Kinome-wide cell-based assays

全激酶组细胞分析

基本信息

批准号：
8780886
负责人：
REENA ZUTSHI
金额：
$ 23.99万
依托单位：
LUCEOME BIOTECHNOLOGIES, LLC
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-01 至 2016-01-14
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8780886
关键词：
Active Sites Adaptor Signaling Protein Apoptosis Architecture Binding Biochemical Biological Biological Assay Cardiovascular Diseases Catalytic Domain Cell Cycle Cell Proliferation Cell division Cell membrane Cell physiology Cells Chemicals Clinic Communities Complex DHFR gene Dasatinib Data Development Diabetes Mellitus Dihydrofolate Reductase Dimerization Disease Drug Design Early identification Economics Escherichia coli Evaluation Failure Fingerprint Genome Goals Human Human Genome Hybrids In Vitro Inflammation Lead Length Libraries Luciferases Luminescent Measurements Measures Mediating Mediator of activation protein Methodology Methods Monitor Neoplasm Metastasis Normal Cell Pathway interactions Permeability Pharmaceutical Preparations Phase Phosphorylation Phosphotransferases Physiology Play Property Protein Kinase Protein Tyrosine Kinase Protein-Serine-Threonine Kinases Reporting Role Scaffolding Protein Serine Services Signal Transduction Testing Threonine Toxic effect Trees Trimethoprim Tyrosine Work base cost cross reactivity design drug candidate drug development drug discovery effective therapy genetic regulatory protein high throughput screening human disease inhibitor/antagonist kinase inhibitor luminescence meetings neoplastic cell public health relevance screening stem success therapeutic target trafficking

项目摘要

DESCRIPTION (provided by applicant): Protein kinases serve as critical mediators of signal transduction in human cells and impact virtually all aspects of cellular physiology, from the coordination of the cell cycle and cell division to apoptosis. The deregulation of kinase mediated signaling is now known to be involved in a variety of human diseases that include, diabetes, inflammation, cardiovascular diseases, tumor cell proliferation and metastasis making them a target for drug development. Structurally, kinases share a very similar architecture at the active-site (ATP-binding domain), making selectivity an issue in drug discovery and development. Thus, the development of rapid and economic methods for profiling drug candidates against a large panel of kinases in whole cells will not only aid in anticipating potential long term toxicit, but also help in understanding kinase targeted polypharmacology and aid in identifying new targets for old compounds. In this Phase I application, we will develop a split-luciferase based luminescent assay for kinase profiling in a cellular setting. These cell-based assays will meet a critical need for directly monitoring the effect of compounds on a target kinase not only to ascertain cell-permeability and cellular toxicity, but also to establish efficacy in the cellular mlieu in the presence of adaptor and regulatory proteins. We will test the generality of the approach for kinases across various groups and evaluate the feasibility of our assay for high throughput screening using a panel of 80 known kinase inhibitors. Our goal is to make cellular kinase profiling assays both easily available and affordable, so that compound profiling in a cellular or native context can be done earlier and thereby lead to early identification of failures, resulting n many more opportunities for success.

描述（由申请人提供）：蛋白激酶是人类细胞信号转导的关键介体，并影响细胞生理学的几乎所有方面，从细胞周期和细胞分裂的配位到凋亡。现在众所周知，激酶介导的信号传导的失调与多种人类疾病有关，包括糖尿病，炎症，心血管疾病，肿瘤细胞增殖和转移，使其成为药物发育的靶标。从结构上讲，激酶在活动位置（ATP结合域）上具有非常相似的结构，使选择性成为药物发现和开发中的问题。因此，开发快速和经济的方法，用于针对整个细胞中大量激酶的候选药物进行分析，这不仅有助于预测潜在的长期毒性，而且还有助于理解激酶靶向的多形能力学，并有助于识别旧化合物的新靶标。在此阶段I应用中，我们将开发一种基于分裂的葡萄酸酶的发光测定，用于在细胞环境中进行激酶分析。这些基于细胞的测定将满足直接监测化合物对靶激酶的影响的关键需求，这不仅是确定细胞 - 渗透性和细胞毒性的影响，而且还可以在衔接蛋白和调节蛋白的存在下在细胞MLIEU中建立功效。我们将测试各组的激酶方法的通用性，并使用80个已知的激酶抑制剂的面板评估我们测定法对高吞吐量筛查的可行性。我们的目标是使细胞激酶分析测定既容易获得又负担得起，以便可以在细胞或本地环境中进行复合谱分析，从而可以尽早确定失败，从而导致更多成功的机会。