Kinome Drug Bioassay Platform

描述(申请人提供)：蛋白激酶在信号转导中起关键作用，其功能障碍是癌症、炎症、糖尿病等疾病的关键因素。Kinome药物发现是制药行业的主要焦点，而高通量筛选(HTS)是发现铅的主要工具。反应生物公司(RBC)的Discovery Dot小型化技术不仅将促进药物发现过程，还将节省数百万美元的成本。RBC已经开发了用途极其广泛的纳升反应微阵列，以服务于超超高温药物发现、大规模IC50测定和大规模IC50选择性/毒性分析市场。这些反应比目前在药物发现中广泛使用的孔板形式小1000到10000倍。通过接触式打印的力量，RBC每周能够打印和激活300多万个反应。目前，单个微阵列可以进行3072个药物筛选反应。在数百个重复组中打印大型化学库以进行激酶和磷酸酶筛选的能力可以推动化学蛋白质组研究、环境毒理学、HIT发现和优先排序、铅选择和ADMET。在初步研究中，RBC已经证明了使用印刷微阵列对人类蛋白激酶(c-src，pKA)进行校准筛选的能力。通过第一阶段的资助，拟议的研究将集中于以下三个目标：目的1利用Z‘&gt；0.6验证5种酪氨酸激酶和5种丝氨酸/苏氨酸激酶的分析，并进一步优化，以实现行业中最低成本的分析；目的2设计、实施和验证适用于基于微阵列的化学文库筛选的激酶和磷酸酶的新的分析方法；目的3使用标准化的筛选文库，对照井板分析，验证微阵列上的激酶和磷酸酶的HTS。这项工作将使第二阶段的活动成为可能，在此阶段，反应生物学将使用针对大型合成和自然文库的通用激酶分析，扩大其在人类基因组中的分析范围。

DESCRIPTION (provided by applicant): Protein kinases play pivotal roles in signal transactions, and dysfunction of them is a key factor for diseases like cancer, inflammation, diabetes etc. Kinome drug discovery is a major focus in pharmaceutical industry, and high throughput screening (HTS) is a major tool for lead discovery. Reaction Biology Corporation's (RBC) Discovery Dot miniaturization technology will not only facilitate the drug discovery process but also save the cost by millions. RBC has developed extremely versatile nanoliter reaction microarrays to serve markets for ultra uHTS drug discovery, large scale IC50 determinations, and large scale IC50 selectivity/toxicity profiling. These reactions are 1000 to 10,000-fold smaller than well plate formats currently used widely in drug discovery. Through the power of contact printing, RBC has the capacity to print and activate over 3 million reactions per week. Currently, 3072 drug screening reactions can be carried out on a single microarray. The ability to print large chemical libraries in hundreds of replicates sets for kinase and phosphatase screening can drive chemical-proteomic research, environmental toxicology, hit discovery and prioritization, lead selection, and ADMET. In preliminary studies, RBC has demonstrated the ability to conduct calibrated screening of human protein kinases (c-src, PKA) using printed microarrays. Through Phase I funding, the proposed research will focus on the following three aims: Aim 1 Validation of 5 tyrosine kinase and 5 serine/threonine kinase assays with Z'>0.6 and further optimization to achieve the lowest cost assays in the industry; Aim 2 Design, implementation, and validation of new assay methods for kinases and phosphatases that are amenable to microarray-based screening of chemical libraries; Aim 3 Validation of kinase and phosphatase HTS on microarrays against well-plate assay using a standardized screening library. This work will enable Phase II activities where Reaction Biology will expand its assay range across the human kinome using a universal kinase assay against large synthetic and natural libraries.