Dopamine transporter gene variant that affects expression in human brain is associated with bipolar disorder:
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1 Dopamine transporter gene variant that affects expression in human brain is associated with bipolar disorder: Implications for ADHD subtypes and drug response in autism. Julia Pinsonneault
2 Genetic Contributions to ADHD Subtypes and Markers of Drug Response in Autism J Pinsonneault, E Hurt, W Sadee & M Aman
3 Autism AD: Severe neurodevelopmental disorder characterized by: impaired social interaction impairments in communication restricted repetitive and stereotyped patterns of behavior, interests, and activities AD, PDD-NOS, and Asperger s are frequently referred to as Autism Spectrum Disorders (ASDs). 40% of children with ASD have clinicallysignificant symptoms of ADHD
4 ADHD ADHD is a common childhood disorder that affects between 5 to 7% of typically-developing children Inattentive Hyperactive Combined Psychostimulant medicine (methlyphenidate, amphetamines) is the standard of care non-stimulant alternative is the selective norepinephrine reuptake inhibitor, atomoxetine. Genetics suggest involvement of dopaminergic, the serotonergic and the norepinephrine systems
5 Hypotheses Experimentally determined functional polymorphisms in dopaminergic, serotonergic and noradrenergic genes associate with response to therapeutic drugs in ASD; Such polymorphisms are also linked to expression of ADHD symptoms in ASD.
6 Dopamine Pathways Serotonin Pathways Functions *Reward (motivation) *Pleasure (euphoria) *Motor function (fine tuning) *Compulsion *Perseveration Functions * Mood * Memory processing * Sleep * Cognition National Institutes of Health
7 Dopamine synaptic signaling
8 Drug abuse Disease relationships Attention deficit disorder (ADHD/ADD) Parkinson disease Tourette syndrome Schizophrenia Bipolar disorder DAT genetic variants have been implicated in association studies of these diseases
9 Genetic variation Single nucleotide polymorphisms (SNPs) Polymorphisms that affect RNA processing, expression and stability may account for diversity between individuals
10 DAT1 gene SLC6A Kb on the minus strand of chromosome 5 Exon Kb rs6350 Exon 2 rs rs6347 Exon 9 Intron 8 VNTR rs37022 rs40184 rs & rs rs27072 Exon 15 rs rs VNTR There are no frequent non-synonymous SNPs Test for regulatory polymorphisms with AEI 3 VNTR: The most studied DAT polymorphism
11 Allelic expression imbalance
12 AEI in human substantia nigra using 2 marker SNPs rs6347 rs27072 We scanned many SNPs for AEI association The best associations are with the 2 marker SNPs themselves. Relationship to VNTR is unclear.
13 Constructs tested in cell culture
14 Allelic expression imbalance of paired co-transfected DNAs in cell culture Minor/Major 9/10 repeat rs6347 rs27072
15 The effect of dilution on allelic expression of rs27072 & WT
16 The effect of dilution on DA uptake Significance levels between WT and rs27072 constructs were determined by T test: * 2.93 E-06, ** 3.22 E-14, *** 2.56 E-29
17 The effect of dilution on luciferase reporter activity
18 rs27072 is functional and concentration dependent At dilute concentrations of DNA ~ low mrna mrna Protein activity At high concentrations of DNA ~ high mrna mrna Protein activity
19 Testing in clinical cohorts Stanley Brain Collection 170 DNA samples 50 bipolar, 60 schizophrenic, 60 controls Bipolar Disorder A. Malhotra (Zucker Hillside Hospital, NY) Schizophrenia A. Bertolino (University of Bari)
20 Clinical associations Bipolar and control subjects from Zucker Hillside Hospital
21 Now we have an additional clinical cohort available: The ADHD symptoms in autism study
22 Hypotheses Experimentally determined functional polymorphisms in dopaminergic, serotonergic and noradrenergic genes associate with response to therapeutic drugs in ASD; Such polymorphisms are also linked to expression of ADHD symptoms in ASD.
23 Functional SNPs DAT rs27072, (rs6347, Intron 8, 9/10 VNTR) DRD2 rs228365, rs and rs TPH2 rs , rs MAOA haplotype
24 Candidate Genes COMT, DAO, DISC1, DRD2, DRD3, SLC6A2, SLC6A3, SLC6A4, ESR1, MAOA, MAOB, HTR2A, TPH2, TH, VMAT2 and CHRNA4
25 Autism study design We expected to recruit approximately 40 subjects with ADHD symptoms and 30 subjects without ADHD symptoms in one year from the above studies.
26 Current recruitment status Subjects collected 63/61 ADHD symptoms: 40 Genotyped subjects 57 No 7 SNPs genotyped 60 Unknown 14 Diagnosis: AD 38 Gender M 56 PDD NOS 14 F 5 Asperger's 7 Race W 54 Unknown 2 Other 7
27 Clinical data Gender, Race, Age at baseline Baseline SNAP- a parent completed rating of ADHD symptoms based on a 0 to 3 rating scale: Not at All = 0, Just A Little = 1, Quite A Bit = 2, and Very Much = 3. The items from the DSM-IV(1994) criteria for Attention-Deficit/Hyperactivity Disorder (ADHD) are included for the two subsets of symptoms: inattention and hyperactivity/ impulsivity. Also, items are included from the DSM-IV criteria for Oppositional Defiant Disorder. Follow up SNAP, to measure the response to medication, if any, prescribed for ADHD symptoms. ADHD responders are defined as children who show at least a 30% reduction, as compared to baseline (BL), on the parent-completed SNAP Aberrant Behavior Checklist (ABC)- is a 58-item rating scale developed for persons with developmental disabilities to study treatment effects and for characterization in developmental disabilities. The checklist contains 5 subscales: 1 Irritability; 2 Lethargy; 3 Stereotypy; 4 Hyperactivity; 5--Inappropriate Speech. Autism Diagnostic Interview-Revised (ADI-R) score for diagnosis of Autism or PDD-NOS, or Asperger s Disorder. The ADI-R evaluates behavior in three main areas: qualities of reciprocal social interaction; communication and language; and restricted and repetitive, stereotyped interests and behaviors. Medication, if any and dosage for the treatment of ADHD symptoms
28 ADHD, yes or no, basic allele test Minor Allele Freq. Chi Squared P Corr/Trend Fisher's Marker R Exact P MAOA_rs MAOA_rs TH_rs MAOA_rs MAOA VNTR SLC18A2_rs DRD3_rs SLC18A2_rs SLC6A3_rs ANODEV P
29 ABC- Hyperactivity score, basic allele test Corr/Trend Corr/Trend Marker P R F Test P MAOA_rs MAOA_rs MAOA_rs MAOA_rs DRD2_rs DRD2 rs ESR1_rs MAOB_rs SLC6A3_rs
30 ABC- Hyperactivity score dominant allele test Minor Marker Corr/Trend R F Test P Allele Freq. Call Rate HWE P DRD3_rs SLC6A3_rs DRD2 rs
31 Future work Refine the phenotypic data Drug response data for subjects ADHD symptom status for all subjects CYP2D6 genotype Continuing to collect samples and genotype.
32 Acknowledgements Cara Grantier, Pam Sayre, Mandy Curtis Mike Aman, Wolfgang Sadee, Beth Hurt The project described was supported by Award Number UL1RR from the National Center For Research Resources. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Center For Research Resources or the National Institutes of Health. Additional support for this project was by UO1 GM
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