Supramolecular Approaches to Protein Inhibitors Activated by MicroRNA-21

MicroRNA-21 激活蛋白质抑制剂的超分子方法

• 批准号: 8466335
• 负责人: Janarthanan Jayawickramarajah
• 金额: $ 27.59万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-05 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8466335
• 关键词: Address; Adverse effects; Antineoplastic Agents; Attenuated; Basic Science; Binding; Biological Assay; Biological Markers; Carbonic Anhydrase II; Cell Line; Cells; Cellular Structures; Chemistry; Chimera organism; Complex; DNA; DNA Structure; DNA-Binding Proteins; Development; Enzymes; Fluorescence; Fluorescence Spectroscopy; Foundations; Goals; Isoenzymes; Knowledge; Lead; Ligands; Malignant Neoplasms; MicroRNAs; Mission; Modification; Molecular Conformation; Oligonucleotides; Oncogenic; Outcome; Patients; Prodrugs; Protein Dynamics; Proteins; Public Health; Research; Resistance; Seminal; Site; Stimulus; Structure; System; Testing; Tissues; Transfection; United States National Institutes of Health; Vertebral column; Vision; Work; aqueous; base; burden of illness; cancer cell; cell growth; chemotherapy; clinically relevant; design; enzyme substrate; esterase; improved; inhibitor/antagonist; innovation; locked nucleic acid; molecular recognition; novel; nuclease; overexpression; protein activation; research study; response; small molecule; stem

DESCRIPTION (provided by applicant): Prodrugs that are triggered by cancer biomarkers are attractive because of their potential to be activated preferentially in cancer cells thereby reducing the serious adverse effects associated with conventional chemotherapy. While numerous cancer biomarkers have been discovered, prodrugs triggered by enzymes have been the main focus because enzymes can activate prodrugs via covalent transformations. Thus, a barrier exists in using non-enzyme biomarkers for prodrug-activation because of our poor understanding of how to develop molecules that selectively recognize, and are activated by, biomarkers that cannot facilitate covalent transformations. The existence of this knowledge gap hinders the full scope of prodrugs that can be developed. Thus the long-term vision of this project is to generate non-covalently triggered prodrugs. The objective of this basic-science application, which is a seminal step in achieving our long term goal, is to establish fundamental supramolecular mechanisms for the non-covalent activation of protein inhibitors by microRNA-21 (miR-21), a highly deregulated, oncogenic, miR. The central hypothesis of this application, which we will test via two complementary design strategies (Aim 1 and 2, respectively), is that miR-21 inducible protein antagonists can be achieved by developing DNA-small molecule chimeras (DCs), that undergo a structural switch from a de-activated conformation to an activated state upon binding sequence specifically to miR-21. The specific aims for this project are to develop, (Aim 1) hairpin-forming DCs de-activated by host/guest interactions and (Aim 2) bidentate DCs de-activated via duplex formation with a guide strand, as miR-21 responsive inhibitors of clinically relevant carbonic anhydrase-II (CA-II). In both aims, we will synthesize and characterize the respective DC systems and will probe their ability to undergo miR-21 induced structure-switching to an activated state via a number of spectroscopic experiments (including UV-vis and fluorescence quenching). Subsequently, the differential ability of the de-activated and active DC conformations to inhibit CA-II will be evaluated (via a colorimetric esterase assay). After optimization of the structure of the DC systems (including enhancing CA-II binding and nuclease resistance), we will transfect DCs into cell-lines over expressing (and control cell-lines under expressing) miR-21 and probe CA-II inhibition-derived cell growth arrest. This proposal is innovative because it develops the first examples of activating protein inhibitors in response to miRs. Further, this project will provide impetus to shift general research into examining non- covalent mechanisms for prodrug activation. This work is significant because (a) it is a seminal step in a continuum of research aimed, in the long-term, at developing miR-21 triggered prodrugs, and (b) it will exert a sustained influence on the field of supramolecular chemistry by enhancing our understanding of how key principles of molecular recognition (such as host/guest interactions and multivalency) in combination with DNA assembly and structure-switching phenomena can be used to develop dynamic protein-binders. )

描述（由申请人提供）：由癌症生物标志物触发的前药很有吸引力，因为它们在癌细胞中优先激活的潜力，从而减少了与常规化学疗法相关的严重不良影响。尽管已经发现了许多癌症生物标志物，但酶触发的前药一直是主要焦点，因为酶可以通过共价转化激活前药。因此，使用非酶生物标志物进行前药激活而存在障碍，因为我们对如何开发如何有选择地识别并被无法促进共价转化的生物标志物进行了有选择性地识别和激活。这种知识差距的存在阻碍了可以开发的前药的全部范围。因此，该项目的长期视野是生成非共价触发前药。这种基本科学应用的目的是实现我们的长期目标的开创性一步，是建立基本的超分子机制，用于通过microRNA-21（miR-21）（一种高度失去管制的，高含量的，致癌的mir）通过microRNA-21（miR-21）对蛋白质抑制剂的非共价激活。该应用的核心假设，我们将通过两种互补设计策略（分别为1和2）进行测试，即可以通过开发DNA-SMALL分子嵌合体（DCS）来实现miR-21诱导蛋白拮抗剂，这些蛋白拮抗剂（DCS）可以从与激活状态进行DE活化的结构转换，从而具有与激活状态的结构切换，从而在与MiR-MiR-21方面具有粘合型的结合状态。该项目的具体目的是开发（AIM 1）由主机/来宾相互作用消除了发夹形成的DC，（AIM 2）双齿DCs通过双工形成与指南链消灭，作为MiR-21响应式抗性抑制剂，用于临床相关的碳氧化和碳氧化碳氧化碳纤维酶-II（CA-II）。在这两个目标中，我们都将合成和表征各自的DC系统，并将通过许多光谱实验（包括UV-VIS和荧光淬灭）探测其将MiR-21诱导的结构切换到激活状态的能力。随后，将评估去激活和活性直流构象抑制CA-II的差异能力（通过比色酯酶测定）。在优化DC系统的结构（包括增强CA-II结合和核酸酶耐药性）之后，我们将通过表达（表达的对照细胞）和探针CA-II抑制衍生的细胞生长的生长量转移到细胞线中。该提议具有创新性，因为它发展了响应miRS激活蛋白质抑制剂的第一个例子。此外，该项目将为将一般研究转移到研究非共价激活机制。 This work is significant because (a) it is a seminal step in a continuum of research aimed, in the long-term, at developing miR-21 triggered prodrugs, and (b) it will exert a sustained influence on the field of supramolecular chemistry by enhancing our understanding of how key principles of molecular recognition (such as host/guest interactions and multivalency) in combination with DNA assembly and structure-switching phenomena can be used to develop动态蛋白质粘合剂。 ）