Targeting Nucleic Acids with an Integrated Vitural and Actual Screen

通过集成的虚拟和实际屏幕靶向核酸

• 批准号: 7194426
• 负责人: Jonathan B. CHAIRES
• 金额: $ 28.75万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-02-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7194426
• 关键词: Adopted; Algorithms; Antineoplastic Agents; Automation; Base Sequence; Binding; Biological Assay; Biological Factors; Cancer Biology; Cells; Computer Simulation; Computer software; DNA; DNA Binding; DNA biosynthesis; Data; Data Set; Development; Development Plans; Dialysis procedure; Diamidine; Docking; Drug Delivery Systems; Equilibrium; Feedback; Fluorescence; Free Energy; G-Quartets; Gene Expression; Generations; Genetic Transcription; Goals; Hybrids; Intercalating Agents; Laboratories; Lead; Libraries; Ligand Binding; Ligands; Liquid substance; Malignant Neoplasms; Maps; Measurement; Measures; Methods; Molecular; Molecular Target; Nucleic Acid Binding; Nucleic Acids; Nucleic acid sequencing; Numbers; Paste substance; Phase; Phase I Clinical Trials; Physical Dialysis; Play; Porphyrins; Publishing; Purpose; Research; Research Personnel; Role; Sampling; Score; Screening procedure; Site; Solutions; Specificity; Structure; System; Telomerase; Testing; Therapeutic Agents; Thermodynamics; Time; Validation; Virtual Library; analytical method; cyanine dye; design; high throughput screening; innovation; intercalation; interdisciplinary approach; interest; novel; novel therapeutics; nucleic acid structure; programs; receptor; repository; small molecule; tool; virtual

DESCRIPTION (provided by applicant): Nucleic acids are under-represented molecular targets for small molecule therapeutic agents, even though recent research shows that unusual nucleic acid structures play a profound role in gene expression and DNA replication. We propose an innovative, interdiscplinary, high-throughput screening approach for the discovery of new nucleic acid-targeted therapeutic agents. Our approach integrates an actual experimental screening assay with a virtual screening assay. The former is a unique competition dialysis assay that we invented for the discovery of small molecules that target specific nucleic acid sequences or structures. In the competiton dialysis assay, an array of designed nucleic acid structures and sequences are dialyzed against a common test ligand solution. At equilibirum, more ligand accumulates in the dialysis cell that contains the preferred structure or sequence. The assay is thermodynamically rigorous, and allows for quantitative measurement of ligand binding free energies. In our integrated approach, an in silico virtual screening assay will be implemented that will contain an array of receptors that is identical to the actual array of nucleic acids used in the competiton dialysis assay. The virtual assay can easily screen millions of compounds for their selective binding to particular nucleic acid structures of functional significance. "Hits" from the virtual screen will be passed to the actual competition dialysis assay for validation. With proposed improvements, the competiton dialysis can rapidly screen hundreds of compounds. Our strategy will be to identify libraries of compounds that selectively recognize particular structures with the virtual screen, then to rigorously verify these "hits" by high-throughput competition dialysis. Specific plans for the development of the competition dialysis assay include : i) implementation of the assay in a 96-well plate format (facilitating automation), ii) reduction of sample volumes, iii) expansion of the biologically significant nucleic acid array, and iv) reduction of throughput time. The virtual and actual screens will be fully integrated by an iterative feedback loop to refine the scoring algorithm, using actual data obtained from the first generation dialysis assay in which the interaction of 126 compounds with 13 nucleic acids structures was studied. Our approach matches the Road Map Initiative goals by its interdisciplinary approach and by its emphasis on expanding the toolbox available for the discovery of new therapeutics.

描述（由申请人提供）：核酸是小分子治疗剂的代表性不足的分子靶标，尽管最近的研究表明，不寻常的核酸结构在基因表达和DNA复制中起着重要作用。我们提出了一种创新的，跨学科的，高通量的筛选方法，用于发现新的核酸靶向治疗药物。我们的方法集成了一个实际的实验筛选测定与虚拟筛选测定。前者是我们发明的一种独特的竞争透析测定法，用于发现靶向特定核酸序列或结构的小分子。在竞争透析测定中，将设计的核酸结构和序列的阵列针对共同的测试配体溶液透析。在平衡时，更多的配体在含有优选结构或序列的透析池中积累。该测定法在化学上是严格的，并且允许定量测量配体结合自由能。在我们的综合方法中，将实施计算机虚拟筛选测定，其将包含与竞争透析测定中使用的实际核酸阵列相同的受体阵列。虚拟检测可以很容易地筛选数百万种化合物，以确定它们与具有功能意义的特定核酸结构的选择性结合。来自虚拟屏幕的“命中”将被传递到实际竞争透析测定以进行验证。通过改进，竞争透析可以快速筛选数百种化合物。我们的策略将是识别化合物库，选择性地识别特定的结构与虚拟屏幕，然后严格验证这些“命中”的高通量竞争透析。开发竞争透析测定的具体计划包括：i）以96孔板形式实施测定（促进自动化），ii）减少样品体积，iii）扩增具有生物学意义的核酸阵列，以及iv）减少通过时间。虚拟和实际筛选将通过迭代反馈循环完全整合，以使用从第一代透析测定中获得的实际数据来完善评分算法，其中研究了126种化合物与13种核酸结构的相互作用。我们的方法符合路线图倡议的目标，其跨学科的方法，并强调扩大工具箱可用于发现新的治疗方法。