Human ion channel pharmacology in droplet bilayer membranes

液滴双层膜中的人体离子通道药理学

• 批准号: 8454192
• 负责人: Jason L. Poulos
• 金额: $ 59.18万
• 依托单位: LIBREDE, INC.
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-15 至 2015-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8454192
• 关键词: Achievement; Address; Adoption; Agreement; Arrhythmia; Artificial Membranes; Automation; Biological Assay; Cardiac; Cardiac Death; Cell membrane; Cells; Contracts; Data; Data Quality; Development; Device or Instrument Development; Dose; Drug Interactions; Drug Targeting; Electrophysiology (science); Equipment; Guidelines; Housing; Human; Industry; Inhibitory Concentration 50; Ion Channel; Libraries; Literature; Market Research; Measurement; Measures; Membrane; Methods; Pain; Performance; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacology; Phase; Physiological Processes; Preparation; Process; Process Measure; Protocols documentation; Reagent; Reporting; Research; Risk; Running; Safety; Simulate; Solutions; System; TRPV1 gene; Technology; Technology Transfer; Testing; Transmembrane Transport; Validation; Work; assay development; base; cost; drug discovery; high throughput screening; instrument; instrumentation; meetings; novel strategies; operation; patch clamp; prototype; public health relevance; relating to nervous system; scale up; screening

DESCRIPTION (provided by applicant): Ion channel drug discovery and safety screening are limited by high cost per data point, low throughput, and complexity. To directly address these industry-wide pain points, Librede Inc. is developing an alternative cell-free technology for ion channel screening, based on the measurement of ion channels in artificial droplet membranes. By eliminating cells at the point of measurement, our technology enables much lower running costs and simplified assay development. In preliminary work, we have used our platform to measure TRPM8 and hERG ion channels and obtain drug IC50 and EC50 values in agreement with state-of-the-art ion channel electrophysiology platforms. In Phase I development of our technology, we created an automation- ready prototype array plate, and verified its reliability and high yield by using it to create over 1000 artificial membranes. In the Phase II work proposed here, we will validate our platform with four ion channels: hERG, Kv7.1, TRPV1, and Cav1.2, selected for their relevance to screening and their degree of complexity. We will optimize the methods of preparing these ion channels from commercially available host cells to achieve highly repeatable and reliable conductance measurements. We will then use these preparations to measure the effects of drugs on the channels' conductance in a dose-dependent manner, generating reliable and repeatable IC50 and EC50 values that we will compare to those obtained from automated patch clamp screening instrumentation. These drug measurements will be performed using the array plates we developed in Phase I, allowing us to evaluate their performance for a simulated screening application. During the proposed work, we will collaborate with the research group of Prof. Jacob Schmidt at UCLA who, with the PI, initially developed the proposed technology. The Schmidt group will aid Librede by initially processing and measuring the ion channel preparations and then transferring this technology to Librede to replicate, optimize, scale-up, and ultimately commercialize. At the completion of this work, we will have validated four highly relevant ion channels pharmacologically in our system and demonstrated its operation in assay conditions. This validation will demonstrate our system's performance to potential end users and allow placement of instrument prototypes with identified beta testers and early adopters. The Phase II work proposed here is a critical step in the development of our new cell-free electrophysiology platform which can address the main industry pain points of cost, throughput, and ease of use (Librede's value proposition), and has the potential to significantly impact ion channel drug discovery and safety screening.

描述（由申请人提供）：离子通道药物发现和安全性筛选受到每个数据点高成本、低通量和复杂性的限制。为了直接解决这些全行业的痛点，Librede Inc.正在开发一种用于离子通道筛选的替代无细胞技术，该技术基于对人工液滴膜中离子通道的测量。通过在测量点消除细胞，我们的技术可以大大降低运行成本并简化分析开发。在初步工作中，我们使用我们的平台测量TRPM8和hERG离子通道，并获得与最先进的离子通道电生理学平台一致的药物IC50和EC50值。在我们技术的第一阶段开发中，我们创建了一个自动化的原型阵列板，并通过使用它创建超过1000个人工膜来验证其可靠性和高产量。在这里提出的第二阶段工作中，我们将用四个离子通道验证我们的平台：hERG，Kv7.1，TRPV 1和Cav1.2，选择它们与筛选的相关性和它们的复杂程度。我们将优化从市售宿主细胞制备这些离子通道的方法，以实现高度可重复和可靠的电导测量。然后，我们将使用这些制剂以剂量依赖性方式测量药物对通道电导的影响，产生可靠且可重复的IC50和EC50值，我们将其与自动膜片钳筛选仪器获得的值进行比较。这些药物测量将使用我们在第一阶段开发的阵列板进行，使我们能够评估它们在模拟筛选应用中的性能。在拟议的工作中，我们将与加州大学洛杉矶分校的雅各布施密特教授的研究小组合作，他与PI一起最初开发了拟议的技术。施密特团队将帮助Librede进行离子通道制备的初步加工和测量，然后将这项技术转让给Librede进行复制、优化、放大并最终商业化。在这项工作完成后，我们将在我们的系统中验证四个高度相关的离子通道，并证明其在测定条件下的操作。该验证将向潜在的最终用户展示我们的系统性能，并允许将仪器原型与已确定的beta测试人员和早期采用者一起放置。本文提出的第二阶段工作是我们开发新的无细胞电生理学平台的关键一步，该平台可以解决成本，吞吐量和易用性（Librede的价值主张）的主要行业痛点，并有可能显着影响离子通道药物发现和安全筛选。