Id Protein Assay for Diagnosis & Drug Screening

用于诊断的 Id 蛋白测定

• 批准号: 6741681
• 负责人: JOHN CHEN
• 金额: $ 9.87万
• 依托单位: BIOCHECK, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-03-09 至 2004-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6741681
• 关键词: angiogenesis inhibitors; antineoplastics; autoradiography; biotechnology; biotherapeutic agent; breast neoplasms; diagnosis design /evaluation; drug discovery /isolation; drug screening /evaluation; enzyme linked immunosorbent assay; genetically modified animals; high throughput technology; laboratory mouse; method development; neoplasm /cancer diagnosis; prostate neoplasms; protein protein interaction; protein quantitation /detection; recombinant proteins; transcription factor

DESCRIPTION (provided by applicant): The Idl &ld3 genes and proteins are required for tumor-associated angiogenesis. We hypothesize that inhibiting the activity of the Idl and/or Id3 transcription activators will retard the growth & metastases of tumors. However, the effort to discover an orally active, small molecule, anti-cancer drug by identifying an molecule that inhibits the action of the Idl & Id3 proteins requires bioanalytical methods to screen for small molecules that inhibit the binding of Idl & Id3 to their natural ligands, the E-proteins, and also to establish which cancers would be most likely to respond to an anti-ld treatment. Such bioanalytical method are not currently available for the Id proteins, but we recently obtained promising preliminary results by monitoring the extent of the interaction of horseradish peroxidase-labeled E47 with Idl captured by an anti-ldl antibody bound to microtiter plates. This procedure resembles a traditional ELISA, but uses a natural protein ligand (E47) to the analyte (Idl) instead of a second antibody to detect capture. When fully developed and validated, this method will be used to both measure Idl & Id3 in the blood of patients with certain advanced cancers to determine whether it has potential as a diagnostic method, and also to evaluate the ability of small molecule inhibitors to reduce Id activity. To advance this effort, we plan to (1) validate the pilot assay with respect to limit of quantitation, precision, accuracy, reproducibility, repeatability and stability, (2) use the validated assay to determine if Id concentrations can be reliably measured in the serum and tissue from different mouse models of human cancer, and (3) use the validated assay, configured with fixed & physiologically relevant concentrations of Idl and E47 to explore disruption of Idl binding to E47 at various concentrations of candidate small molecule inhibitors of binding. It is expected that completion of the research described in this grant will result in an assay useful for high throughput screening of chemical libraries for anti-ld activity, and in the selection of those cancers, and perhaps stages of cancer, where anti- Id treatment is most likely to be effective. The assay would, at a minimum, have the potential to be developed into a practical diagnostic/prognostic product to select patients for clinical trials and to monitor their response to anti-ld therapy. It is also possible that the assay could be a commercial product depending on the technical and medical success achieved.

描述（由申请人提供）：Id1&ld3基因和蛋白质是肿瘤相关血管生成所必需的。我们假设抑制Id1和/或Id3转录激活剂的活性将延缓肿瘤的生长和转移。然而，通过鉴定抑制 Idl 和 Id3 蛋白作用的分子来发现口服活性小分子抗癌药物的努力需要生物分析方法来筛选抑制 Idl 和 Id3 与其天然结合的小分子。配体、E-蛋白，并确定哪些癌症最有可能对抗 ld 治疗产生反应。这种生物分析方法目前不适用于 Id 蛋白，但我们最近通过监测辣根过氧化物酶标记的 E47 与结合到微量滴定板的抗 ldl 抗体捕获的 Idl 相互作用的程度，获得了有希望的初步结果。该过程类似于传统的 ELISA，但使用分析物 (Idl) 的天然蛋白质配体 (E47) 而不是第二抗体来检测捕获。当完全开发和验证后，该方法将用于测量某些晚期癌症患者血液中的 Idl 和 Id3，以确定其是否具有作为诊断方法的潜力，并评估小分子抑制剂降低 Id 的能力活动。为了推进这项工作，我们计划 (1) 在定量限度、精密度、准确性、再现性、重复性和稳定性方面验证试点测定，(2) 使用经过验证的测定来确定是否可以在实验中可靠地测量 Id 浓度。来自不同人类癌症小鼠模型的血清和组织，以及（3）使用经过验证的测定法，配置有固定且生理相关浓度的 Idl 和 E47，以探索不同浓度的候选小分子抑制剂对 Idl 与 E47 结合的破坏。绑定。预计本次资助中描述的研究的完成将产生一种可用于高通量筛选化学文库抗Id活性的测定方法，并可用于选择那些癌症，也许还有癌症的阶段，其中抗Id治疗最有可能有效。该检测至少有可能被开发成实用的诊断/预后产品，以选择患者进行临床试验并监测他们对抗ld疗法的反应。该检测也有可能成为商业产品，具体取决于所取得的技术和医学成功。