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%H"RESEARCH UPDATE IN PHENYLKETONURIA##" 9Dr. Maureen Cleary Great Ormond Street Hospital NHS Trust9 o"Blood-brain barrier studies in PKU" Blood-brain barrier studies in PKU Large Neutral amino acids Essential fatty acids supplementation Biopterin treatment Ammonia lyase Gene therapy Nc       <rBlood-brain barrier studies  Phenylketonuria0  Monitor metabolic control by blood phe Preferable to measure phe at site of action (brain) rather than point of delivery (blood)N$    @ #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~a#"Ta#"  T 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 28       )1H-Magnetic Resonance Spectroscopy in PKU**() Non-invasive assessment of changes in brain metabolism Initial reports measure N-Acetyl-Aspartate, NA Choline, inositol, creatinine (8OpZ          )1H-Magnetic Resonance Spectroscopy in PKU**() ; Normal NAA, choline, creatinine Suggests no demyelination$<. \          -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      '        MRS and brain phe$  V1995 Detection and quantitation methodology of brain metabolites in patients with PKU&RR2   6  MRS: normal (   MRS: PKU (  Measurement of phe$  Present in relatively small quantities Cf NAA, choline Need to use  difference spectroscopy i.e. subtract spectra from non PKU controls &'f'fN'     T MRS of brain phe studies in PKU2    b17 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)&*l  C  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~     >  %   Blood- brain relationship        jSuggests Intervariability of brain phe Explains different outcomes Only really explains unusual patients6 <& <&N      C MRS blood:brain0  Pietz et al., (1999) linear blood: brain 4:1 Moller et al.,(2000) saturated at higher phe levels Moats et al. (2000) ?? exponential ZZZZZ!ZZZZZ!j  *  $  0 Blood-brain relationships      Large Neutral amino acids  =s LNAA and PKU  H Large neutral amino acids compete for entry to brain with phenylalanine$;   !Large neutral amino acids and PKU! F Administer large quantities of LNAA and reduce phe entry to the brain20      Large Neutral AminoAcids and PKU!!02     What is the evidence that it should work? Earlier studies (animals or functional testing) Later studies (humans) using Magnetic Resonance Spectroscopy ,+ZqZ+q  "" LNAA and PKU  Cerebral protein synthesis reduced in hyperphe state in rats Improves upon supplementing with LNAA Binek-Singer & Johnson, 1982 &ddF&  6   ##CLNAA and PKU: effect of supplements on brain amino acids in animalsDD$D Rats phe hydroxylase inhibited Phe load Phe load + LNAA LNAA group had lower brain phe and similar blood phe Andersen & Avins, 1976:OO          )        $$!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., 1985L+"I+"I      #        2 ''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 6AXAXj,  .     ((&LNAA, PKU and MRS: Pietz et al. (1999)2   Rise in brain phe occurred after loading This rise blocked when LNAA taken with phe load EEG spectra abnormalities not seen when LNAA ingestedN  A  < )) LNAA study  Brain phe after oral phe load mean preload 252 mean post load 6 hrs 344 mean post load 12 hrs 377 Brain phe after oral phe + LNAA mean preload 226 mean post load 6 hrs 235 mean post load 12 hrs 210 TZKZ ZKZK K~     W     S *7Further considerations Is MRS sufficiently robust tool for intervention studies? What are the relationships between BB phe entry and actual brain tissue phe levels? Na     +8LNAA, MRS and MOUSE PAHENU-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)lZZ6Z +       i   ,9 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  Z 2   -: 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  /pEssential fatty acids in PKU  0ZEssential fatty acids in PKU 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?<ZZZkZZk~,      /   #  1 1qEssential 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 NQ  '    >tPKU and biopterin$    .=PKU 3 Phenylalanine Tyrosine Biopterin metabolism*N        2eBiopterin responsive PKU   3Should we be treating some patients with biopterin?44(*)    3fRole 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           (   &   4gBiopterin in PKU   PRecent observation that biopterin may benefit phe hydroxylase deficient patientsb          5hBiopterin 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 8 ]  ? 6i 7jBiopterin and PKU    Suggest loading test in all patients However newborn failed loading test patients have subsequently been found to be responsive 8kBiopterin and pku8     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*   o 9lAlternative therapiesAmmonia lyase therapy Recombinant phenylalanine ammonia lyase converts phe to trans-cinnamic acid in the gut reduces plasma phe by approx 50% in PKU mouse2 ]t    :m New therapiesAmmonia lyase treatment may be useful needs further studies to test safety may still need some diet may be many years before available&ooy?uPKU and Gene therapy  ;nAlternative therapies GENE THERAPY Adv/RSV-hPAH infused into portal vein of PAHenu2 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 <  ,F  )   @v 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 underwayN"   F   '   0` ` ̙33` 333MMM` ff3333f` f` f` 3>?" dd@,|?" dd@   " @ ` n?" dd@   @@``PR    @ ` ` p>> $(    6~ P ~ T Click to edit Master title style! !  0~  ~ RClick to edit Master text styles Second level Third level Fourth level Fifth level!     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PCr   P <ċh QCho  H P 0޽h ? 3̙33  0 2* T(  Tx T c $ o@    T XAvolspr1pP0P  T 0ر~` p  Qphe 2  T 0 q  QNAA 2 H T 0޽h ? f̙33  0 0X8(  Xx X c $x    X dAtf05040sum3rp  X 0z  ]NAA  2f  X 0  @ Uphe 2f RB X@ s *D@0 H X 0޽h ? f̙33  0 @\0(  \x \ c $臲P   x \ c $  H \ 0޽h ? 3̙33  0 P`0(  `x ` c $P   x ` c $d  H ` 0޽h ? 3̙33  0 `d0(  dx d c $P   x d c $  H d 0޽h ? 3̙33  0 91ph(  hx h c $ԝ   x h c $   LB h c $DP LB h c $DP  P  h 6p  wPietz et al.,(1999)0$$   h 6$P `p D0 RB h s *D0p H h 0޽h ? 3̙339 0 ply(  l l N|gֳgֳ ?` WMagnetic Resonance Spectroscopy   $d l Hxgֳgֳ ?  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)  [  s    [  s  p d       T H l 0޽h ? ̙33 0 7/p(  p p N4gֳgֳ ?` WMagnetic Resonance Spectroscopy   $ p H0gֳgֳ ? :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> \  K    H p 0޽h ? ̙33p  0  t(  tx t c $  H t 0޽h ? 3̙33  0 x6(  x~ x s *乲P   x x c $   H x 0޽h ? ̙33;  0 |{(  |x | c $4Dz   x | c $ Ȳ   LB | c $D@@@ RB | s *D @p LB | c $D @ LB | c $D LB | c $D   LB  | c $DP LB  | c $DP PLB  | c $D@ LB  | c $D@ @ LB  | c $D@ LB | c $D`p | Bв $@ uPietz et al.,(1999)0$   | 6dֲN n vMoller et al.,(2000)0$   | 6ڲP `p hMoats et al., (2000)0 H | 0޽h ? 3̙33v 0 &(  ~  s *p  H  0޽h ? 3̙33  0 $(  r  S P   r  S   H  0޽h ? ff̙33  0 0(  x  c $HP   x  c $   H  0޽h ? 3̙33  0 0(  x  c $P   x  c $t  H  0޽h ? ̙33  0 6(  ~  s *\ P   x  c $4 }  H  0޽h ? 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'>3___PPT10i. p٠+D=' = @B +  0 P<(  ~  s *P   ~  s *\  H  0޽h ? f33  0 `<(  ~  s *P   ~  s *  H  0޽h ? f3̙33  0 p$(  r  S HP   r  S    H  0޽h ? ff̙33  0 0(  x  c $   x  c $P  H  0޽h ? 33̙33  0  $(  r  S ,P   r  S   H  0޽h ? ff̙33r`U^|`ZdRf JhBj @npqsuw:|ă=~ Ƣ0("0 ' ^zHvhnbDfn@Oh+'00 `h   $RESEARCH UPDATE IN PHENYLKETONURIASupportDavid Stening4Microsoft PowerPoint@px@TۼU@2|x^Gg  @  yf--$xx--'@Times New Roman-. $2 0RESEARCH UPDATE IN ."System0-@Times New Roman-. 2 <PHENYLKETONURIA.-@Times New Roman-. f"2 K:Dr. Maureen Cleary.-@Times New Roman-. f@2 V&Great Ormond Street Hospital NHS Trust.-՜.+,0X    On-screen ShowGOSHP 7 ;Times New Roman GaramondArialDefault Design#RESEARCH UPDATE IN PHENYLKETONURIA#Blood-brain barrier studies in PKUBlood-brain barrier studiesPhenylketonuria$1H-Magnetic Resonance Spectroscopy Magnetic Resonance SpectroscopySlide 7*1H-Magnetic Resonance Spectroscopy in PKUSlide 9*1H-Magnetic Resonance Spectroscopy in PKU.PKU and Magnetic Resonance Spectroscopy (MRS)MRS and brain phe Slide 13 MRS: normal MRS: PKUMeasurement of phe MRS of brain phe studies in PKU MRS results Blood- brain relationship Slide 20 Slide 21 Slide 22MRS blood:brainBlood-brain relationshipsLarge Neutral amino acids LNAA and PKU"Large neutral amino acids and PKU!Large Neutral AminoAcids and PKU LNAA and PKUDLNAA and PKU: effect of supplements on brain amino acids in animals"LNAA and cerebral function in PKULNAA, PKU and MRS'LNAA, PKU and MRS: Pietz et al. (1999) LNAA studyFurther considerationsLNAA, MRS and MOUSE Conclusions ConclusionsEssential fatty acids in PKUEssential fatty acids in PKUEssential fatty acids in PKUPKU and biopterinPKUBiopterin responsive PKURole of biopterin in PKUBiopterin in PKUBiopterin in PKU Slide 48Biopterin and PKUBiopterin and pkuAlternative therapiesNew therapiesPKU and Gene therapyAlternative therapies Conclusions  Fonts UsedDesign Template Slide Titles7%_n 0David SteningDavid Stening  !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~     %Root EntrydO)Pictures"Current UserSummaryInformation(PowerPoint Document(DocumentSummaryInformation8