Isogenic Cell-Based Screen:Cancer-Targeting Catenin(RMI)

描述（由申请人提供）：在广泛的常见人类癌症中发现了B-连环蛋白癌基因的激活突变，包括结肠直肠癌、肝细胞癌、子宫内膜癌、甲状腺癌等。激活的癌基因如B-连环蛋白是一个有吸引力的癌症药物靶点，因为它们代表了在癌细胞中发现而不是在患者正常组织中发现的所需的遗传功能获得。与此想法一致，我们先前已通过实验证明，靶向致癌B-连环蛋白可能是有效的抗癌策略（Mol Can Ther 1：1355，2002）。在本申请中，我们提出实施基于细胞的筛选以鉴定小分子和天然产物先导化合物，其对携带致癌B-连环蛋白等位基因的人结肠癌细胞具有特异性细胞毒性/细胞抑制作用。为此，我们将采用一组成对的基因靶向的人类癌细胞，其差异仅在于存在或不存在突变（致癌）B-连环蛋白的内源性等位基因。由于细胞在其他方面是等基因的，特异性杀死或抑制亲本细胞（携带致癌B-连环蛋白）生长的化合物，而不是其中致癌B-连环蛋白已通过基因靶向缺失的衍生物，是作为B-连环蛋白靶向先导化合物的良好候选物。这些命中将在各种二次筛选中进一步验证，以证明重现性和普遍性。一旦通过筛选大约4000种化合物证明了可行性，我们将申请将筛选转移到NIH Roadmap筛选中心进行高通量筛选工作。

DESCRIPTION (provided by applicant): Activating mutations in the B-catenin oncogene are found in a wide range of common human cancers, including colorectal carcinoma, hepatocellular carcinoma, endometrial carcinoma, thyroid carcinoma, and others. Activitated oncogenes such as B-catenin are an appealing cancer drug target, since they represent a required genetic gain-of-function found in cancer cells but not in the patient's normal tissues. Consistent with this idea, we have previously experimentally demonstrated that targeting oncogenic B-catenin is likely to be an effective anticancer strategy (Mol Can Ther 1:1355, 2002). In this application we propose to implement a cell-based screen to identify small molecule and natural product lead compounds that are specifically cytotoxic/cytostatic to human colon cancer cells harboring an allele of oncogenic B-catenin. To do this we will employ a paired set of gene-targeted human cancer cells that differ only in the presence or absence of their endogenous allele of mutant (oncogenic) B-catenin. Since the cells are otherwise isogenic, compounds that specifically kill or inhibit the growth of parental cells (harboring oncogenic B-catenin) but not the derivatives in which oncogenic B-catenin has been deleted by gene targeting are good candidates as B-catenin targeted lead compounds. Such hits will be further validated in a variety of secondary screens to demonstrate reproducibility and generalizability. Once feasibility has been demonstrated by screening approximately 4000 compounds, we will apply to transfer the screen to an NIH Roadmap screening center for high throughput screening efforts.