Proteomic Biosignatures of Chronic Drug Exposure

描述（由申请方提供）：基于与慢性药物暴露特异性相关的长期血浆蛋白变化的生物特征可用于识别易重复使用药物的患者，从而促进早期干预。我们将使用大鼠长期暴露于成瘾药物和差异蛋白质组表达来测试找到这种特征的可行性。根据14天递增剂量方案（慢性暴露），大鼠将接受可卡因或非成瘾性多巴胺转运蛋白抑制剂或吗啡或生理盐水治疗，并在末次药物治疗后75天内收集血浆。蛋白质组学分析将涉及逐步耗尽丰富的血浆蛋白，然后通过液相色谱法与质谱联用，以分析由非常低丰度蛋白质组成的耗尽的蛋白质组和中等丰度的结合蛋白质。其改变的表达维持数周（例如30天）并最终恢复至基线的蛋白质将是候选生物标志物，其可用于在成瘾药物不再容易测量时检测近期暴露。将通过在不需要蛋白抗体的定量多重测定中进行多反应监测来验证最佳候选物。将检查蛋白质组学变化与慢性药物治疗的行为和神经化学后遗症的时间相关性，并将平行测量。将使用统计分析和数据挖掘来确定具有所需特性的生物特征。一项成功的可行性研究将刺激并为人类受试者样本的类似研究提供信息。 公共卫生相关性：对慢性药物暴露的分析生物标志物（生物特征）的需求尚未得到满足，这将有助于识别易重复用药的患者。有效的测试可以成为常规医疗保健的一部分，以确定需要治疗药物使用障碍的患者，并减轻吸毒成瘾的个人和社会负担。

DESCRIPTION (provided by applicant): A biosignature based on long-term plasma protein changes specifically associated with chronic drug exposure can be used to identify patients predisposed to repeated drug use and thereby facilitate early intervention. We will test the feasibility of finding such a signature using chronic exposure of rats to addictive drugs and differential proteomic expression. Rats will receive treatment with cocaine or a non-addictive dopamine transporter inhibitor, or morphine, or saline according to a 14 day escalating dose schedule (chronic exposure) and plasma collected up to 75 days following last drug treatment. Proteomic analysis will involve stepwise depletion of abundant plasma proteins followed by liquid chromatography coupled online with mass spectrometry to analyze both the depleted proteome consisting of very low abundance proteins and the bound proteins of intermediate abundance. Proteins whose altered expression is maintained for several weeks, e.g. 30 days, and eventually return to baseline would be candidate biomarkers that could be used to detect recent exposure at a time when the addictive drug can no longer be readily measured. Top candidate will be validated by multiple reaction monitoring in a quantitative multiplexed assay that does not require antibodies to the proteins. Proteomic changes will be examined for temporal correlation with behavioral and neurochemical sequelae of chronic drug treatment that will be measured in parallel. Statistical analysis and data mining will be used to identify a biosignature with the desired properties. A successful feasibility study would stimulate and inform similar studies on human subject samples. PUBLIC HEALTH RELEVANCE: There is an unmet need for analytical biomarkers, a biosignature, of chronic drug exposure that would help to identify patients predisposed to repeated drug. An effective test could become part of routine medical care that would identify patients in need of treatment for substance use disorders and mitigate the personal and societal burdens of drug addiction.