Genome-Wide DNA Methylation Patterns in Persistent Attention-Deficit/Hyperactivity Disorder and in Association With Impulsive and Callous Traits

Abstract

Attention-deficit/hyperactivity disorder (ADHD) is a neurodevelopmental disorder that often persists into adulthood. ADHD and related personality traits, such as impulsivity and callousness, are caused by genetic and environmental factors and their interplay. Epigenetic modifications of DNA, including methylation, are thought to mediate between such factors and behavior and may behave as biomarkers for disorders. Here, we set out to study DNA methylation in persistent ADHD and related traits. We performed epigenome-wide association studies (EWASs) on peripheral whole blood from participants in the NeuroIMAGE study (age range 12-23 years). We compared participants with persistent ADHD (n = 35) with healthy controls (n = 19) and with participants with remittent ADHD (n = 19). Additionally, we performed EWASs of impulsive and callous traits derived from the Conners Parent Rating Scale and the Callous-Unemotional Inventory, respectively, across all participants. For every EWAS, the linear regression model analyzed included covariates for age, sex, smoking scores, and surrogate variables reflecting blood cell type composition and genetic background. We observed no epigenome-wide significant differences in single CpG site methylation between participants with persistent ADHD and healthy controls or participants with remittent ADHD. However, epigenome-wide analysis of differentially methylated regions provided significant findings showing that hypermethylated regions in the APOB and LPAR5 genes were associated with ADHD persistence compared to ADHD remittance (p = 1.68 * 10 −24 and p = 9.06 * 10 −7 , respectively); both genes are involved in cholesterol signaling. Both findings appeared to be linked to genetic variation in cis. We found neither significant epigenome-wide single CpG sites nor regions associated with impulsive and callous traits; the top-hits from these analyses were annotated to genes involved in neurotransmitter release and the regulation of the biological clock. No link to genetic variation was observed for these findings, which thus might reflect environmental influences. In conclusion, in this pilot study with a small sample size, we observed several DNA-methylation-disorder/trait associations of potential significance for ADHD and the related behavioral traits. Although we do not wish to draw conclusions before replication in larger, independent samples, cholesterol signaling and metabolism may be of relevance for the onset and/or persistence of ADHD.

Epigenome-wide association study of attention-deficit/hyperactivity disorder in adults

“…We assessed whether probes in different categories: (i) showing a statistically significant proportion of methylation variance explained by additive genetic effects as reported by Zeng et al 47 ; (ii) probes identified in previous EWASs on exposure to adverse live events [48][49][50] ; (iii) probes identified in previous EWASs on ADHD 21,22 or ADHD symptoms 6,9 or (iv) probes located in ADHDassociated loci identified through GWAS 4,29,51 showed, on average, a stronger association with adult ADHD than other methylation sites by regressing our EWAS test statistics (Z score ) on each CpG category as described by van Dongen et al 9 :…”

“…One identified methylation changes associated with ADHD symptomatology that did not remain significant when results were meta-analyzed across cohorts 9 . The second one found hypermethylated regions in genes involved in fatty acid metabolism and fatty acid oxidation pathways associated with ADHD persistence when compared to remittance 21 . In the childhood period, Wilmot et al analyzed a population cohort of school-age boys and found lower methylation levels at the VIPR2 gene in ADHD subjects compared to their age-and sex-matched controls 10 , results that were recently replicated in the largest EWAS on ADHD in children conducted so far 22 .…”

“…Studies of DNA methylation profiles in ADHD have been conducted using peripheral blood, cord blood, buccal samples or saliva 6,[9][10][11][20][21][22][23][24][25][26][27][28] . Candidate gene studies have revealed differential methylation patterns in genes involved in the dopaminergic, serotoninergic and neurotrophic systems, including SLC6A4, DRD4, COMT, ANKK1, BDNF, or NGFR, associated with ADHD symptomatology and severity [23][24][25][26][27][28] .…”

“…Candidate gene studies have revealed differential methylation patterns in genes involved in the dopaminergic, serotoninergic and neurotrophic systems, including SLC6A4, DRD4, COMT, ANKK1, BDNF, or NGFR, associated with ADHD symptomatology and severity [23][24][25][26][27][28] . Seven epigenome-wide association studies (EWASs) on ADHD have been run to date, with sample sizes ranging from 54 subjects for clinical samples 21 to 4,689 individuals in a meta-analysis considering ADHD symptomatology in general population 9 , yielding nonoverlapping findings across them 6,[9][10][11][20][21][22] . There is limited research on adults using this approach, given that most of the EWASs have focused on pediatric samples 6,10,11,20,22 .…”

“…There is limited research on adults using this approach, given that most of the EWASs have focused on pediatric samples 6,10,11,20,22 . To the best of our knowledge, only two studies evaluated methylome-wide patterns on adults 9,21 . One identified methylation changes associated with ADHD symptomatology that did not remain significant when results were meta-analyzed across cohorts 9 .…”