Synapse loss induced by HIV-1 proteins in the presence of ART and drugs of abuse

翻译后摘要：在ART和药物滥用的存在下，HIV-1蛋白诱导的突触丢失 HIV-1相关的神经认知障碍（HAND）影响几乎一半的感染者。认知 这些患者的损伤与突触树突损伤相关。研究HIV-1相关突触 变化，高含量分析（HCA）测定被开发来量化功能性突触 培养物中海马神经元之间。这种高度自动化的检测方法对HIV-1非常敏感 蛋白质和神经炎症。在该提案中，使用HCA测定来检查浓度-以及 HIV-1蛋白、抗逆转录病毒药物和滥用药物的时间依赖性突触毒性， 组合描述。第一个目标将确定HIV-1蛋白质是否联合协同作用， 增加突触毒性。利用HCA的灵敏度、通量和定量严谨性 检测所有HIV-1蛋白将单独或组合进行检测。浓度响应数据将使 鉴定通过组合各种药物产生的拮抗性、相加性和协同性突触毒性。 proteins.第二个目标是确定抗逆转录病毒药物是否产生突触毒性。有 联合抗逆转录病毒治疗（cART）可能会引起一些直接的神经毒性，这一点越来越受到关注。与更多 FDA批准的20多种抗逆转录病毒药物用于治疗HIV-1， 可能是微妙的，全面评估其潜在的神经毒性，特别是在存在 HIV-1神经毒素缺乏。所有FDA批准的抗逆转录病毒药物和抗逆转录病毒组合将进行突触 临床相关浓度下的毒性。这些结果将确定ART药物具有损害 突触功能第三个目标是确定滥用药物是否会产生突触毒性。40%的艾滋病毒- 1感染者在过去一年内使用非法药物，药物滥用是感染的危险因素。 神经系统功能障碍。一项关于药物滥用如何影响 缺乏HIV-1蛋白和ART药物组合的毒性。代表主要类别的原型药物 将在大脑中检测到的中毒剂量的浓度下测试滥用药物的突触毒性。 第四个目标是评估HIV-1蛋白和药物组合的突触毒性。最有效的代理商 目标1-3中所确定的各项措施将一并进行测试。目标是识别HIV-1蛋白、ART 药物和滥用药物，导致突触损失与高效力。神经毒性组合将是 评价了协同作用、既往暴露于药物滥用后的脱敏作用以及 延长治疗。预期的结果将确立使用HCA测定来鉴定 来自神经炎性药物组合的协同CNS毒性。这些研究将提供一个 为今后研究突触丢失的机制和潜在逆转奠定了基础。新识别 毒性将通过后续体内研究进行验证，并可为HAND患者的临床护理提供信息。

Abstract: Synapse loss induced by HIV-1 proteins in the presence of ART and drugs of abuse HIV-1 associated neurocognitive disorder (HAND) affects almost half of infected individuals. Cognitive impairment in these patients correlates with synaptodendritic damage. To study HIV-1 associated synaptic changes, a high content analysis (HCA) assay was developed to quantify functional glutamatergic synapses between hippocampal neurons in culture. This highly automated assay is exquisitely sensitive to HIV-1 proteins and neuroinflammation. In this proposal, the use of the HCA assay to examine the concentration- and time-dependent synaptic toxicity of HIV-1 proteins, antiretroviral drugs, and drugs of abuse, alone and in combination is described. The first aim will determine whether HIV-1 proteins in combination synergize to increase synaptic toxicity. Taking advantage of the sensitivity, throughput and quantitative rigor of the HCA assay all HIV-1 proteins will be tested singly and in combinations. Concentration response data will enable identification of antagonistic, additive, and synergistic synaptic toxicities produced by combining the various proteins. The second aim will determine whether antiretroviral drugs produce synaptic toxicity. There is growing concern that combined antiretroviral therapy (cART) may elicit some direct neurotoxicity. With more than 20 antiretroviral drugs approved by the FDA for HIV-1 and the expectation that ART-induced neurotoxicity might be subtle, a comprehensive assessment of their potential neurotoxicity, particularly in the presence of HIV-1 neurotoxins, is lacking. All FDA approved ART drugs and ART combinations will be tested for synaptic toxicity at clinically relevant concentrations. These results will identify ART drugs with a propensity to impair synaptic function. The third aim will determine whether drugs of abuse produce synaptic toxicity. 40% of HIV- 1 infected individuals used illegal drugs within the past year and substance abuse is a risk factor for neurological dysfunction in these patients. A comprehensive survey of how drugs of abuse influence the toxicity of HIV-1 proteins and ART drugs in combination is lacking. Prototype drugs representing major classes of abused drugs will be tested for synaptic toxicity at concentrations detected in the brain at intoxicating doses. The fourth aim will evaluate HIV-1 protein and drug combinations for synaptic toxicity. The most potent agents identified in Aims 1-3 will be tested in combination. The goal is to identify combinations of HIV-1 proteins, ART drugs, and drugs of abuse that induce synapse loss with high potency. Neurotoxic combinations will be evaluated for synergistic effects, desensitizing effects following prior exposure to drugs of abuse, and effects of prolonged treatment. The anticipated results will establish the feasibility of using HCA assays to identify synergistic CNS toxicities from combinations of neuroinflammatory agents. These studies will provide a foundation for future studies of the mechanism and potential reversal of synapse loss. Newly identified toxicities will be validated with follow up in vivo studies and could inform clinical care of HAND patients.