De novo protein neurotoxins for ion channels

离子通道的从头蛋白神经毒素

• 批准号: 8883982
• 负责人: Steve A N Goldstein
• 金额: $ 31.28万
• 依托单位: BRANDEIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8883982
• 关键词: Affect; Affinity; American; Arrhythmia; Atrial Fibrillation; Bacteriophages; Beta Cell; Binding; Cardiac; Cell surface; Cells; Clinical; Cloning; Diabetes Mellitus; Diagnosis; Disease; Disulfides; Diversity Library; Drug Targeting; Elan brand of omega-conopeptide MVIIA; Fertility; Generations; Gland; Glucose; Grant; Health; Heart; Heart Atrium; Human; Insulin-Dependent Diabetes Mellitus; Investigation; Ion Channel; Ion Channel Protein; Kv1.3 potassium channel; Kv1.5 potassium channel; Kv2.1 channel; Laboratories; Lead; Left; Libraries; Ligands; Membrane; Methodology; Methods; Morbidity - disease rate; Neurotoxins; Non-Insulin-Dependent Diabetes Mellitus; Orphan; Pancreas; Peptide Hydrolases; Peptides; Phage Display; Pharmaceutical Preparations; Population; Potassium; Potassium Channel; Preparation; Proteins; Protons; Public Health; Reporting; Resistance; Role; Scorpions; Solutions; Sorting - Cell Movement; Source; Specificity; Sperm Capacitation; Spiders; Structure; Surface; Targeted Toxins; Technology; Testing; Therapeutic; Therapeutic immunosuppression; Toxin; Validation; Variant; Venoms; Voltage-Gated Potassium Channel; Work; base; biophysical techniques; combinatorial; gene cloning; improved; inhibitor/antagonist; insulin secretion; interest; male; mortality; preclinical study; public health relevance; receptor; scaffold; sensor; success; synthetic peptide; therapeutic target; tool; treatment strategy; voltage

 DESCRIPTION (provided by applicant): Protein neurotoxins are among the most powerful tools in the arsenal for investigation of membrane receptors and are now used as well for treatment of disease. This grant seeks, first, to advance our method of creation of de novo toxins by targeting potassium and proton channels that lack these useful ligands; second, to use de novo toxins to assess the potential of Kv1.5 and Kv2.1 channels as targets for treatment of two diseases of broad public impact- Atrial Fibrillation (AF) and Type 2 Diabetes (T2D), respectively. Success on these two fronts will allow wider use of these tools to produce toxins for other targets and motivate analogous work directed towards other types of membrane receptors important to human health and disease. To date, toxins have been derived primarily from limited sources: crude venoms, venom gland gene cloning, and laboratory synthesis of known toxins. Therefore, most receptors of interest lack a recognized, specific, high-affinity ligand. This state-of-affairs is easily understood: neither the purpose of toxins in the wild nor screens of venoms favor target specificity. Here, these problems are avoided by cloning toxins based on desired attributes using a new, high- throughput method we developed to generate a selective inhibitor of Kv1.3 channel pore. Specific Aim 1 seeks to improve the method by creating highly-diverse libraries on native scaffolds that bind to ion channel pores or voltage sensors, first using purified ion channel receptors, and then using channels on cell surfaces so the many receptors that do not retain their structure when purified can also be targeted. A recently identified channel for protons (Hv1) is used here to develop tools for voltage sensors (and to test directly the postulated role of this orphan receptor in male fertility). Specific Aim seeks de novo toxins directed to the Kv1.5 potassium channel. This orphan receptor is widely pursued as a potential therapeutic target for AF and lack of specific blockers has left unclear fundamental questions about its role in the heart and precluded direct tests of its utility as a drg target. Specific Aim 3 seeks de novo toxins specific for the Kv2.1 channel. Kv2.1 inhibition enhances glucose-stimulated insulin secretion from pancreatic -cells in a SUMO-dependent manner and inhibitors that do not cross inhibit are sought as potential drugs for T2D. De novo toxins for Kv1.5 and Kv2.1 are not only useful to validate the channels as drug targets but offer potential as therapeutic leads. Thus, small, disulfide-stabilized peptide toxins in use as drugs (Byetta(r), Integrilin(r), and Prialt(r)) have proven effective, protease resistant and non-immunogenic.

 描述（由申请人提供）：蛋白质神经毒素是研究膜受体的最强大工具之一，现在也用于治疗疾病。这首先寻求资助以推进我们从头创造毒素的方法。通过靶向缺乏这些有用配体的钾和质子通道；其次，使用 de novo 毒素来评估 Kv1.5 和 Kv2.1 通道作为治疗广大公众两种疾病的靶点的潜力；影响 - 心房颤动（AF）和 2 型糖尿病（T2D），这两个方面的成功将允许更广泛地使用这些工具来为其他目标产生毒素，并激发针对对人类健康重要的其他类型的膜受体的类似工作。迄今为止，毒素主要来自有限的来源：粗毒液、毒腺基因克隆和已知毒素的实验室合成。因此，大多数感兴趣的受体缺乏公认的、特定的、这种情况很容易理解：野生毒素的目的和毒液的筛选都不利于目标特异性，通过使用新的、高亲和力的基于所需属性的克隆毒素可以避免这些问题。我们开发的用于生成 Kv1.3 通道孔选择性抑制剂的吞吐量方法具体目标 1 旨在通过在与离子通道孔或电压传感器结合的天然支架上创建高度多样化的库来改进该方法。纯化的离子通道受体，然后使用细胞表面的通道，因此也可以靶向许多纯化后不保留其结构的受体（Hv1），这里使用最近确定的质子通道（Hv1）来开发电压传感器工具（并用于开发电压传感器）。具体目的是寻找针对 Kv1.5 钾通道的从头毒素。这种孤儿受体被广泛作为 AF 的潜在治疗靶点，但目前尚不清楚具体的阻断剂。基本问题具体目标 3 寻找对 Kv2.1 通道具有特异性的新毒素，以增强胰腺  细胞中葡萄糖刺激的胰岛素分泌。 SUMO 依赖性方式和不交叉抑制的抑制剂被寻求作为 Kv1.5 和 Kv2.1 的从头毒素的潜在药物。验证这些通道作为药物靶标，但提供了作为治疗先导化合物的潜力。因此，用作药物的小二硫键稳定肽毒素（Byetta(r)、Integrilin(r) 和 Prialt(r)）已被证明是有效的、具有蛋白酶抗性且具有抗蛋白酶活性。非免疫原性。