The Story of Phenylketonuria and the Path to Treatment

Genetics Webinar slides - February 8th, 2022

1. The Story of Phenylketonuria and the Path to Treatment Stephanie Sacharow, MD NERGN & Weitzman Institute Genetics Webinar Series 2/8/2022

2. Learning Objectives : Identify the milestones that lead to interest in intellectual disability syndromes and the discovery of PKU Understand how newborn screening began and evolved, and the impact of newborn screening Review the path to treatment and prevention of intellectual disability in affected individuals Discuss conventional dietary therapy and novel therapies for PKU, as well as new treatments in clinical trials

3. Discovery of PKU

4. Discovery of PKU Dr. Asbjørn Følling : Norway, 1934 Ferric chloride test Found phenylpyruvic acid in the urine of siblings with intellectual disability Phenylalanine Tyrosine Phenylpyruvic acid

5. Pearl S. Buck Nobel- and Pulitzer-prize winning novelist Pearl S. Buck wrote about her own daughter, who was disabled by PKU Published 1950

6. Discovery of PKU Horst Bickel, German physician, working in England on PhD in Biochemistry, 1951-2 Implemented PKU diet – during his Pediatric fellowship

7. President Kennedy meets with the McGrath Family

8. Summary of Clinical Features of Untreated PKU Global developmental delay and variable intellectual disability + Autistic features Epilepsy Musty odor Eczema Decreased skin and hair pigmentation Parkinson-like features (particularly in adults) *All avoidable with treatment

9. Robert Guthrie 1958-1962

10. Newborn screening for PKU

11. Metabolic Conditions Tested by Recent NBS Disorders of fatty acid oxidation 2,4-Dienoyl-CoA reductase deficiency Carnitine acylcarnitine translocase deficiency Carnitine palmitoyltransferase I deficiency Carnitine palmitoyltransferase II deficiency Carnitine transport defect Electron transfer flavoprotein deficiency ETF ubiquinone oxidoreductase deficiency Long-chain L-3-OH acyl-CoA dehydrogenase def. Medium-chain acyl-CoA dehydrogenase deficiency Medium-chain L-3-OH acyl-CoA dehydrogenase def. Medium chain ketoacyl -CoA thiolase deficiency Short-chain acyl-CoA dehydrogenase deficiency Trifunctional protein deficiency Very long-chain acyl-CoA dehydrogenase def. Disorders of amino acid metabolism Arginase deficiency Argininosuccinate lyase deficiency Argininosuccinate synthase deficiency Maple syrup urine disease (MSUD) Citrin deficiency Cystathionine  -synthase deficiency Methionine adenosyltransferase deficiency Mitochondrial ornithine trasport defect (HHH) Phenylalanine hydroxylase deficiency (PKU) Defects of biopterin metabolism Fumarylacetoacetase deficiency Tyrosine aminotransferase deficiency Disorders of organic acid metabolism 2-CH3 butyryl -CoA dehydrogenase deficiency 2-CH3 3-OH butyryl -CoA dehydrogenase deficiency 3-OH 3-CH 3 glutaryl -CoA lyase deficiency 3-CH 3 crotonyl -CoA carboxylase deficiency 3-CH 3 glutaconyl -CoA hydratase deficiency Isobutyryl -CoA dehydrogenase deficiency Isovaleryl -CoA dehydrogenase deficiency Glutaryl -CoA dehydrogenase deficiency Malonyl -CoA carboxylase deficiency Methylmalonyl -CoA mutase deficiency Disorders of cobalamin metabolism  - ketothiolase deficiency Multiple carboxylase deficiency Propionyl -CoA carboxylase deficiency Other enzyme disorders Galactosemia Biotinidase Deficiency Pompe Disease (pilot) Mucopolysaccharidosis Type I (Pilot) X-linked Adrenoleukodystrophy (Pilot) Collective incidence 1:2-4,000 newborns Clinical impact (US): Up to 2,000 cases/year

12. Overview of Phenylketonuria (PKU) Phenylalanine Hydroxylase (PAH) deficiency Autosomal recessive Elevated Phe  cognitive impairment and neuropsychiatric symptoms (https://southeastgenetics.org/userfiles/images/PKU%20pathway%20dianne%20-%20adria%20edit3%20-without%20ohh20.png)

13. PKU Cofactor Disorders

14. Phe Neurotoxicity Factors related to toxicity Concentration of Phe Deficiency of other LNAA Untreated PKU  Altered white and gray matter morphology Microcephaly Inhibited growth of the cortex and disrupted myelination A – C , FLAIR ( A ), trace diffusion-weighted ( B ), and ADC av  ( C ) images show extensive white matter abnormalities with restricted diffusion

15. Pathophysiology of PKU: Summary of potential mechanisms of neurocognitive impairment by high phenylalanine concentrations

16. PKU Treatments Dietary intervention Severe restriction of dietary Phe Phe -free amino acid–modified medical foods (https ://southeastgenetics.org/userfiles/images/PKU%20pathway%20dianne%20-% 20adria%20edit3%20-without%20ohh20.png)

17. PKU Diet http://depts.washington.edu/pku/about/diet.html

18. Efficacy of PKU Diet Normal intellectual quotients (I.Q .) I.Q. gap when compared to their non-PKU siblings Lower scores for certain neuropsychological functions Executive function

19. PKU Treatments Cofactor therapy with sapropterin (https ://southeastgenetics.org/userfiles/images/PKU%20pathway%20dianne%20-% 20adria%20edit3%20-without%20ohh20.png)

20. Cofactor therapy - sapropterin dihydrochloride Approved by the FDA - December 2007 Improves Phe tolerance in about 40% of PKU patients Milder genotype are associated with higher s apropterin responsiveness requires trial period to evaluate its effectiveness .

21. LAT-1 Transporter for LNAA Phe is transported into the brain by the L-amino acid transporter 1 ( LAT-1) The binding of a LNAA to a transporter is a competitive process LAT-1 has the highest affinity for Phe Phe  disturbance in neurotransmitter production Large Neutral Amino Acids

22. Glycomacropeptide Glycomacropeptide (GMP) is a whey-based protein that is produced when making cheese. Dietary protein containing a minimal amount of Phe contains 2.5-5 mg Phe /g protein Alternative to the amino acid based medical foods Examples: BetterMilk , PhenylAde GMP

23. Phenylalanine ammonia lyase or “PAL” PAL (Phenylalanine ammonia lyase ) is an injectable plant, fungi, and bacterial enzyme that breaks down Phe PAH (Phenylalanine Hydroxylase) is the enzyme that is not working in PKU patients Enzyme s ubstitution enzyme therapy for PKU FDA-approved 2018 Trademark as Palynziq ( Pegvaliase ) Trans- cinnamic acid Phenylalanine

24. Pegvaliase Phenylalanine Ammonia Lyase (PAL) enzyme Recombinant Anabaena variabilis phenylalanine ammonia lyase Pegylation https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/phenylalanine-ammonia-lyase rAV -PAL Monomer (MW 64,000) Pegvaliase-pqpz ( rAV -PAL–PEG) ~150,000 atoms (MW 1,200,000) Longo N et al. Lancet. 2014;384:37-44. Image credit : Peerview.com

25. Outcomes Overview- PAL Clinic at BCH 45 active patients have been on PAL >1 year 44 show response to treatment Some have fluctuating response (e.g. 27 2.616mg/dl) Most on a regular diet Discontinuations for various reasons adverse events, anxiety, variable response

26. Results: Phe decrease over time PRISM STUDY 261 participants Mean ( SD) blood Phe 1232.7 (386.4) μmol /L at baseline 564.5 (531.2) μmol /L at 12 months 311.4 (427) μmol /L at 24 months Phe decreased from baseline - 51.1% and 68.7%, respectively BCH PAL Clinic 20 study patients at close-out (~30 total) 24 naïve on treatment, 10 reached 12 months on treatment Mean blood Phe 1146 μmol /L at baseline 534 μmol /L at 12 months (for 10 patients who reached 12 months on treatment) 306 μmol /L at lowest point Phe decreased from baseline – 53%

27. Side-effects Data PRISM Study The most common AEs were: Arthralgia (70.5%) Injection-site reaction (62.1%) Injection-site erythema (47.9%) Headache (47.1%) BCH Population The most common AEs were: Injection-site reaction (includes erythema) 23/26 (88%) Arthralgia 18/26 (69%) Rash 18/26 (69%) Fatigue 12/26 (46%) Headache 9/26 (35%) GI symptoms 7/26 (27%) Fever/Chills 6/26 (23%) Hair loss 5/26 (19%) Lymph nodes 4/26 (15%) Dizziness 2/26 (8%) STEROID needed 11/26 (42%) *Auto-injectable epinephrine required due to risk of anaphylaxis

28. Future therapies (in clinical trials)

29. Gene therapy: Genome editing

30. Gene therapy: Gene transfer https://www.nhlbi.nih.gov/health-topics/genetic-therapies

31. Summary of therapeutic approaches for phenylketonuria (PKU) Metabolites 2014 , 4 , 1007-1017; doi:10.3390/metabo4041007

32. Acknowledgements Harvey Levy Program for Phenylketonuria and Related Conditions Olaf Bodamer, MD, PhD, Associate Chief, Division of Genetics and Genomics Gerry Berry, MD, Director of the Metabolism Program Harvey Levy, MD, Emeritus Director of the PKU Program Susan Waisbren, PhD, Emeritus Co-Director of the PKU Program Stephanie Sacharow, MD, Program Director Boston Children’s PAL Clinic Team & Leadership Stephanie Sacharow, MD, Medical Director Ann Wessel, MS, RD, LDN Leslie Martell, MS, RD, LDN Suzanne Hollander, MS, RD, LDN Krista Viau, PhD, RD, CSP Amy Kritzer, MD Benjamin Goodlett, PhD Harvey Levy, MD Fran Rohr, MD, RD, LDN