Mechanism of Action for n-3 PUFA antidepressant properties

 描述(由申请人提供)：假设G蛋白Gs与脂筏相关的程度是抑郁症和治疗反应的指标。最初的观察表明这一生物特征是在死亡组织中进行的，我们将这项工作扩展到体内和体外的初步数据，表明通过简单的洗涤剂提取确定Gs的RAFT定位是抑郁和抗抑郁反应的生物标志物。有研究表明，n-3多不饱和脂肪酸既有抗抑郁作用，也有协同某些抗抑郁药物的作用。为了测试这一点，我们将使用一种神经胶质细胞系以及来自抑郁受试者的淋巴母细胞，这些受试者要么对抗抑郁药物有效，要么无效。这应该为抗抑郁剂的反应提供一个预测模型，无论该药是鱼油、西酞普兰还是某种组合。我们将通过测量在NIMH资助的临床试验期间收集的血细胞膜中Gs的脂筏分布来测试该生物标记物的适用性，该临床试验显示n-3PUFA增强了抗抑郁反应。拟议的实验还试图将机械性的临床前研究与临床研究结合起来，以提出和测试生物标记物。临床前工作试图确定用抗抑郁剂±n-3多不饱和脂肪酸体外治疗是否将Gs转移到质膜的非RAFT组分，在那里它更有效地激活腺苷环化酶。我们认为，一种具有抗抑郁作用的化合物集中在富含胆固醇的膜结构域(脂筏)，并破坏Gs在这些结构域中的锚定。一种可能的机制是直接修改Gs。这将通过膜组分的生化分级、光漂白后的荧光恢复(FRAP)和细胞成像来研究。由于大多数抗抑郁药物治疗(节省氯胺酮)在疗效之前需要一段时间延迟，并且这可以在细胞中复制，因此将使用荧光Gs融合蛋白实时监测Gs对抗抑郁药物的反应的易位。如果n-3PUFA单独或与SSRIs一起显示出这些抗抑郁活性的生物学特征，将通过检查n-3PUFA(±SSRIs)抗抑郁效果的临床研究中采集的血液来解释这一点。初始数据 这项研究表明，与单独使用安慰剂或SSRI相比，n3 PUFA+SSRI具有更强的抗抑郁反应。最后，我们将通过生化和成像研究来测试来自已知对西酞普兰有反应(或缺乏反应)的抑郁症患者的淋巴母细胞。一个目标是开发一种内容丰富的屏幕，它可能会前瞻性地建议给定潜在疗法的有效性。拟议研究的成功完成还将表明，在抗抑郁治疗中补充n-3多不饱和脂肪酸对于缩短治疗开始时间和/或减少抗抑郁药的总剂量是有用的。他们还将为建立一种低成本的血液生物标记物铺平道路，该标记物可以在广泛的药物范围内同时用于抑郁症和抗抑郁剂的疗效。

 DESCRIPTION (provided by applicant): It is hypothesized that the degree to which the G protein, Gs, is associated with lipid rafts is an indicator of both depression and therapeutic response. The original observations suggesting this biosignature were made in postmortem tissue, and we have extended this work with in vivo and in vitro preliminary data suggesting that raft localization of Gs, determined by simple detergent extraction, is a biomarker of both depression and antidepressant response. There are suggestions that n-3 PUFA either have antidepressant activity or can synergize the action of some antidepressant drugs. To test this, we will use a glial cell line as well as lymphoblasts from depressed subjects who were either antidepressant responsive or unresponsive. This should provide a predictive model for the response to an antidepressant agent, whether that agent is fish oil, citalopram, or some combination. We will test the applicability of the biomarker by measuring the lipid raft distributin of Gs in membranes from blood cells collected during an NIMH-funded clinical trial showing n-3 PUFA augmentation of antidepressant response. The proposed experiments also attempt to mesh mechanistic preclinical studies with a clinical study to propose and test a biomarker. The preclinical work seeks to determine whether in-vitro treatment with antidepressants ± n-3 PUFA shifts Gs into non-raft fractions of the plasma membrane, where it more effectively activates adenylyl cyclase. We suggest that a compound with antidepressant efficacy concentrates in cholesterol-rich membrane domains (lipid rafts) and disrupts the anchoring of Gs within those domains. One possible mechanism for this is direct modification of Gs. This will be studied by biochemical fractionation of membrane components, fluorescence recovery after photobleaching (FRAP) and by cellular imaging. Since most antidepressant therapy (save ketamine) requires a time lag prior to therapeutic efficacy and this can be replicated in cells, translocation of Gs in response to antidepressant agents will be monitored in real time using a fluorescent Gs fusion protein. Should n-3 PUFA show these biological hallmarks of antidepressant activity, alone or in combination with SSRIs , this will be translated by examining blood taken in a clinical study of the antidepressant efficacy of n-3 PUFA (± SSRIs). Initial data from this study show a greater antidepressant response to n3 PUFA + SSRI than to placebo or SSRI alone. Finally, we will test, with the biochemical and imaging studies, lymphoblasts, derived from depressed patients with known response (or lack thereof) to citalopram. One goal is to develop a high content screen that might suggest, prospectively, the effectiveness of a given potential therapy. Successful completion of the proposed studies will also indicate the usefulness of n-3 PUFA supplementation to antidepressant therapy for decreasing time of therapeutic onset and/or lowering overall antidepressant dose. They will also pave the way toward establishing a low-cost blood biomarker for both depression and antidepressant efficacy over a broad range of agents.