Discovery of novel openers of the understudied human drug target Kir6.1

发现正在研究的人类药物靶标 Kir6.1 的新开启子

• 批准号: 10580933
• 负责人: TIMOTHY J CARDOZO
• 金额: $ 46.61万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10580933
• 关键词: 3-Dimensional; Agonist; Algorithms; American; Amino Acids; Analgesics; Astrocytes; Binding; Biological; Biological Assay; CCL2 gene; CRSP3 gene; Cell Line; Cells; Cessation of life; Chemicals; Complex; Computer Models; Coupled; Cryoelectron Microscopy; Cysteine; Data; Databases; Drug Targeting; Electrophysiology (science); Engineering; Family; Future; Genome; Guns; Helping to End Addiction Long-term; Homology Modeling; Human; Hypersensitivity; Ion Channel; Lead; Libraries; Ligands; Link; MAPK8 gene; Malignant Bone Neoplasm; Maps; Membrane; Metabolic; Methods; Modeling; Molecular Conformation; Molecular Target; Muscle; Nerve; Neurons; Nociception; Nociceptors; Outcome; Overdose; Pain; Pain management; Pharmaceutical Preparations; Phenotype; Phosphatidylinositol 4,5-Diphosphate; Phosphotransferases; Physiological; Pinacidil; Postoperative Pain; Postoperative Period; Probability; Proteins; Public Health; Quantitative Structure-Activity Relationship; Rattus; Regulation; Reporting; Resolution; Rest; Rewards; Route; Services; Site; Skin; Spinal; Spinal Cord; Spinal Ganglia; Structural Models; Structure; Structure-Activity Relationship; Surgical incisions; Testing; Thallium; United States National Institutes of Health; Validation; Vehicle crash; X-Ray Crystallography; addiction; antagonist; assay development; base; cancer pain; chemical group; cheminformatics; chronic pain; drug discovery; druggable target; falls; in silico; in vitro testing; in vivo; inhibitor; innovation; inward rectifier potassium channel; new therapeutic target; non-opioid analgesic; novel; opioid use disorder; pain reduction; pain sensation; painful neuropathy; palmitoylation; patch clamp; preclinical study; prescription opioid; public health emergency; response; screening; small molecule; success; three dimensional structure; three-dimensional modeling; virtual; virtual library; virtual model

Project Summary Drug overdoses now kill more Americans per year than vehicle crash and gun deaths combined. Overdose is a common outcome of opioid use disorder (OUD), which itself has been suspected to evolve from routine prescription treatment of pain with opioid medications. This public health crisis has created a need for the discovery of non-opioid medications to treat pain without eliciting rewarding effects that lead to addiction. New molecular targets for pain therapies offer the best route towards elusive non-opioid medications to treat certain kinds of pain, a fact that is recognized by the present FOA, for which this project is proposed. The ATP- sensitive inward rectifier potassium channel 8 (Kir6.1/KCNJ8) has been linked to the control of postoperative and cancer pain, but little else is known about this ion channel in the context of pain. Specifically, Kir6.1 openers/agonists may be novel analgesics, but drug-like, selective openers/agonists of any KATP family channel have never been identified. Kir6.1 is also considered an understudied protein in terms of its validation as a druggable target generally and is on the list of qualified drug targets for this FOA. We recently investigated the regulation by palmitoylation of Kir6 channels. Our studies identified a druggable site utilized by the palmitoylation mechanism, which provides an avenue to discover drug-like, small molecule opener/agonists of these channels. We proved the principle that we could find selective openers of Kir6.2 by targeting this site using in silico/virtual library screening (VLS) coupled with electrophysiological testing in engineered cell lines. Here, we propose to use the same approach to screen for selective, drug-like openers of Kir6.1 and establish preliminary structure-activity relationships (SAR) between channel opening and specific chemical determinants in the ligands. Specifically, we will build highly accurate homology models of alternative conformations of the Kir6.1/SUR2B complex and use ultra-large VLS to find the maximum number of diverse chemical entities that fit with the target pocket in its agonist, antagonist and unknown conformations (Aim 1). These conformations will be refined based on the observed activity of hit ligands. We will also develop three orthogonal assays to test hit compound identified by VLS: a) a thallium flux cellular primary screening assay, b) patch clamping of Kir6.1 and c) an LPS-stimulated, human astrocyte MCP-1/p-JNK elevation phenotypic assay (Aim 2). Finally, SAR will be derived for each of the three assays, and a meta-SAR across the three, which can serve as an advanced starting point for future drug discovery projects, will be inferred using cheminformatics methods (Deliverable). A specific 3D structural agonist site on KATP channels has not previously been identified, so this proposal is highly innovative.

项目概要 现在，每年因吸毒过量而死亡的美国人比车祸和枪支死亡的总和还多。过量 是阿片类药物使用障碍 (OUD) 的常见结果，其本身被怀疑是从常规演变而来 使用阿片类药物处方治疗疼痛。这场公共卫生危机需要 发现非阿片类药物可以治疗疼痛，但不会引起导致成瘾的奖励效应。新的 疼痛治疗的分子靶标为难以捉摸的非阿片类药物治疗某些疾病提供了最佳途径 目前的 FOA 已经认识到这一事实，为此项目提出了建议。 ATP- 敏感的内向整流钾通道 8 (Kir6.1/KCNJ8) 与术后控制有关 和癌症疼痛，但人们对这种离子通道在疼痛方面的作用知之甚少。具体来说，Kir6.1 开放剂/激动剂可能是新型镇痛药，但任何 KATP 家族的药物样、选择性开放剂/激动剂 通道从未被识别。 Kir6.1 就其验证而言也被认为是一种尚未研究的蛋白质 通常作为可药物靶点，并且在该 FOA 的合格药物靶点列表中。我们最近调查了 Kir6 通道棕榈酰化的调节。我们的研究确定了一个可成药的位点 棕榈酰化机制，为发现药物样小分子开启剂/激动剂提供了途径 这些渠道。我们证明了通过针对该位点可以找到Kir6.2的选择性开启子的原理 使用计算机/虚拟文库筛选 (VLS) 以及工程细胞系中的电生理测试。 在这里，我们建议使用相同的方法来筛选 Kir6.1 的选择性药物样开启剂，并建立 通道开放与特定化学物质之间的初步构效关系（SAR） 配体中的决定因素。具体来说，我们将建立替代品的高精度同源模型 Kir6.1/SUR2B 复合物的构象并使用超大 VLS 来找到不同的最大数量 与目标口袋的激动剂、拮抗剂和未知构象相匹配的化学实体（目标 1）。 这些构象将根据观察到的命中配体的活性进行细化。我们还将开发三 用于测试 VLS 鉴定的命中化合物的正交测定：a) 铊通量细胞初步筛选测定， b) Kir6.1 膜片钳和 c) LPS 刺激的人星形胶质细胞 MCP-1/p-JNK 表型升高 测定（目标 2）。最后，将为这三种测定中的每一种导出 SAR，以及跨这三种测定的元 SAR， 这可以作为未来药物发现项目的高级起点，将使用以下方式推断 化学信息学方法（可交付成果）。 KATP 通道上的特定 3D 结构激动剂位点尚未 此前已被确定，因此该提案具有很强的创新性。