Development of in vivo probes to study the function of TRIP8b in cognition

摘要 认知功能障碍是与慢性应激相关的神经精神疾病的共同特征。这些 症状可能会使人虚弱，尽管严重抑郁障碍等疾病的发病率很高， 治疗选择相对较少。为了更好地解决这些症状，新的分子靶点是 需要的。超极化激活的环核苷酸门控(HCN)通道在调节 海马体的兴奋性，这是一个与认知密切相关的大脑区域。避免心脏副作用 与拮抗HCN通道(也在心脏中表达)有关，我们小组专注于 在TRIP8b上，HCN通道的神经元特异性亚单位。在我们最近发布的报告中(Lyman等人，在 在《科学转化医学》杂志上发表的一篇文章中，我们证实了化学遗传学扰乱了这种相互作用 背侧海马区TRIP8b和HCN之间的相互作用足以挽救脑损伤后的认知损害 慢性压力。总而言之，这些结果与一种假设是一致的，即一种破坏 TRIP8b-HCN的相互作用可以挽救慢性应激后出现的认知功能障碍。为了 利用这一发现，我们团队最近进行了一次高通量筛查，以确定小分子 TRIP8b-HCN相互作用的抑制剂。我们确定了一个显示出有希望的活性的候选支架 作为TRIP8b抑制剂。这个复合系列是我们目前提交的文件的基础，在该文件中，我们 建议进一步优化药物化学，开发用作化学探针的TRIP8b抑制剂 目的：研究TRIP8b在体内的作用。在我们附上的初步数据中，我们显示我们的抑制剂有效地破坏了 TRIP8b与HCN在细胞内的相互作用毒性可忽略不计。这种抑制产生了健壮的 体内和体外对hcn通道介导的Ih电流的影响。为了扩展这项工作，我们 提出三个目标。在目标1中，我们将进行迭代药物化学优化以开发 具有改进的效力和药学特性的化合物。在我们的产品中表现良好的化合物 生化和生物物理分析将在目标2中推进到细胞分析，以确定它们是否 能够破坏TRIP8b-HCN的相互作用。在目标3中，我们将利用体内的化合物来研究 它们对海马区功能的影响。将对有希望的化合物进行研究，以了解它们的救援能力 慢性社会失败导致的认知缺陷，这是慢性压力的动物模型。这些实验 将使我们能够开发可以用作化学探针的TRIP8b抑制剂来研究HCN的作用 慢性应激后认知功能障碍的调节途径。

Abstract Cognitive dysfunction is a common feature of neuropsychiatric diseases related to chronic stress. These symptoms can be debilitating, and despite the high prevalence of illnesses like Major Depressive Disorder, there are relatively few treatment options. To better address these symptoms, novel molecular targets are needed. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels play a critical role in regulating excitability in the hippocampus, a brain region critically involved in cognition. To avoid cardiac side effects associated with antagonizing HCN channels (which are also expressed in the heart), our group has focused on TRIP8b, a neuron-specific subunit of HCN channels. In our recently published report (Lyman et al, in press at Science Translational Medicine), we established that chemogenetically disrupting the interaction between TRIP8b and HCN in the dorsal hippocampus is sufficient to rescue impairments in cognition after chronic stress. Combined, these results are consistent with the hypothesis that a small molecule that disrupts the TRIP8b-HCN interaction could rescue the cognitive dysfunction seen after chronic stress. In order to capitalize on this discovery, our group recently executed a high throughput screen to identify small molecule inhibitors of the TRIP8b-HCN interaction. We identified one candidate scaffold which shows promising activity as a TRIP8b inhibitor. This compound series serves as the basis for our current submission in which we propose further medicinal chemistry optimization to develop TRIP8b inhibitors for use as chemical probes in order to study TRIP8b in vivo. In our enclosed preliminary data, we show that our inhibitor potently disrupts the interaction between TRIP8b and HCN in cells with negligible toxicity. This inhibition produces robust effects on Ih, the current mediated by HCN channels both in vivo and in vitro. To expand upon this work, we propose three aims. In Aim 1, we will perform iterative medicinal chemistry optimization to develop compounds with improved potency and pharmaceutical characteristics. Compounds that perform well in our biochemical and biophysical assays will be advanced to cellular assays in Aim 2 in order to determine if they are capable of disrupting the TRIP8b-HCN interaction. In Aim 3 we will utilize our compounds in vivo to study their effect on hippocampal function. Promising compounds will be investigated for their ability to rescue cognitive deficits that result from chronic social defeat, an animal model of chronic stress. These experiments will enable us to develop TRIP8b inhibitors that can be used as chemical probes to study the role of HCN channels in regulating cognitive dysfunction after chronic stress.