Proteomic biosignatures of withdrawal from cocaine in rhesus monkeys

描述（由申请人提供）：在美国，可卡因滥用和依赖仍然是一个主要的公共卫生问题，但在对患有成瘾障碍的个人进行适当诊断和治疗方面仍然存在重大障碍。研究工作的主要重点是发展和改进毒理学分析方法，以检测可卡因，作为最近可卡因使用情况的一种措施，并查明表明易滥用可卡因的遗传标记。虽然这两种方法本身都很有用，但仍有一个未满足的医疗需求，即识别外围生物标志物，以协助客观诊断，识别患者亚群，并监测进展和对治疗的反应。此外，生物标志物也可以作为与慢性可卡因使用相关的神经生物学变化的指标。为此，我们将利用我们在与可卡因滥用相关的蛋白质组学研究方面的良好记录，识别血浆生物标志物，并将其与恒河猴慢性可卡因自我给药戒断过程中奖赏相关脑区发生的生化变化联系起来。这些研究将利用博士的科学专业知识。Porrino和Nader以及他们正在进行的NIDA资助研究的资源（R01DA009085）。在试验1中，血浆样本将在100天的可卡因自我给药期间纵向收集，并在30天的停药期间收集。样品将使用无偏互补蛋白质组学方法（2D-DIGE和iTRAQ/MDLC与MALDI ToF/ToF分析）和适当的验证方法进行分析。将使用多变量统计分析方法来描述一组有限的蛋白质，这些蛋白质在摄入和停药以及急性和慢性停药之间具有最大程度的可区分性。Aim 2的研究将探讨在戒除可卡因1天和30天时，可卡因诱导的腹侧纹状体NMDA和AMPA受体及相关蛋白的失调。此外，我们将使用靶向蛋白质组学方法，使用iTRAQ /MDLC和MALDI ToF/ToF分析来评估可卡因戒断对腹侧纹状体膜组分中NR1亚基和相关复合物的影响。在Aim 1中鉴定的外周生物标志物的表达将与Aim 2中鉴定的神经生物学变化相关。我们预计这些策略将提供可卡因使用/戒断和相关神经元失调的初步生物特征，最终可用于治疗可卡因成瘾的诊断和预后工具。

DESCRIPTION (provided by applicant): Cocaine abuse and dependence in the United States continues to be a major public health concern yet significant obstacles remain for the appropriate diagnosis and treatment of individuals suffering from addictive disorders. Research efforts have focused primarily on the development and refinement of toxicological assays for detecting cocaine as a measure of recent cocaine use and on the identification of genetic markers indicative of a predisposition to cocaine abuse. While both approaches are useful in their own right, a continuing unmet medical need is the identification of peripheral biological markers that can assist with the objective diagnosis, identification of subsets of patients, and monitor progression and response to treatment. Moreover, biomarkers could also be used as indicators of neurobiological changes associated with chronic cocaine use. To this end, we will use our well developed track record in proteomics research relevant to cocaine abuse to identify and correlate plasma biomarkers with biochemical changes occurring in reward relevant brain regions during withdrawal from chronic cocaine self-administration in rhesus monkeys. These studies will leverage the scientific expertise of Drs. Porrino and Nader and the resources of their ongoing NIDA funded study (R01DA009085). In Aim 1, plasma samples will be collected longitudinally during a 100 day period of cocaine self- administration through a thirty day period of withdrawal. Samples will be analyzed using unbiased complimentary proteomic methods (2D-DIGE and iTRAQ/MDLC with MALDI ToF/ToF analysis) and appropriate validation methods. Multivariate statistical analysis approaches will be used to delineate a finite set of proteins with the greatest degree of discriminability between intake and withdrawal and between acute and chronic withdrawal. Studies in the Aim 2 will explore the cocaine induced dysregulation of NMDA and AMPA receptors and associated proteins in the ventral striatum at one and thirty days of withdrawal from cocaine. In addition, we will use a targeted proteomic approach to evaluate the effects of cocaine withdrawal on the NR1 subunit and associated complex in membrane fractions from the ventral striatum using iTRAQ /MDLC with MALDI ToF/ToF analysis. The expression of peripheral biomarkers identified in Aim 1 will be correlated with neurobiological changes identified in Aim 2. We anticipate these strategies will provide a preliminary biosignature of cocaine use/withdrawal and correlated neuronal dysregulation that ultimately can be used as a diagnostic and prognostic tool in the treatment of cocaine addiction. PUBLIC HEALTH RELEVANCE: A continuing need in addiction medicine is the identification of peripheral biological markers that are indicative of prior chronic drug use and are correlated with changes in the brain that mediate reward, craving and relapse. Using a rhesus monkey model of cocaine self-administration withdrawal, we propose to define a proteomic biosignature in plasma that is correlated with neurobiological changes occurring in response to cocaine withdrawal. The proposed studies are intended to delineate a preliminary set of peripheral biomarkers that can be further evaluated in the rhesus monkey model and in human behavioral laboratory studies before implementing as potential diagnostic and prognostic tool at the clinic.