Connecting the functional effects of drugs to how they change PPAR gamma

将药物的功能效应与其改变 PPAR gamma 的方式联系起来

• 批准号: 8767700
• 负责人: Travis Shane Hughes
• 金额: $ 9万
• 依托单位: SCRIPPS FLORIDA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8767700
• 关键词: 3T3-L1 Cells; Acute; Adipocytes; Adverse effects; Affect; Affinity; Agonist; Animals; Antidiabetic Drugs; Area; Arts; Atherosclerosis; Binding; Biology; Biophysics; Calorimetry; Cell Line; Cells; Cluster Analysis; Collaborations; Complex; DNA; Data; Development; Diabetes Mellitus; Diabetes prevention; Dimerization; Drug Design; Drug Prescriptions; Drug Targeting; Drug effect disorder; Entropy; Environment; FDA approved; Family; Florida; Fluorine; Fracture; Gene Expression; Genes; Genetic Transcription; Glyburide; Goals; Grant; Heart failure; Human; Inflammation; Knowledge; Length; Ligand Binding Domain; Ligands; Light; Link; Location; Maintenance; Manuscripts; Maps; Measurement; Measures; Mentors; Metformin; Methods; Modeling; Molecular Conformation; Motion; Movement; Names; Non-Insulin-Dependent Diabetes Mellitus; Nuclear Magnetic Resonance; Nuclear Receptors; Osteogenesis; Output; PPAR gamma; PPARBP gene; Patients; Peptides; Peroxisome Proliferator-Activated Receptors; Pharmaceutical Preparations; Phase; Physics; Play; Prediabetes syndrome; Preparation; Production; Proteins; RXR; Research; Research Institute; Research Personnel; Role; Sampling; Site; Solutions; Structure; Surface; Techniques; Testing; Thermodynamics; Time; Titrations; Training; Weight Gain; Woman; Work; bone cell; bone loss; cost; diabetic; flexibility; functional outcomes; immune function; immunoregulation; improved; insulin sensitizing drugs; lipid biosynthesis; molecular dynamics; prevent; programs; public health relevance; receptor function; research study; shape analysis; simulation; transcription factor

DESCRIPTION (provided by applicant): Connecting the functional effects of drugs to how they change PPAR? The most effective drugs for treating and preventing Type II diabetes are those that bind to a protein named PPAR?. PPAR? is a transcription factor critical for the production and maintenance of adipocytes and bone cells and it affects immune function. Some PPAR? dependent effects can be beneficial to people with diabetes and some are not. The challenge is to develop PPAR? binding drugs that retain the unique and robust anti-diabetic effects but reduce the side effects of heart failure, weight gain and bone loss. Current research indicates that it may be possible to achieve separation of unwanted and wanted effects by targeting PPAR? with the right drug. This concept is supported by the fact that animal studies show some new PPAR? drugs activate distinct gene sets from currently prescribed drugs. However, how different drugs uniquely change PPAR? in order to produce these drug specific effects are unknown. Development of improved anti-diabetic PPAR? drugs is more likely when it is known how drugs change PPAR? and how these changes produce functional changes such as changes in gene expression. Understanding how ligands produce function in PPAR? and closely related proteins is the long term goal of the principle investigator (PI). This knowledge will aid in development of better drugs for the whole family of PPAR? like proteins (nuclear receptors) which are the target of ~13% of FDA approved drugs. Recently we have discovered that a large region of PPAR? exists in at least two conformations in solution alone or when bound to less efficacious drugs, however one PPAR conformation is detected when bound to a ligand that induces high transcription (Hughes et al. 2012). Importantly, these data are time consuming and expensive to obtain and only indicate that internal movement exists with little other detail. To better understand the link between PPAR?s internal motion and its function in cells we have developed low- cost, rapid NMR methods that can be used on large complexes and a NMR line shape analysis program which reveal in detail the range of conformations present (i.e. the conformational ensemble) at one site within the flexible region of PPAR?. These methods reveal conformational complexity that would be very difficult to observe using other NMR methods (manuscript in preparation) and allow characterization of a sufficient number of PPAR? drugs to draw statistically meaningful conclusions about any correlation between drug induced changes to PPAR? and changes in gene expression in cells. During the independent phase (aim 2) of the project the amount of NMR probe locations will be expanded (from the single current location) to get a more complete picture of PPAR?'s conformational ensemble in different areas. Additionally we will study PPAR? in the two main forms that it is found in the body 1) full length PPAR? (FL-PPAR?) and 2) the full-length heterodimer complex, which consists of PPAR?, RXR? and DNA. This work will reveal in unprecedented detail how ligands affect the range of related structures that comprise the conformational ensemble of PPAR?. However, they will not detect ligand-induced changes in PPAR?'s small fast movements (i.e. conformational entropy) that may be critical to how ligands produce effects in humans. To study these movements the PI will be trained in using molecular dynamics simulations. These simulations will be checked against experiment where possible using NMR. These data will be used to estimate the average regional change in the small fast internal movement of PPAR? (conformational entropy) that occurs when a drug binds PPAR. All of these measurements of drug-induced changes in PPAR? will be tested for correlation with functional outcomes such as dimerization with FL-RXR¿, recruitment of coregulator peptides and gene expression in adipocytes. In order to fully utilize these recent advances and to build the best possible model of how PPAR dynamics and conformation leads to function the PI needs protected time for training in molecular dynamics simulation. The advisory team, which includes Dr. Cheatham, will provide expert guidance in this area. During the mentored portion of the grant (aim 1) the PI will continue to receive guidance in NMR and protein molecular dynamics simulations from collaborators (Drs. Art Palmer and Mark Rance), in addition to the PI's primary mentor Dr. Kojetin. The PI will also receive training in the methods and analysis of PPAR? drug effects on target gene expression in cells from Dr. Griffin (co-mentor). The Scripps Research Institute in Florida (TSRI) has seven groups (Nettles, Griffin, Kojetin, Kameneka, Solt, Rousch and Smith) that study nuclear receptors. Four of these groups are currently using different approaches and methods for answering important questions about PPAR?. This makes collaboration natural and provides an excellent environment in which to receive the training necessary to sustain independent research in this area. The PI has degrees in physics and biology which has allowed him to quickly acquire expertise in many areas of nuclear magnetic resonance (NMR) of proteins and several other biophysical and biology techniques during his 3 years of training at TSRI and provides a breadth of training that will be essential for connecting the biophysics and thermodynamics of PPAR movement and structure to functional outcomes.

描述（由适用提供）：将药物的功能效应与它们如何更改PPAR联系起来？治疗和预防II型糖尿病的最有效药物是与称为PPAR的蛋白质结合的药物。 ppar？是对脂肪细胞和骨细胞的生产和维持至关重要的转录因子，它会影响免疫功能。一些ppar？依赖性影响可能对糖尿病患者有益，有些则无济于事。挑战是开发PPAR吗？保留独特且强大的抗糖尿病作用但减少心力衰竭，体重增加和骨质损失的副作用的结合药物。当前的研究表明，可以通过靶向PPAR来实现不需要和想要的效果的分离？与合适的药物。动物研究表明一些新的PPAR的事实支持了这个概念？药物激活当前处方药的不同基因集。但是，不同的药物如何唯一改变PPAR？为了产生这些药物特异性效果，尚不清楚。开发改善的抗糖尿病PPAR？当知道药物如何改变PPAR时，药物更有可能？这些变化如何产生功能变化，例如基因表达的变化。了解配体如何在PPAR中产生功能？密切相关的蛋白质是原理研究者（PI）的长期目标。这些知识将有助于为整个PPA家族开发更好的药物吗？像蛋白质（核受体）一样，是约13％的FDA批准药物的靶标。最近，我们发现PPAR的大部分地区？至少存在于溶液中或与效率较低的药物结合时，至少存在两个构象，但是当绑定到诱导高转录的配体时，检测到一种PPAR构象（Hughes等，2012）。重要的是，这些数据耗时且获得昂贵，并且仅表明内部运动存在很少的细节。为了更好地理解PPAR的内部运动及其在细胞中的功能之间的联系，我们开发了低成本的快速NMR方法，该方法可用于大型复合物和NMR线形状分析程序，该程序揭示了PPAR柔性区域内的一个位置的构象范围（即构型集合）。这些方法揭示了使用其他NMR方法（准备中的手稿）很难观察到的概念复杂性，并允许表征足够数量的PPAR？关于药物引起的PPAR变化之间有任何相关性的统计意义的药物吗？以及细胞中基因表达的变化。在项目的独立阶段（AIM 2）期间，NMR探针位置的数量将扩大（从单个当前位置），以更完整地了解PPAR？在不同区域的构象合奏。另外，我们将学习PPAR？在体内发现的两种主要形式中1）全长PPAR？ （fl-ppar？）和2）全长异二聚体复合物，哪个由ppar？，rxr组成？和DNA。这项工作将以前所未有的细节揭示配体如何影响构成PPAR构象合奏的相关结构的范围。但是，他们不会检测到配体诱导的PPAR的变化，这对于配体如何在人类中产生影响至关重要，这可能是至关重要的。为了研究这些运动，PI将在使用分子动力学模拟方面进行训练。这些模拟将在可能的情况下使用NMR检查实验。这些数据将用于估计PPAR小快的内部运动的平均区域变化？当药物结合PPAR时发生的（构象熵）。所有这些测量药物诱导的PPAR变化的测量？将测试与功能结果的相关性，例如与fl-rxr dimerization，Coregulator肽的募集以及脂肪细胞中基因表达的相关性。为了充分利用这些最新进展，并建立了最好的模型 PPAR动力学和会议如何导致功能功能，PI需要受保护的时间进行分子动力学模拟的训练。包括Cheatham博士在内的咨询团队将在该领域提供专家指导。在赠款的修订部分（AIM 1）期间，PI将继续从合作者（Art Palmer博士和Mark Rance博士）获得NMR和蛋白质分子动力学模拟指导，此外PI的主要导师Kojetin博士。 PI还将接受PPAR方法和分析的培训吗？药物对Griffin博士（Co-Intor）细胞中靶基因表达的影响。佛罗里达州的Scripps研究所（TSRI）有七组（Nettles，Griffin，Kojetin，Kameneka，Solt，Rousch和Smith），研究了核接收器。这些小组中的四个目前正在使用不同的方法和方法来回答有关PPAR的重要问题。这使得协作自然，并提供了一个绝佳的环境，可以在该环境中接受必要的培训，以维持该领域的独立研究。 The PI has degrees in physics and biology which has allowed him to quickly acquire expert in many areas of nuclear magnetic resonance (NMR) of proteins and several other biophysical and biology techniques during his 3 years of training at TSRI and provides a breadth of training that will be essential for connecting the biophysics and thermodynamics of PPAR movement and structure to functional outcomes.