Leveraging genomic risk factors for major depressive disorder to provide mechanistic insights and predictive neurobiological markers

Major Depressive Disorder (MDD) is a disabling, common psychiatric disorder and the leading cause of global disability. A complex combination of genetic and environmental factors gives rise to MDD, although the exact aetiology has not been identified. Genome-wide association studies (GWAS) have established that MDD has a moderate heritability of approximately 37%. MDD has in the past also been associated with abnormalities of white matter microstructure, which represents the brain’s connectivity network. This network is also moderately heritable, providing rationale to investigate its relationship to MDD genetic risk. Over recent years, there has been considerable progress in establishing genetic contributions to MDD. These advances can be harnessed, in combination with neuroimaging and epigenomics, to understand the neurobiology of the disorder. This has only recently become possible at sufficient scale with the availability of large publicly available datasets including genomic, epigenomic, and neuroimaging data. In the current thesis, I therefore aimed to leverage genetic, epigenetic, and neuroimaging data in two large datasets, UK Biobank (N range: 6,400 – 14,800) and Generation Scotland: Scottish Family Health Study (N = 625). Specifically, I aimed to uncover links between white matter microstructure, as measured by fractional anisotropy and mean diffusivity, and (i) differential gene expression as indexed by expression quantitative trait loci (eQTLs) scores in chapter 2; here, decreased white matter integrity was found to be associated with 6 scores regulating genes previously reported to be implicated in neurological and neuropsychiatric disorders, while 2 scores regulating neurodevelopment-linked genes were associated with increased white matter integrity; (ii) MDD genetic risk stratified by the NETRIN1 Signalling Pathway, previously implicated in MDD, indexed by polygenic risk scores (PRS) in chapter 3; results indicated novel associations between the pathway-focussed PRS and decreased white matter integrity in thalamic radiations, as well as several association fibres, including superior and inferior longitudinal fasciculus; (iii) a novel wholegenome epigenetic risk score for MDD, which uncovered an association with MDD, but no significant associations with changes in white matter microstructure (chapter 4). The overall aim of the thesis was to use advanced genomic techniques to stratify genetic function and risk and explore epigenetic risk for MDD in order to identify novel links to structural brain connectivity. Overall, the three studies provide a strong rationale for integrating neuroimaging, genomic and epigenomic data. Specifically, findings in chapter 2 indicate the importance of DCAKD, SLC35A4, SEC14L4, SRA1, PLEKHM1, UBE3C, NMT1, and CPNE1, not previously found by conventional GWAS approaches. This suggests that integrating neuroimaging and genetic expression data may uncover novel associations that inform disease- or trait-specific genetic links to brain connectivity. Chapter 3 results provide a rationale for investigating the NETRIN1 Signalling Pathway and emphasise the role of thalamic connections in MDD within this biological pathway, indicating that novel associations with brain connectivity may be uncovered at a more focused level when stratifying MDD risk by biology. Finally, results from chapter 4 indicate that epigenetics play an important role in MDD risk, although further analysis including larger-scale epigenetic and neuroimaging data should be carried out to uncover the role of epigenetics in relation to brain phenotypes

Structural neuroimaging measures and lifetime depression across levels of phenotyping in UK biobank

Depression is assessed in various ways in research, with large population studies often relying on minimal phenotyping. Genetic results suggest clinical diagnoses and self-report measures of depression show some core similarities, but also important differences. It is not yet clear how neuroimaging associations depend on levels of phenotyping. We studied 39,300 UK Biobank imaging participants (20,701 female; aged 44.6 to 82.3 years, M = 64.1, SD = 7.5) with structural neuroimaging and lifetime depression data. Past depression phenotypes included a single-item self-report measure, an intermediate measure of ‘probable’ lifetime depression, derived from multiple questionnaire items relevant to a history of depression, and a retrospective clinical diagnosis according to DSM-IV criteria. We tested (i) associations between brain structural measures and each depression phenotype, and (ii) effects of phenotype on these associations. Depression-brain structure associations were small (β < 0.1) for all phenotypes, but still significant after FDR correction for many regional metrics. Lifetime depression was consistently associated with reduced white matter integrity across phenotypes. Cortical thickness showed negative associations with Self-reported Depression in particular. Phenotype effects were small across most metrics, but significant for cortical thickness in most regions. We report consistent effects of lifetime depression in brain structural measures, including reduced integrity of thalamic radiations and association fibres. We also observed significant differences in associations with cortical thickness across depression phenotypes. Although these results did not relate to level of phenotyping as expected, effects of phenotype definition are still an important consideration for future depression research

Early-life inflammatory markers and subsequent psychotic and depressive episodes between 10 to 28 years of age

Inflammation is implicated in depression and psychosis, including association of childhood inflammatory markers on the subsequent risk of developing symptoms. However, it is unknown whether early-life inflammatory markers are associated with the number of depressive and psychotic symptoms from childhood to adulthood. Using the prospective Avon Longitudinal Study of Children and Parents birth cohort (N = up-to 6401), we have examined longitudinal associations of early-life inflammation [exposures: interleukin-6 (IL-6), C-reactive protein (CRP) levels at age 9y; IL-6 and CRP DNA-methylation (DNAm) scores at birth and age 7y; and IL-6 and CRP polygenic risk scores (PRSs)] with the number of depressive episodes and psychotic experiences (PEs) between ages 10–28 years. Psychiatric outcomes were assessed using the Short Mood and Feelings Questionnaire and Psychotic Like Symptoms Questionnaires, respectively. Exposure-outcome associations were tested using negative binomial models, which were adjusted for metabolic and sociodemographic factors. Serum IL-6 levels at age 9y were associated with the total number of depressive episodes between 10 and 28y in the base model (n = 4835; β = 0.066; 95%CI:0.020–0.113; pFDR = 0.041) which was weaker when adjusting for metabolic and sociodemographic factors. Weak associations were observed between inflammatory markers (serum IL-6 and CRP DNAm scores) and total number of PEs. Other inflammatory markers were not associated with depression or PEs. Early-life inflammatory markers are associated with the burden of depressive episodes and of PEs subsequently from childhood to adulthood. These findings support a potential role of early-life inflammation in the aetiology of depression and psychosis and highlight inflammation as a potential target for treatment and prevention

Automated Classification of Depression from Structural Brain Measures across Two Independent Community-based Cohorts

ACKNOWLEDGEMENTS: This study was supported and funded by the Wellcome Trust Strategic Award ‘Stratifying Resilience and Depression Longitudinally’ (STRADL) (Reference 104036/Z/14/Z), and the Medical Research Council Mental Health Pathfinder Award ‘Leveraging routinely collected and linked research data to study the causes and consequences of common mental disorders’ (Reference MRC-MC_PC_17209). MAH is supported by research funding from the Dr Mortimer and Theresa Sackler Foundation. The research was conducted using the UK Biobank resource, with application number 4844. Structural brain imaging data from the UK Biobank was processed at the University of Edinburgh Centre for Cognitive Ageing and Cognitive Epidemiology (CCACE) http://www.ccace.ed.ac.uk/), which is a part of the crosscouncil Lifelong Health and Wellbeing Initiative (MR/K026992/1). CCACE received funding from Biotechnology and Biological Sciences Research Council (BBSRC), Medical Research Council (MRC), and was also supported by Age UK as part of The Disconnected Mind project. This work has made use of the resources provided by the Edinburgh Compute and Data Facility (ECDF) (http://www.ecdf.ed.ac.uk/)Peer reviewedPublisher PD

Epigenome-wide association study of global cortical volumes in Generation Scotland:Scottish Family Health Study

Funding This work was supported by the Wellcome Trust [104036/Z/14/Z]. Acknowledgements Generation Scotland received core support from the Chief Scientist Office of the Scottish Government Health Directorates [CZD/16/6] and the Scottish Funding Council [HR03006] and is currently supported by the Wellcome Trust [216767/Z/19/Z]. Genotyping of the GS:SFHS samples was carried out by the Genetics Core Laboratory at the Edinburgh Clinical Research Facility, University of Edinburgh, Scotland and was funded by the Medical Research Council UK and the Wellcome Trust (Wellcome Trust Strategic Award “STratifying Resilience and Depression Longitudinally” (STRADL) Reference 104036/Z/14/Z). MCB is supported by a Guarantors of Brain Non-clinical Post-Doctoral Fellowship. AMM is supported by the Wellcome Trust (104036/Z/14/Z, 216767/Z/19/Z, 220857/Z/20/Z) and UKRI MRC (MC_PC_17209, MR/S035818/1). KLE is supported by the NARSAD Independent Investigator Award (Grant ID: 21956). JMW is supported by UK Dementia Research Institute which is funded by the MRC, Alzheimer’s Research UK and Alzheimer’s Society, by the Fondation Leducq (16 CVD 05), and the Row Fogo Centre for Research Into Ageing and the Brain (BRO- D.FID3668413). This work is part of a project that has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 847776.Peer reviewedPublisher PD

Methylome-wide association study of early life stressors and adult mental health

The environment and events that we are exposed to in utero, during birth and in early childhood influence our future physical and mental health. The underlying mechanisms that lead to these outcomes are unclear, but long-term changes in epigenetic marks, such as DNA methylation, could act as a mediating factor or biomarker. DNA methylation data were assayed at 713 522 CpG sites from 9537 participants of the Generation Scotland: Scottish Family Health Study, a family-based cohort with extensive genetic, medical, family history and lifestyle information. Methylome-wide association studies of eight early life environment phenotypes and two adult mental health phenotypes (major depressive disorder and brief resilience scale) were conducted using DNA methylation data collected from adult whole blood samples. Two genes involved with different developmental pathways (PRICKLE2, Prickle Planar Cell Polarity Protein 2 and ABI1, Abl-Interactor-1) were annotated to CpG sites associated with preterm birth (P < 1.27 × 10(−9)). A further two genes important to the development of sensory pathways (SOBP, Sine Oculis Binding Protein Homolog and RPGRIP1, Retinitis Pigmentosa GTPase Regulator Interacting Protein) were annotated to sites associated with low birth weight (P < 4.35 × 10(−8)). The examination of methylation profile scores and genes and gene-sets annotated from associated CpGs sites found no evidence of overlap between the early life environment and mental health conditions. Birth date was associated with a significant difference in estimated lymphocyte and neutrophil counts. Previous studies have shown that early life environments influence the risk of developing mental health disorders later in life; however, this study found no evidence that this is mediated by stable changes to the methylome detectable in peripheral blood

Epigenome-wide association study of alcohol consumption in N = 8161 individuals and relevance to alcohol use disorder pathophysiology:identification of the cystine/glutamate transporter SLC7A11 as a top target

Alcohol misuse is common in many societies worldwide and is associated with extensive morbidity and mortality, often leading to alcohol use disorders (AUD) and alcohol-related end-organ damage. The underlying mechanisms contributing to the development of AUD are largely unknown; however, growing evidence suggests that alcohol consumption is strongly associated with alterations in DNA methylation. Identification of alcohol-associated methylomic variation might provide novel insights into pathophysiology and novel treatment targets for AUD. Here we performed the largest single-cohort epigenome-wide association study (EWAS) of alcohol consumption to date (N = 8161) and cross-validated findings in AUD populations with relevant endophenotypes, as well as alcohol-related animal models. Results showed 2504 CpGs significantly associated with alcohol consumption (Bonferroni p value < 6.8 × 10(−8)) with the five leading probes located in SLC7A11 (p = 7.75 × 10(−108)), JDP2 (p = 1.44 × 10(−56)), GAS5 (p = 2.71 × 10(−47)), TRA2B (p = 3.54 × 10(−42)), and SLC43A1 (p = 1.18 × 10(−40)). Genes annotated to associated CpG sites are implicated in liver and brain function, the cellular response to alcohol and alcohol-associated diseases, including hypertension and Alzheimer’s disease. Two-sample Mendelian randomization confirmed the causal relationship of consumption on AUD risk (inverse variance weighted (IVW) p = 5.37 × 10(−09)). A methylation-based predictor of alcohol consumption was able to discriminate AUD cases in two independent cohorts (p = 6.32 × 10(−38) and p = 5.41 × 10(−14)). The top EWAS probe cg06690548, located in the cystine/glutamate transporter SLC7A11, was replicated in an independent cohort of AUD and control participants (N = 615) and showed strong hypomethylation in AUD (p < 10(−17)). Decreased CpG methylation at this probe was consistently associated with clinical measures including increased heavy drinking days (p < 10(−4)), increased liver function enzymes (GGT (p = 1.03 × 10(−21)), ALT (p = 1.29 × 10(−6)), and AST (p = 1.97 × 10(−8))) in individuals with AUD. Postmortem brain analyses documented increased SLC7A11 expression in the frontal cortex of individuals with AUD and animal models showed marked increased expression in liver, suggesting a mechanism by which alcohol leads to hypomethylation-induced overexpression of SLC7A11. Taken together, our EWAS discovery sample and subsequent validation of the top probe in AUD suggest a strong role of abnormal glutamate signaling mediated by methylomic variation in SLC7A11. Our data are intriguing given the prominent role of glutamate signaling in brain and liver and might provide an important target for therapeutic intervention

Hair glucocorticoids are associated with childhood adversity, depressive symptoms and reduced global and lobar grey matter in Generation Scotland

ACKNOWLEDGEMENTS We would like to thank all of the Generation Scotland participants for their contribution to this study. We also thank the research assistants, clinicians and technicians for their help in collecting the data. Generation Scotland received core support from the Chief Scientist Office of the Scottish Government Health Directorates [CZD/16/6] and the Scottish Funding Council [HR03006] and is currently supported by the Wellcome Trust [216767/Z/19/Z]. This study was also supported and funded by the Wellcome Trust Strategic Award ‘Stratifying Resilience and Depression Longitudinally’ (STRADL) (Reference 104036/Z/14/Z). We acknowledge the support of the British Heart Foundation (RE/18/5/34216). CG is supported by the Medical Research Council and the University of Edinburgh through the Precision Medicine Doctoral Training Programme. MCB is supported by a Guarantors of Brain Non-Clinical Post-Doctoral Fellowship. JMW is funded by the UK Dementia Research Institute which is funded by the UK Medical Research Council, Alzheimer’s Research UK and Alzheimer’s SocietyPeer reviewedPublisher PD