Kinome-wide Cell-Based Assays

全激酶组细胞检测

基本信息

批准号：
9053369
负责人：
REENA ZUTSHI
金额：
$ 87.37万
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 2017-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9053369
关键词：
Academia Active Sites Address Adverse effects Apoptosis Architecture Binding Binding Proteins Binding Sites Biochemical Biological Assay Cardiovascular Diseases Cell Cycle Cell Membrane Permeability Cell Proliferation Cell membrane Cell physiology Cells Cleaved cell Client Clinic Clinical Cloning Communities Complex Cysteine Data Development Diabetes Mellitus Disease Distal Drug Design Drug Efflux Drug Transport Early identification Evaluation Failure Family Fingerprint Glean Goals Human Human Genome Hybrids In Vitro Individual Industry Inflammation Lead Length Letters Libraries Luciferases Luminescent Measurements Measures Mediating Monitor Neoplasm Metastasis Permeability Pharmaceutical Preparations Phase Phosphorylation Phosphotransferases Play Protein Kinase Proteins Reporting Rheumatoid Arthritis Role Scaffolding Protein Serine Services Signal Transduction Threonine Toxic effect Tyrosine Validation assay development base cost cross reactivity design drug development drug discovery effective therapy high throughput screening human disease infancy inhibitor/antagonist kinase inhibitor luminescence meetings mutant neoplastic cell novel therapeutics oncology phase 1 study public health relevance screening small molecule stem success therapeutic target

项目摘要

DESCRIPTION (provided by applicant): The human genome encodes 518 protein kinases that catalyze the phosphorylation of client proteins. Signal transduction mediated by protein impacts virtually all aspects of cellular physiology, from the coordination of the cell cycle and cll division to apoptosis. Not surprisingly, the deregulation of kinases is implicated in many diseases including diabetes, inflammation, cardiovascular diseases, tumor cell proliferation and metastasis, making them a target for drug development. Small molecule drug discovery targeting protein kinases has seen enormous success in the past decade with over 25 drugs approved for oncology and most recently rheumatoid arthritis with many more in the pipeline for diverse indications. However, the similarity of the ATP binding active site in kinases targeted by small molecules continues to make selectivity a significant concern in drug discovery and development, since a promiscuous drug is expected to give rise to undesired adverse-effects, especially for long term therapy. In this Phase II application, we will develop a split-luciferase based luminescent assay for kinome-wide screening and profiling of inhibitors in a cellular setting. Currently, there are several kinome scale in vitro biochemical assays that are used for screening and identification of kinase inhibitors, but these assays do not provide any information on the ability of a molecule to cross cell membranes or engage its desired target in a cell. In addition, most biochemical assays use the catalytic kinase domain, thereby limiting the identification of potentially more selective allosteric inhibitors that bind sites distal to the AT-binding site. We hypothesize that the development of kinome wide cell-based assays, which are in its infancy, will meet a critical need. The cell-based kinome assays proposed herein will allow for directly monitoring the effect of compounds on a target kinase to not only to ascertain cell-permeability and cellular toxicity, but also to establish efficacy in the cellular milieu in the presence of other ATP binding proteins. Our goal is to render 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 in many more opportunities for success and new and safer clinical candidates.

描述（由适用提供）：人类基因组编码518个蛋白激酶，可催化客户蛋白的磷酸化。蛋白质介导的信号转导影响细胞生理学的几乎所有方面，从细胞周期和CLL分裂的协调到凋亡。毫不奇怪，激酶的放松管制与许多疾病有关，包括糖尿病，炎症，心血管疾病，肿瘤细胞增殖和转移，使它们成为药物发育的靶标。在过去的十年中，靶向蛋白激酶的小分子药物发现靶向蛋白激酶的成功增强了成功，超过25种用于肿瘤学和最近的类风湿关节炎的药物，其中有更多的潜水员适应性。但是，小分子靶向的激酶中ATP结合活性位点的相似性继续使选择性成为药物发现和发育的重要问题，因为预计滥交药物会引起不良的不良影响，尤其是长期治疗。在此II期应用中，我们将开发一种基于分裂的脂肪酶的发光测定法，以在细胞环境中对抑制剂进行抑制剂的筛查和分析。当前，有几种用于筛查和鉴定激酶抑制剂的体外生化测定法，但这些测定没有提供有关分子跨细胞膜或在细胞中接合其所需靶标的能力的任何信息。此外，大多数生化测定方法都使用催化激酶结构域，从而限制了潜在的更有选择性的变构抑制剂，该抑制剂结合了与AT结合位点远端的位点结合。我们假设，尚处于起步阶段的基于Kinome宽的细胞测定法的发展将满足至关重要的需求。本文提出的基于细胞的Kinome测定法将允许直接监测化合物对靶激酶的影响，不仅可以确定细胞 - 透露率和细胞毒性，还可以在存在其他ATP结合蛋白的情况下确定细胞环境的效率。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分析，从而可以尽早确定失败，从而获得更多成功的机会以及新的和更安全的临床候选者。