Study Report
Basic Info
| Reference |
Kent L, 200212232786
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| Citation |
Kent L., Doerry U., Hardy E., Parmar R., Gingell K., Hawi Z., Kirley A., Lowe N., Fitzgerald M., Gill M. and Craddock N. (2002) "Evidence that variation at the serotonin transporter gene influences susceptibility to attention deficit hyperactivity disorder (ADHD): analysis and pooled analysis." Mol Psychiatry, 7(8): 908-12.
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| Study Design |
family-based |
| Study Type |
Candidate-gene association study |
| Sample Size |
113 trios |
| Predominant Ethnicity |
Caucasian |
| Population |
United Kingdom, Ireland |
| Gender |
103 (91%) male and 10 (9%) female |
| Age Group |
Children/Adolescents
:
aged 5-16 years, mean age 11.1 years (SD=3.0)
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Detail Info
| Summary |
Two published studies have implicated variation at a polymorphism in the promoter of the serotonin transporter (5HTT; hSERT) in influencing susceptibility to ADHD. Consistent with these results, this study also found a trend for the long allele of the promoter polymorphism to influence susceptibility to ADHD in a sample of 113 ADHD parent proband trios (P=0.13). A pooled analysis of current, and the published results demonstrated a significant over representation of the long allele of the promoter in ADHD probands compared to controls (P=0.008). They also examined two other 5HTT polymorphisms (the VNTR in intron 2 and the 3' UTR SNP). TDT analysis demonstrated preferential transmission of the T allele of the 3' UTR SNP (P=0.04). In addition, ETDT analysis of haplotypes demonstrated significant preferential transmission of haplotypes containing the T allele of the 3' UTR SNP with the long allele of the promoter polymorphism (P=0.004) and the 10 repeat of the VNTR (P=0.03). This study provides further evidence for the possible involvement of the serotonin transporter in susceptibility to ADHD. |
| Total Sample |
Both the full sample of 113 trios and the subset of 90 trios who had been interviewed with the CAPA were analysed. Probands were Caucasian, born in the United Kingdom or Ireland. Of the 113 probands, 91 (81%) were combined type, nine (8%) were inattentive subtype and 13 (11%) were hyperactive/impulsive subtype. In terms of comorbidity 44 (39%) were comorbid for oppositional defiant disorder, and 17(15%) for conduct disorder. |
| Sample Collection |
This collaborative study was approved by the relevant research ethics committees. Written informed consent was obtained from parents, and children either provided written informed consent or assent. The sample was recruited from child psychiatry clinics in Birmingham, United Kingdom and Ireland and consisted of 113 (60 from Ireland, 53 from Birmingham) parent-proband trios. |
| Diagnosis Description |
All probands in this study fulfilled DSM-IV6 diagnostic criteria for ADHD. All of the Birmingham sample and 62% of the Irish sample were interviewed employing the Child and Adolescent Psychiatric Assessment (CAPA). Consistent interview procedures were employed across the two centres, with researchers from each centre receiving a common training in the use of the CAPA. The remaining 38% of the Irish sample met criteria for DSM-IV ADHD following consensus diagnosis according to procedures published previously. Children with a significant learning disability, autistic spectrum disorder or significant medical condition such as epilepsy were excluded. |
| Technique |
Up to 20 ml of venous blood or a cheek swab was obtained from each subject. Genomic DNA was extracted from either venous blood or cheek swabs according to standard procedures. For the promoter polymorphism, touchdown PCR amplification was performed. The PCR products were visualised on 2% agarose gel: a short allele of 484 bp and a long allele of 528 bp. The VNTR polymorphism amplification was performed by PCR. The PCR products were visualised on 2.5% agarose gel: a 9-repeat allele of 250 bp, a 10-repeat allele of 267 bp and a 12-repeat allele of 300 bp. The 3' UTR G/T SNP was amplified with a touch down procedure. The product (208 bp) was digested for 3h with MseI and the resulting polymorphic fragments were separated on a 3% sea agarose gel into alleles G (27 bp and 181 bp) and T (27 bp, 52 bp and 129 bp). |
| Analysis Method |
Genotype frequencies for each polymorphism were compared to frequencies expected under Hardy¨CWeinberg equilibrium using a X2 goodness-of-fit test. The Transmission Disequilibrium Test and the extended Transmission Disequilibrium Test (ETDT) was employed to test for evidence of distorted transmission. Haplotypes between marker pairs were constructed where phase could be determined. Evidence for linkage disequilibrium between markers was examined by X2 analysis in addition to calculating D, the coefficient of disequilibrium and D', the normalised disequilibrium coefficient. Pooled analysis was performed according to the method described by Woolf. |
| Result Description |
Genotype frequencies in parents did not demonstrate any significant departure from Hardy-Weinberg equilibrium and were similar for previously published studies. Each possible polymorphism pair demonstrated highly significant moderate LD. TDT analysis demonstrated a trend associated between the long allele of the promoter polymorphism and ADHD in the expected direction (one-tailed P=0.07) but this does not reach statistical significance. Results for the independent trios demonstrate significant preferential transmission of the T allele of the 3' UTR SNP (P=0.04). There was nominally significant preferential transmission of two haplotypes containing the 3' UTR T allele: the 10-repeat VNTR/T allele of the 3' UTR haplotype (uncorrected P=0.05) and the long promoter/T allele of the 3' UTR haplotype (uncorrected P=0.0007). Significant preferential transmission of the 10-repeat VNTR/long promoter haplotype was demonstrated (uncorrected P=0.02), but the overall allele-wise TDT result for this haplotype was not significant. After correction, the promoter/3' UTR haplotype transmission remains significant. The haplotype consisting of the long promoter allele-10 repeat VNTR allele-T allele 3' UTR was preferentially transmitted (P=0.006) based on HAPMAX. Pooled analysis demonstrated a significant over representation of the long/long genotype in the ADHD population, and under representation of the short/short genotype with an overall odds ratio of 1.33 (95% CI=1.06-1.66, P=0.01). Allele frequencies also demonstrate an over representation of the long allele in the probands (P=0.008). |
Other variant reported by this study (count: 3)
| Variant Name |
Allele Change |
Risk Allele |
Statistical Values |
Author Comments |
Result of Statistical Analysis |
| 5HTTLPR |
short/long |
long |
TDT P-value>0.05
TDT P-value>0.05
|
No significant evidence was found for biased transmission |
Non-significant
|
| SLC6A4 3'-UTR G/T |
G/T |
T |
TDT P-value=0.04
TDT P-value=0.04
|
significant preferential transmission of the T allele |
Significant
|
| SLC6A4 intron2 VNTR |
9, 10, 12 repeat |
10 repeat |
TDT P-value>0.05
TDT P-value>0.05
|
No significant evidence was found for biased transmission |
Non-significant
|
Genes reported by this study (count: 1)
| Gene |
Statistical Values/Author Comments |
Result of Statistical Analysis |
| SLC6A4 |
ETDT haplotype analysis: P=0.004 for Promoter/3'UTR, P=0.03 ......
ETDT haplotype analysis: P=0.004 for Promoter/3'UTR, P=0.03 for VNTR/3'UTR. Current study demonstrated preferential transmission of the T allele of the 3' UTR polymorphism in the serotonin transporter in a sample of 113 ADHD trios in addition to preferential transmission of haplotypes containing the T allele of the 3' UTR, as well as the long allele of the promoter polymorphism (a finding also supported by the pooled analysis).
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Significant
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