Res Proj 2: Molecular Imaging Strategies to Study Cdc25A Regulation in vivo

Res Proj 2：研究 Cdc25A 体内调节的分子成像策略

• 批准号: 7287031
• 负责人: HELEN M PIWNICA-WORMS
• 金额: $ 24.57万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-28 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7287031
• 关键词: Animals; Cell Cycle; Cell Cycle Checkpoint; Cell Cycle Regulation; Cell Line; Cells; Chimeric Proteins; Clinical Trials; Combined Modality Therapy; Condition; DNA Damage; Diagnosis; Drug Delivery Systems; Enzymes; Exposure to; Firefly Luciferases; Future; Goals; Grant; Holoenzymes; Human; Image; Imaging Techniques; Imaging technology; Knock-in Mouse; Life; Luciferases; Malignant Neoplasms; Mammals; Monitor; Mus; Order Coleoptera; Protein Serine/Threonine Phosphatase; Protein phosphatase; Proteins; Range; Reagent; Regulation; Regulatory Pathway; Specificity; Therapeutic; Therapeutic Intervention; Treatment Protocols; anti-cancer therapeutic; cancer cell; cancer therapy; high throughput screening; high throughput technology; in vivo; inhibitor/antagonist; malignant breast neoplasm; molecular imaging; mouse model; novel; response; stable cell line; tumor

Our broad goals include delineating how the cell division cycle is regulated, determining how cancer cells derail cell cycle regulatory pathways, and ultimately using this information for diagnosing and treating human cancers. We propose to focus our efforts on the Cdc25A protein phosphatase as this enzyme is a key regulator of the cell division cycle in mammals and is overproduced in a wide range of human tumors. Stable cell lines and mouse colonies will be generated that enable Cdc25A regulation to be studied in cells and living mice using molecular imaging technologies. In particular, cell lines that inducibly express a fusion protein between human Cdc25A and firefly luciferase (Cdc25A-FLuc) will be used in a high throughput screen to identify the serine/threonine protein phosphatase holoenzymes that regulate the stability of Cdc25A in vivo. In principal the proteins identified in this study could provide novel targets for imaging agents and therapeutic intervention in cancer treatment. In addition, knock-in mice will be generated that express a fusion protein between endogenous Cdc25A and click beetle red luciferase (Cdc25A-CBRLuc) from the Cdc25A locus. In this way endogenous Cdc25A protein levels can be monitored non-invasively and repetitively in living mice under steady state conditions and in response to DMAdamaging agents and drugs targeting cell cycle checkpoints. In addition, molecular imaging strategies will be applied to mouse models of breast cancer to validate target specificity of rational anti-cancer therapeutic regimens in combination therapies. The reagents and results obtained in these studies will be useful in future clinical trials that combine DNA damaging agents with novel Chk1 inhibitors or novel checkpoint abrogators. The proposed studies are expected to enhance our understanding of basic principles of cell cycle control in mammals and may impact future therapeutic strategies for cancer treatment.

我们的广泛目标包括描述细胞分裂周期是如何调节的，确定癌细胞如何 破坏细胞周期调控途径，并最终利用这些信息进行诊断和治疗 人类癌症。我们建议将重点放在 Cdc25A 蛋白磷酸酶上，因为这种酶是一种 哺乳动物细胞分裂周期的关键调节因子，在多种人类肿瘤中过量产生。 将产生稳定的细胞系和小鼠集落，以便在细胞中研究 Cdc25A 调节 和使用分子成像技术的活体小鼠。特别是，诱导表达融合的细胞系 人类 Cdc25A 和萤火虫荧光素酶 (Cdc25A-FLuc) 之间的蛋白质将用于高通量 筛选以确定调节稳定性的丝氨酸/苏氨酸蛋白磷酸酶全酶 体内 Cdc25A。原则上，本研究中确定的蛋白质可以为成像提供新的靶点 癌症治疗中的药物和治疗干预。此外，将产生敲入小鼠 表达内源性 Cdc25A 和点击甲虫红色荧光素酶 (Cdc25A-CBRLuc) 之间的融合蛋白 来自 Cdc25A 基因座。通过这种方式，可以非侵入性地监测内源性 Cdc25A 蛋白水平 并在稳态条件下对活体小鼠重复进行，并响应 DMA 损伤剂和 针对细胞周期检查点的药物。此外，分子成像策略将应用于小鼠 乳腺癌模型以验证合理抗癌治疗方案的靶标特异性 联合疗法。这些研究中获得的试剂和结果将有助于未来的临床 将 DNA 损伤剂与新型 Chk1 抑制剂或新型检查点废除剂相结合的试验。这 拟议的研究有望增强我们对细胞周期控制基本原理的理解 哺乳动物，并可能影响未来的癌症治疗策略。