| Summary |
The 4p15.31 region was shown to be associated with ADHD in several GWASs. In the present study we also report association of the 4p15.31 locus with Cluster B and Cluster C PD (personality disorders) as identified by a pooled genome-wide association study in 400 individuals suffering from PD. The gene coding for the Kv channel-interacting protein 4 ( KCNIP4) is located in this region. KCNIP4 is an interaction partner of prese nilin and plays a role in a negative feedback loop in the Wnt/¦Â-catenin pathway. Thus, we reasoned it to be a promising candidate gene for ADHD as well as for PD. To clarify the role of KCNIP4in those disorders, we conducted candidate gene based association studies in 594 patients suffering from adult ADHD and 630 PD patients as compared to 974 healthy control individuals. In the adult ADHD sample, six single markers and one haplotype block revealed to be associated with disease (p values from 0.0079 to 0.049). Seven markers within the KCNIP4 gene showed an association with PD (p values from 0.0043 to 0.0437). The results of these studies suggest a role of KCNIP4in the etiology of ADHD, PD and other comorbid disorders. |
| Total Sample |
For association tests of KCNIP4 with adult ADHD and comorbid disorders: 49 of 56 SNPs within or near the KCNIP4 gene passed quality control for association testing in 594 patients suffe ring from aADHD and 974 healthy controls from Germany. For association tests of KCNIP4 and childhood ADHD: Pedigree disequilibrium testing for 51 SNPs which passed quality check was performed with genotyping data gained from 171 families (113 trios, 43 quartets and 15 multisibling) consisting of 592 individuals in total. |
| Sample Collection |
For the mass array based SNP genotyping the healthy control group recruited in the Lower Franconia area and in Munich in Germany consisted of 974 healthy volunteers of Caucasian origin. A total of 594 patients suffering from aADHD were available for genotyping. The family-based childhood ADHD sample were from Wurzburg. |
| Diagnosis Description |
All aADHD patients were evaluated by experienced psychiatrists in an established four step procedure. First, other physical and mental conditions that could explain the symptoms more adequately such as addictive disorders, bipolar disorders, schizophrenia or mental retardation were excluded. Second, adult manifestation of ADHD was assessed according to the DSM-IV criteria by a semi-structured diagnostic interview. Informative input from partners, relatives and friends was also collected. The matching of psychometric, psychopathological and biographical information was proven. To ensure diagnostic validity probands were examined by more than one experienced investigator at least at two time points. Third, onset before the age of 7 years was proven by retrospective diagnosis (which was confirmed by a family member and certificates, wherever possible). Fourth, anamnestic information demonstrate that the symptoms are a lifelong condition and definitely do not have an episodic course. Furthermore, quantitative personality traits had been assessed in the aADHD and personality disorder samples by using two established questionnaires (NEO-PI R and TPQ). All children with ADHD were assessed by experienced child psychiatrists using also full semi-structured interview (Kiddie-Sads-PL-German Version or Kinder-DIPS) and parent and teacher ADHD DSM-IV based rating scales to ensure pervasiveness of symptoms. Exclusion criteria were an intelligence quotient <80, comorbid autistic disorders or somatic disorders (hyperthyroidism, epilepsy, neurological diseases, severe head trauma, etc.), primary affective disorders, autism, Tourette syndrome and other severe primary psychiatric disorders. |
| Technique |
The SNP IDs (rs numbers) of SNPs located within the exons of the KCNIP4 gene and their flanking intronic sequences plus the 5' and 3' flanking sequences were downloaded from the HapMap genome browser. Tagging SNP selection was performed using the tagger algorithm of the Haploview software for pairwise tagging with r-square threshold >0.8 and a minor allele frequency >5% to tag all exonic and the promoter regions, yielding a total of 56 SNPs. SNP genotyping was conducted using the MassArray system. PCR was performed using iPlex chemistry as recommended in the MassArray iPlex standard operating procedure and a thermocycler. Primer sequences can be obtained on request. After spotting the samples onto a SpectroCHIP (Sequenom) using the SequenomMassARRAY Nanodispenser, mass spectrometric analyses were carried out on a BrukerAutoflex time-of-flight mass spectrometer. |
| Analysis Method |
Mass spectra were all checked by eye for quality control before the statistical evaluation. Inconsistent genotypes in the family sample were revealed and removed by means of PedCheck. Single SNP association tests were carried out using generalized linear models (GLM) implemented in the R function glm() Family-based association analysis was carried out using pedigree disequilibrium test. |
| Result Description |
For association tests of KCNIP4 with adult ADHD and comorbid disorders: Six SNPs showed a nominally significant association with aADHD with p values <0.05. In addition, in the adult ADHD sample, eight markers were found to be associated with comorbid disorders compared to controls (p values 0.0067-0.0471). Haplotype testing revealed one of the nine haploblocks to be associated with adult ADHD (p=0.0079). The risk haplotype showed an odds ratio of 1.99. None of the SNPs within this haploblock showed an association with aADHD on the single marker level. For association tests of KCNIP4 and childhood ADHD: Two SNPs indicated a nominally significant association with cADHD (rs16869915, p=0.0477 and rs358834, p=0.0411). However, the association of rs876477 with cADHD could not be replicated. Nine markers showed an overtransmission of one allele for comorbid disorders, but only two of them acquired a sufficient significance level (rs11726872: learning disability p=0.0038; rs6447994: oppositional defiant disorder p=0.0023). |
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