Antonio M. Persico Unit of Child & Adolescent Neuropsichiatry Lab. of Mol. Psychiatry & Neurogenetics University “Campus Bio-Medico” , Roma & Mafalda Luce Center for Pervasive Developmental Disorders, Milan

Translational pediatric psychopharmacology

27th ECNP Congress Berlin (Germany) - October 20, 2014 Priority list for drug development in pediatric psychopharmacology

12

10

8

6

4

2

0

TNM expert meeting, Child Psychopharmacology Network, ECNP 2013 Translational pediatric psychopharmacology

Patient Patients (genetic syndrome) (different syndromes, similar mechanisms)

Drug development

Molecular models Cellular models Animal models Chemical drug modelling Bioinformatic analysis Induced Pluripotent Stem Cells (iPSCs) in personalized molecular medicine c-Myc, Sox2, Oct ¾, KLF4

Differentiated cells iPS cells Syndrome Pathophysiology Drug Therapeutic target Clinical trials by NCT n. Rett syndrome [MeCP2] Abnormal regulation of gene expression, 01253317, 01777542 impairing neuritic sprouting and synaptogenesis (1-3) IGF1 Enhance neuritic sprouting 22q13 deletion/Phelan-McDermid Disrupted scaffolding of the post-synaptic [Mecasermin, Increlex] and synaptogenesis 01525901 Syndrome [SHANK3] elements, leading to reduced dendritic spines and synaptogenesis Fragile X syndrome [FMR1] Increased translation in dendritic spines MPEP mGLUR5 antagonism None Fenobam 01806415 STX107 01325740, 00965432 AFQ056 [Mavoglurant] 01357239, 01253629, 01482143, 01348087,

01433354, 00718341 RO4917523 01750957, 01015430, 01517698 STX209 [Arbaclofen] GABA-B receptor 00788073, 01282268, agonism 01555333 (terminated), 01325220 CX516 [Ampalex] Positive allosteric 00054730 modulation of AMPA receptors Fragile X syndrome and idiopathic Microglial activation Microglial inhibition 00409747 autism [neuroinflammation]. Minocycline Increased expression and activity of MMP9 MMP9 inhibition 01053156, 0858689 Tuberous Sclerosis [TSC1/TSC2] Disinhibition of the mTOR pathway Rapamycin [Sirolimus] mTOR inhibition 00457808

Everolimus [RAD001, 01289912, 01070316, Afinitor] 01730209, 01713946 Autism with macrocephaly (PTEN) None Neurofibromatosis (NF1) Disinhibition of RAS activity & mTOR Lovastatin Ras activity inhibition 00352599 pathway Genetic variants in OXTR Inadequate action of Oxytoxin Oxytocin Enhance Oxytocin activity 01337687, 01788072, 01624194, 01308749, 01183221, 1256060 CNVs affecting 15q11-13 implicating Excitatory effect GABAergic neurons due to Bumetanide Reinforcement of 01078714 GABRB3, GABRA5 and GABRG3 abnormally elevated intracellular chloride GABAergic inhibition via reduction of intracellular chloride levels Vorstman et al, Psychopharmacol, 231:1063-78, 2014 PNAS 106: 2029-34, 2009 PNAS 111: 4596-601, 2014

N=12, 4-wk multiple ascending dose (40-120 mg/kg twice daily) and an open-label 20-wk extension at the maximum dose PTEN inactivation yields tumors, overgrowth, and autism or intellectual disability Ref. Mut. carriers De novo Clinical phenotype mutations Butler et al., 3/18 (13m, 5f) with H93R (exon 4) Extreme macrocephaly 2005 macrocephaly D252G (exon 7) and macrosomy 16,6% F241S (exon 7)

Butler MG et al, J Med Genet, 42:318, 2005 Kwon CH et al, Neuron, 50:377, 2006 PTEN and the mTOR pathway

mRNA translation Cell proliferation Ma & Blenis, Mol Cell Biol 10:307, 2009 Rapamycin recovers the PTEN -/- phenotype

Zhou J et al, J Neurosci 29:1773, 2009 Rapamycin recovers the PTEN -/- phenotype

Zhou J et al, J Neurosci 29:1773, 2009 Everolimus has been approved for TS

Renal angiomyolipomas

Subependymal giant-cell astrocytomas

Treatment-refractory seizures Facial angiofibromas The pathophysiology of fragile X syndrome

Levenga et al, Trends Mol Med 16:516, 2010 Jacquemont et al, Psychopharmacol 231:1237, 2014 The challenges of clinical trials in fragile X syndrome Challenges and limitations Solutions 1) Patient heterogeneity Use of genetic and epigenetic biomarkers to stratify patients 2) Lack of reliable markers to according to underlying predict response & severe mechanism(s) and target drug side effects therapy 3) Outcome measures Modify existing scales, create display low sensitivity new ad hoc scales, choose appropriate clinical endpoints 4) Short trial duration Longer trial duration, more complex designs including drug 5) Pharmacological + non-pharmacological intervention only intervention combined Jacquemont et al, Psychopharmacol 231:1237, 2014 Scales sensitive to change in FXS studies

Jacquemont et al, Psychopharmacol 231:1237, 2014 Biomarker panels for targeted therapies

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36

Metabolomics

Proteomics

Transcriptomics (coding & non)

Methylomics

Genomics: CNVs, SNPs, repeats

Biological foundations of neurodevelopmental disorders Early intensive behavioral treatment for children aged < 2 years and ½: 20 hrs/wk (2 hrs twice a day x 5 days/wk x 2 yrs) + parent training

Mullen Scale for Early Learning Vineland Adaptive Behavior Scale Severe adverse reactions in cognitive disability

100

/ml) ng

( 80

60

levels

40

blood

20 Prolactin 6+6 5+5 4+4 2+2 1+1 Risperidone dosage in drops (1 mg = 17 drops) Array-CGH in NHS child psychiatry • Blood drawn from 334 families (3 families/wk) • Diagnostic report provided to 124 families (100 simplex e 24 multiplex) • Patients N=147

1 Autism Cognitive disability 2 Genetic syndrome tbd Learning disability 3 Language disorder ADHD Childhood SCZ 7 Developmental delay Memory deficits Depression 8 Dyspraxia Catatonia & epilepsy Bipolar & epilepsy 14 95 Rett syndrome Neurofibromatosis Multiple sclerosis DESR Binge eating Polymicrogyria Prader-Willi Array-CGH outcome (N=147 patients)

No variant 9% Certainly causal CNV 15% Common Negative Positive variant 14 22 51/147 60/147 25% 37 (34%) 38 (41%) 36 Probably causal CNV 26%

Possibly causal CNV of uncertain significance 25% V.M., 23 y.o., autism, cognitive disability, epilepsy

Crom Banda Inizio (bp) Fine (bp) Lunghezza (bp) Sonde Dup/Del Log2 ratio Geni Origine Dup totali in DGV Dup simili in DGV Del totali in DGV Del simili in DGV PRAMEF1, PRAMEF11, LOC649330, 1 p36.21 12.846.934 12.912.625 65.691 3 Del -1,303 HNRNPCL1 Comune ereditata dalla madre 38 17 50 15 RNVU1-18;RNU1-27P;RNU1- 28P;RNU1-3;RNU1-2;RNU1-1;RNU1- 1 p36.13 17.051.180 17.257.997 6 0,585 4,LOC100132147;LOC101927806;LOC4 206.817 Dup 40570,CROCC Comune ereditata dal padre 27 5 26 4 1 q21.3 152.556.449 152.586.281 29.832 3 Del -1,097 LCE3C Comune ereditata dalla madre 6 5 26 22 1 q31.3 196.386.440 196.711.149 324.709 26 Dup 0,296 KCNT2, CFH De novo 21 0 41 0 LOC101930107,LINC00152;MIR4435- 2 p11.2 87.370.476 88.018.726 9 0,418 648.250 Dup 1HG Comune ereditata dal padre 56 4 28 3 3 q26.1 162.556.223 162.594.653 38.430 3 Dup 5,840 Comune ereditata dal padre 13 10 22 21 4 q13.2 69.392.545 69.462.438 69.893 6 Del -3,290 UGT2B17, UGT2B15 Comune ereditata dalla madre 23 14 39 15 OCLN, GTF2H2C, GTF2H2D, LOC100272216, GUSBP3, SERF1A, SERF1B, SMN1, SMN2, LOC100170939, GTF2H2B, SMA5, 5 q13.2 68.849.594 70.369.959 1.520.365 3 Del -1,045 LOC100049076, NAIP, GTF2H2 Comune ereditata dalla madre 113 4 104 5 5 q33.1 151.792.610 151.839.164 46.554 3 Dup 0,955 Rara ereditata parzialmente dal padre 1 0 2 1 6 q14.1 78.979.172 79.023.328 44.156 3 Dup -0,798 Comune ereditata dalla madre 22 17 41 26 8 p11.22 39.237.438 39.374.789 137.351 12 Del -3,461 ADAM5P, ADAM3A Comune ereditata dal padre 26 11 38 23 11 q11 55.372.753 55.453.023 80.270 8 Del 0,585 OR4C6,OR4P4,OR4S2 Comune presente in ambedue i genitori 18 11 53 25 RNF10, POP5, CABP1, MLEC, 12 q24.31 121.006.068 121.273.301 267.233 25 Dup 0,481 UNC119B, ACADS, SPPL3 Rara ereditata dal padre 2 1 9 0 OR11H12, POTEG, POTEM, OR11H2, OR4Q3, OR4M1, OR4N2, OR4K2, 14 q11.2 19.376.762 20.427.242 1.050.480 15 Dup 0,441 OR4K5, OR4K1 De novo 82 3 58 5 14 q11.2 22.368.864 22.964.922 596.058 53 Dup -0,286 Rara ereditata dal padre 20 0 72 2 HERC2P3,GOLGA6L6,MIR1268A,OR4 q11.1- N4,GOLGA8CP,POTEB2;LOC10028896 15 20.394.220 22.669.111 55 0,585 q11.2 6;POTEB;POTED,POTEB2;POTEB,LOC 2.274.891 Del 646214,OR4M2 Comune ereditata dal padre 200 9 197 5 15 q13.3 31.972.646 32.438.943 466.297 26 Dup 0,400 CHRNA7 Comune ereditata dal padre 37 11 27 5 TP53TG3, TP53TG3B, LOC653550, 16 p11.2 32.573.808 33.961.233 1.387.425 17 Dup -0,608 SLC6A10P, LOC390705 Comune ereditata dalla madre 150 3 200 4 17 q12 34.437.475 34.475.514 38.039 4 Del -0,742 Comune ereditata dal padre 34 22 10 8 17 q21.31 44.221.743 44.345.038 123.295 8 Del -0,558 KIAA1267, LOC644246 Comune presente in ambedue i genitori 30 19 15 9 IGF2BP1, B4GALNT2, GNGT2, ABI3, 17 q21.32 47.069.742 47.318.413 248.671 16 Del 0,538 PHOSPHO1 Rara ereditata dal padre 2 0 13 0 22 q11.23 24.347.959 24.390.254 42.295 5 Dup 0,874 LOC391322, GSTT1, GSTTP2 Comune ereditata dal padre 19 18 33 19 X q28 154.396.991 154.425.684 28.693 3 Dup -4,572 Rara ereditata dalla madre 5 5 3 1 D.M., 20 y.o., high functioning autism

Crom Banda Inizio (bp) Fine (bp) Lunghezza (bp) Sonde Dup/Del Log2 ratio Geni Origine Dup totali in DGV Dup simili in DGV Del totali in DGV Del simili in DGV RNVU1-18RNU1-27PRNU1-28PRNU1-3RNU1-2RNU1-1RNU1- p36.13 17.051.180 17.257.997 6 0,585 1 206.817 Dup 4,LOC100132147LOC101927806LOC440570,CROCC Comune ereditata dal padre 27 5 26 4 2 p11.2 87.370.476 88.018.726 648.250 9 Dup 0,418 LOC101930107,LINC00152MIR4435-1HG Comune ereditata dal padre 56 4 28 3 4 q13.2 69.392.545 69.462.438 69.893 6 Del -3,134 UGT2B17, UGT2B15 Comune ereditata dalla madre 23 14 39 15 5 p15.33 715.757 806.629 90.872 3 Dup 0,585 ZDHHC11,ZDHHC11B Comune ereditata dalla madre 52 29 42 20

OCLN, GTF2H2C, GTF2H2D, LOC100272216, GUSBP3, SERF1A, SERF1B, SMN1, SMN2, LOC100170939, GTF2H2B, SMA5, LOC100049076, NAIP, 5 q13.2 68.849.594 70.369.959 1.520.365 3 Del -1,032 GTF2H2 Comune ereditata dalla madre 113 4 104 5 5 q33.1 151.792.610 151.839.164 46.554 3 Dup 0,874 Rara ereditata parzialmente dal padre 1 0 2 1 6 q14.1 78.979.172 79.023.328 44.156 3 Del -0,835 Comune ereditata dalla madre 22 17 41 26 8 p11.22 39.237.438 39.374.789 137.351 12 Del -3,530 ADAM5P, ADAM3A Comune ereditata dal padre 26 11 38 23 11 q11 55.372.753 55.453.023 80.270 8 Dup 0,585 OR4C6,OR4P4,OR4S2 Comune presente in ambedue i genitori 18 11 53 25 12 q24.31 121.006.068 121.273.301 267.233 25 Dup 0,479 RNF10, POP5, CABP1, MLEC, UNC119B, ACADS, SPPL3 Rara ereditata dal padre 2 1 9 0 OR11H12, POTEG, POTEM, OR11H2, OR4Q3, OR4M1, OR4N2, OR4K2, 14 q11.2 19.376.762 20.414.232 1.037.470 14 Dup 0,486 OR4K5, OR4K1 De novo 81 3 58 5 HERC2P3,GOLGA6L6,MIR1268A,OR4N4,GOLGA8CP,POTEB2LOC10028 0,585 15 q11.1-q11.2 20.394.220 22.669.111 2.274.891 55 Dup 8966POTEBPOTED,POTEB2POTEB,LOC646214,OR4M2 Comune ereditata dal padre 200 9 197 5 15 q13.2 30.943.903 31.004.749 60.846 4 Del -0,637 Comune non presente nei genitori 16 13 16 7 15 q13.3 32.021.733 32.510.863 489.130 27 Dup 0,466 CHRNA7 Comune ereditata dal padre 51 10 45 5 16 p11.2 32.573.808 33.625.989 1.052.181 15 Del -0,633 TP53TG3, TP53TG3B, LOC653550, SLC6A10P, LOC390705 Comune ereditata dalla madre 121 7 131 4 17 q12 34.437.475 34.475.514 38.039 4 Del -0,692 Comune ereditata dal padre 34 22 10 8 17 q21.31 44.221.743 44.345.038 123.295 8 Del -0,568 KIAA1267, LOC644246 Comune presente in ambedue i genitori 30 19 15 9 17 q21.32 47.069.742 47.304.479 234.737 15 Dup 0,526 IGF2BP1, B4GALNT2, GNGT2, ABI3, PHOSPHO1 Rara ereditata dal padre 2 0 13 0 22 q11.23 24.347.959 24.390.254 42.295 5 Dup 0,857 LOC391322, GSTT1, GSTTP2 Comune ereditata dal padre 19 18 33 19 X q28 154.396.991 154.425.684 28.693 3 Del -4,729 Rara ereditata dalla madre 5 5 3 1 Biomarkers and molecular psychopharmacology in Autism Spectrum Disorder

Biomarker panel

Vorstman et al, Psychopharmacol, 231:1063-78, 2014 Pathophysiology-driven Child Psychopharmacology

Adult-derived & non-specific Comorbidities psychopharmacology

Personalized Core symptoms molecular drug therapy Thank you!

Carla Lintas Roberto Sacco Antonio M. Persico

Valerio Napolioni Stefano Gabriele

Sarah F. Hastings Ignazio S. Piras