Single-Cell Protein Activity Profiling in Breast Cancer Cells and Tissues

乳腺癌细胞和组织中的单细胞蛋白质活性分析

• 批准号: 10413516
• 负责人: Raymond E Moellering
• 金额: $ 37.35万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-08 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10413516
• 关键词: Address; Advanced Development; Benchmarking; Biological; Biological Markers; Biopsy; Breast; Breast Cancer Cell; Breast Cancer Patient; Cancer cell line; Cell Line; Cells; Chemicals; Clinic; Clinical; Complex; Data; Development; Diagnosis; Disease; Early Intervention; Elements; Enzymes; Family; Future; Goals; Grant; Human; Image; In Situ; Individual; Knowledge; Ligation; Malignant Neoplasms; Mammary Neoplasms; Measurement; Measures; Messenger RNA; Methods; Microfluidics; Molecular; Molecular Probes; Molecular Profiling; Neoplasm Circulating Cells; Neoplasm Metastasis; Operative Surgical Procedures; Organism; Outcome; Patient Care; Patients; Phosphotransferases; Positioning Attribute; Primary Neoplasm; Protein Family; Proteins; Proteome; Proteomics; Publishing; Punch Biopsy; Reagent; Reporting; Research; Resolution; Sampling; Services; Severity of illness; Signal Transduction; Specimen; Technology; Testing; Therapeutic; Tissue Sample; Tissues; Tumor Tissue; Validation; Visualization; anticancer research; base; cancer cell; chemoproteomics; clinical application; design; detection platform; enzyme activity; human disease; improved; innovation; malignant breast neoplasm; molecular phenotype; novel; protein biomarkers; single cell proteins; translational applications; treatment response; tumor

Project Summary Advances in molecular profiling technologies have enabled the identification of ever-increasing numbers of disease biomarkers, each raising the potential for more precise, and possibly even patient-specific, diagnoses. Current methods of quantifying protein biomarkers, however, are slow and laborious, often require large quantities of biopsied tissues, and ultimately fail to provide a complete, intact quantification of functional protein activity. To address these limitations, we have developed an innovative activity dependent proximity ligation platform (ADPL), with the ultimate goal of obtaining multiplexed, ultrasensitive quantification of protein activity directly in complex biological samples. In the context of cancer specimens, we hypothesize that defining a molecular profile based on protein activity – the end result of biologic signaling – can more precisely and accurately inform diagnoses and treatment regimes, allowing early intervention and improving clinical outcomes. This proposal outlines the advanced development and validation of two reliable, high-resolution ADPL-based platforms, with the power to alternately visualize and quantify dozens of active enzymes simultaneously from tumor tissue, cells and biofluids. Our current knowledge and expertise in the use of this integrated platform will be leveraged in the service of the following aims: 1) development of novel chemoproteomic probes for multiplexed, family-wide activity mapping of cancer-relevant enzymes, including kinases; 2) advanced development of on-tissue Exchange-ADPL and on-chip single-cell ADPL (scADPL) platforms for simultaneous, high resolution visualization and quantification of a panel of active enzymes in situ; and 3) applying these technologies to breast cancer tissues and cells to create accurate and precise activity signatures and identify potential correlations between these signatures and clinical outcomes. To accomplish these aims we will develop new chemical proteomic molecular probes, validate a panel of ADPL reagents for cancer-associated enzyme biomarkers, and perform rigorous, metric-driven benchmarking analyses in cell lines and tumor tissues. Completion of this project will validate ADPL platforms for use in diverse cancers, provide the first single-cell activity profiles of breast tumors, and provide novel correlations between breast cancer tumor activity signatures, disease severity and response to therapy.

项目摘要 分子分析技术的进步已使数量不断增加 疾病生物标志物，每种都提高了更精确甚至可能特定于患者的诊断的潜力。 然而，当前量化蛋白质生物标志物的方法缓慢而费力，通常需要大的 数量活检组织，最终无法提供功能蛋白的完整，完整的定量 活动。为了解决这些局限 平台（ADPL），其最终目标是获得蛋白质活性的多重超敏化定量 直接在复杂的生物样品中。在癌症标本的背景下，我们假设定义 基于蛋白质活性的分子谱 - 生物信号传导的最终结果 - 可以更精确，并且 准确地为诊断和治疗方案提供了信息，从而可以提早干预并改善临床结果。 该提案概述了两个可靠的高分辨率基于ADPL的高级开发和验证 平台，只需从 肿瘤组织，细胞和生物流体。我们目前在使用此集成平台方面的知识和专业知识将 在以下目的的服务方面掌握：1）为新的化学蛋白质组问题发展 包括激酶在内的癌症相关酶的多路复用，全家的活性图； 2）高级 开发组织内交易所-ADPL和片上单细胞ADPL（SCADPL）平台，以简单， 高分辨率的可视化和定量一组活性酶原位； 3）应用这些 乳腺癌组织和细胞的技术，以创建准确，精确的活动特征并确定 这些特征与临床结果之间的潜在相关性。为了实现这些目标，我们将发展 新的化学蛋白质组学分子问题，验证与癌症相关酶的ADPL试剂板 生物标志物，并在细胞系和肿瘤组织中进行严格的公制基准测试分析。 该项目的完成将验证ADPL平台用于潜水员癌症，提供第一个单细胞 乳腺肿瘤的活性谱，并提供乳腺癌肿瘤活性特征之间的新相关性， 疾病的严重程度和对治疗的反应。