Direct assessment of GPCR-transducer coupling and G protein subtype bias

GPCR-转导器耦合和 G 蛋白亚型偏差的直接评估

• 批准号: 10239055
• 负责人: Nevin Alan Lambert
• 金额: $ 33.88万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10239055
• 关键词: Address; Adrenergic Receptor; Adverse effects; Agonist; Arrestins; Binding; Biological Assay; Bioluminescence; Cells; Clinical; Communities; Complex; Coupled; Couples; Coupling; Cyclic AMP; Data Set; Development; Drug Prescriptions; Drug Targeting; Energy Transfer; Engineering; Enzymes; Event; FDA approved; Family; Future; G-Protein-Coupled Receptors; GPR3 gene; GTP-Binding Protein alpha Subunits, Gs; GTP-Binding Proteins; Goals; Guanosine Triphosphate Phosphohydrolases; Health; Heterotrimeric GTP-Binding Proteins; Hormones; Human; Individual; Laboratories; Lead; Ligands; Measures; Methods; Molecular; Monitor; Neurotransmitters; Optics; Orphan; Pharmaceutical Preparations; Pharmacology; Physiological; Protein Family; Protein Subunits; Proteins; Receptor Signaling; Reporting; Second Messenger Systems; Signal Pathway; Signal Transduction; Standardization; Structure; Testing; Therapeutic; Therapeutic Agents; Time; Transducers; Visual; Work; arrestin 2; arrestin3; base; cell behavior; drug candidate; drug development; drug discovery; improved; receptor; receptor coupling; response; side effect; tool; treatment response

PROJECT SUMMARY/ABSTRACT G protein-coupled receptors (GPCRs) are important targets of hormones, neurotransmitters and approximately one-third of FDA-approved drugs. These receptors signal by coupling to transducer proteins from four families of heterotrimeric G proteins and a family of four arrestins. It is now recognized that individual GPCRs produce complex signaling responses by interacting with several different transducer subtypes, i.e. that signaling is “pluridimensional”. The broad selectivity of a given GPCR for different transducer proteins defines it's ability to influence cellular behavior, therefore it is important to know which tranducers can be activated by each receptor. This information can be surprisingly difficult to obtain, in large part because transducer coupling is most often inferred indirectly using assays that monitor events that are well downstream from receptors, such as accumulation of second messengers. It is also understood that certain activating ligands (agonists) can specifically promote GPCR coupling to some transducers at the expense of others, a phenomenon most often referred to as “biased agonism”. Biased agonists are especially promising candidates for drug development, and have rapidly advanced into the clinical setting, because they are able to produce therapeutic responses without also producing adverse effects. Almost all biased agonists discovered to date select between G proteins and arrestins as classes, and there are very few ligands that are thought to select between different G protein subtypes. Again, a critical roadblock to discovery and development of G protein subtype-selective drugs is the reliance on downstream signaling assays, which lack the necessary sensitivity to detect subtle ligand bias and are susceptible to signal crosstalk. We will address these problems using a combination of advanced optical tools that allow direct assessment of receptor-transducer coupling. Specifically, we will use improved cellular and cell-free assays based on engineered mini G (mG) proteins to: 1) comprehensively profile GPCR transducer coupling for >200 non-sensory GPCRs, and 2) characterize and discover new G protein subtype-selective ligands. These studies will generate and test hypotheses that are central to pluridimensional signaling, will make a powerful new set of sensitive transducer assays available to the scientific community, and will advance understanding of G protein subtype-biased ligands as potential therapeutic agents.

项目总结/摘要 G蛋白偶联受体（GPCRs）是激素、神经递质和神经递质的重要靶点， 三分之一的FDA批准的药物。这些受体通过与来自四个家族的转导蛋白偶联来发出信号 异源三聚体G蛋白和四个抑制蛋白家族。现在认识到，单个GPCR产生 通过与几种不同的换能器亚型相互作用，即信号传导是复杂的信号传导反应。 “多维”。给定的GPCR对不同转导蛋白的广泛选择性定义了它的能力， 影响细胞行为，因此，重要的是要知道哪些传感器可以被每个激活 受体的这一信息可能令人惊讶地难以获得，在很大程度上是因为换能器耦合是不可预测的。 最常使用监测受体下游事件的测定法间接推断， 作为第二信使的积累。还应理解，某些活化配体（激动剂）可 特别是促进GPCR耦合到一些换能器，而牺牲其他换能器，这种现象最常见的是 被称为“偏向激动”。偏性激动剂是药物开发的特别有希望的候选物， 并迅速进入临床环境，因为它们能够产生治疗反应， 而不会产生不利影响。迄今为止发现的几乎所有偏向性激动剂都选择G 蛋白质和抑制蛋白作为类别，并且很少有配体被认为在不同的G 蛋白亚型同样，发现和发展G蛋白亚型选择性 药物依赖于下游信号分析，缺乏必要的灵敏度来检测微妙的 配体偏置并且对信号串扰敏感。我们将使用以下组合来解决这些问题： 先进的光学工具，允许直接评估受体-换能器耦合。具体来说，我们将使用 基于工程化的miniG（mG）蛋白的改进的细胞和无细胞测定，以：1）全面地 用于>200个非感测GPCR的轮廓GPCR换能器耦合，以及2）表征和发现新的G 蛋白质亚型选择性配体。这些研究将产生和测试的假设是核心， 多维信号，将使一个强大的新的一套敏感的换能器测定可用于 科学界，并将推进对G蛋白亚型偏向配体的理解， 治疗剂。