peak which is markedly elevated in individuals with PKU compared to normal spectra MRS can show reduction in this peak when interventions occur such as LNAA application MRS can show some unusual individuals who have low brain phe and are ‘protected’

38 Conclusions Blood:brain barrier relationship not clear Extent of inter-individual variability not clear Safety of long term LNAA not proven To use the technique in dynamic studies need clarity of these changes through the day

39 Essential fatty acids in PKU

40 Diet low in animal protein –low intake alpha-linolenic acid –low docoshexanoic acid –importance in brain cell membrane Infant aminoacid formulae can be supplemented with PUFA’s Should children’s formulae also be supplemented?

41 Essential fatty acids in PKU AA product supplemented with fatty acids Children had higher levels of DHA than unsupplemented group Considered more palatable than unsupplemented formula

42 PKU and biopterin

43 PKU Phenylalanine Tyrosine Biopterin metabolism

44 Biopterin responsive PKU Should we be treating some patients with biopterin?

45 Role of biopterin in PKU Biopterin co-factor for phe hydroxylase Inborn errors of biopterin detected by PKU screening programme On biopterin those patient usually no longer need phe restriction

46 Biopterin in PKU Recent observation that biopterin may benefit phe hydroxylase deficient patients

47 Biopterin in PKU Hyperphe rather than classical PKU Mutations with residual activity Frequently (but not exclusively) missense mutations within the coding region for the catalytic domain


49 Biopterin and PKU Suggest loading test in all patients However newborn failed loading test patients have subsequently been found to be responsive

50 Biopterin and pku Cost of diet v. cost of biopterin Who would benefit? Does it benefit those with severe PKU? Is it safe in pregnancy? Trial later this year 2004

51 Alternative therapies Ammonia lyase therapy Recombinant phenylalanine ammonia lyase converts phe to trans-cinnamic acid in the gut reduces plasma phe by approx 50% in PKU mouse

52 New therapies Ammonia lyase treatment –may be useful –needs further studies to test safety –may still need some diet –may be many years before available

53 PKU and Gene therapy

54 Alternative therapies GENE THERAPY –Adv/RSV-hPAH infused into portal vein of PAH enu2 mice –phe levels normalised with sufficient dose –comparable to 10-20% enzyme activity –successful only in short term –could not be duplicated due to immune response to vector

55 Conclusions Research is fairly active in PKU Biopterin trial will find some individuals with milder PKU who may benefit form Biopterin treatment Enzyme treatment is underway

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