(Epi)genetic Risk Architectures of Opioid-Dependent Brain

With nearly 20,000 overdose deaths in 2014 alone, opioid addiction (OA) has emerged as one of the most pressing public health crises in recent US history. One-fifth of individuals who try heroin develop an addiction to opioids. Genetics is a major contributor to OA with an estimated 60% heritability, only somewhat less than schizophrenia (80%) which has recently seen substantial gains in identified underlying genetics. Yet, studies to date have failed to uncover most of the genes that predispose individuals to OA, leaving the overwhelming fraction of OA heritability unexplained. The proposed study takes a novel, integrated "omics-based" strategy to investigate the molecular basis of OA and uncover both genetic and epigenetic factors associated with opioid addiction. The premise for our approach is founded on studies from our labs, and others, implicating regulatory variation in common traits and diseases, including those associated with complex brain phenotypes like addiction. We have collected the largest known cohort of postmortem brains from addicts who overdosed on opioids, along with matched control brains from non-users. From both cases and control specimens, we will isolate cells from 2 regions of the brain closely implicated in the addiction phenotype: the Prefrontal Cortex (PFC) and the Nucleus Accumbens (NAc). In Aim 1, we propose ChIP-seq studies to identify regulatory elements that distinguish cases from controls and define the opioid addiction phenotype. The regulatory differences will be connected to their gene targets through high resolution in situ Hi-C. In Aim 2, we propose QTL-based approaches to identify genetic variants that underlie the regulatory differences (hmQTLs). We also propose eQTL analyses to identify genetic variants that underlie differences in transcript levels between cases and controls. Aim 3 leverages the largest heroin addiction GWAS meta-analysis to date to test the hypothesis that SNPs in regions associated with chromatin and expression differences between cases and controls define novel loci for predisposition to OA. Each Aim has the potential for discovery independent of the others (differential HM, RNAexp, QTLs, and variant-phenotype associations) but their synergy is the most powerful component of the proposed study: identifying regulatory pathways that generate, not only phenotype associations, but hypothesized mechanisms for those associations which can be the focus of new opioid addiction prevention and treatment development.]]>

2014年美国阿片类药物成瘾（opioid addiction, OA）相关过量死亡人数就近2万，已然成为近年来美国公共卫生领域最紧迫的危机之一。五分之一尝试海洛因的个体最终会发展为阿片类药物成瘾。遗传是OA的主要易感因素，其估计遗传力达60%，仅略低于精神分裂症（80%）——而后者在潜在致病遗传学研究领域近年已取得显著进展。但迄今为止的相关研究尚未探明绝大多数使个体易患OA的基因，导致OA遗传力的绝大部分仍无法得到解释。

本研究提出一种新颖的整合性“组学（omics-based）”策略，用以探究阿片类药物成瘾的分子机制，同时揭示与阿片类药物成瘾相关的遗传与表观遗传因素。本研究的设计理念基于本课题组及其他团队的相关研究，这些研究证实调控变异与诸多常见性状及疾病相关，包括成瘾这类复杂脑表型相关疾病。

我们已收集到目前已知规模最大的阿片类药物过量成瘾死者的死后脑组织队列，同时匹配了非使用者的对照脑组织样本。我们将从病例组与对照组标本的两个与成瘾表型密切相关的脑区——前额叶皮层（Prefrontal Cortex, PFC）与伏隔核（Nucleus Accumbens, NAc）——中分离细胞。

在目标1中，我们计划开展染色质免疫沉淀测序（ChIP-seq）研究，以识别区分病例组与对照组的调控元件，并明确阿片类药物成瘾的表型特征。这些调控差异将通过高分辨率原位Hi-C技术与对应的基因靶标建立关联。

在目标2中，我们计划采用基于数量性状位点（QTL）的分析方法，识别介导调控差异的遗传变异（组蛋白修饰QTL, hmQTL）。同时我们还将开展表达数量性状位点（eQTL）分析，以识别导致病例组与对照组转录水平差异的遗传变异。

目标3将借助目前规模最大的海洛因成瘾全基因组关联研究（GWAS）荟萃分析数据集，验证如下假说：位于与病例组和对照组间染色质及表达差异相关区域的单核苷酸多态性（SNPs），可定义新的OA易感位点。

每个目标均具备独立的发现潜力（包括差异组蛋白修饰、RNA表达、QTL分析以及变异-表型关联分析），但三者的协同效应才是本研究最具效力的核心：不仅能识别与表型相关的调控通路，还能阐明这些关联背后的潜在机制，为新型阿片类药物成瘾预防与治疗手段的开发提供研究方向。