Cellularly resolved molecular pathway assessment in biopsies via spectral imaging

通过光谱成像进行活检中的细胞解析分子途径评估

基本信息

批准号：
7298872
负责人：
RICHARD M. LEVENSON
金额：
$ 22.79万
依托单位：
CAMBRIDGE RESEARCH AND INSTRUMENTATION
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-09-28 至 2009-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7298872
关键词：
Address Affect Algorithms Animals Antibodies Antigens Archives Area Attention Automation Avastin BAY 54-9085 BIK gene Benign Biological Assay Biological Preservation Biology Biometry Biopsy British Columbia CD34 gene Calibration Cell Nucleus Cell Surface Receptors Cell surface Cells Certification Classification Clinical Clinical Laboratory Information Systems Clinical Research Clinical Trials Colon Carcinoma Computer software Computers Condition Correlative Study Coupled Data Detection Development Diagnosis Discrimination Disease Drug Industry Elements Employment Energy Transfer Engineering Ensure Epidermal Growth Factor Receptor Epitopes Evaluation Event Excision Fixatives Fluorescence Formalin Funding Future Generations Goals Guidelines Harvest Histocompatibility Testing Hour Human Image Image Analysis Imagery Imaging technology Immunofluorescence Immunologic Immunohistochemistry Individual Industry Kidney Label Learning Length Life Light Lighting Localized Location MEKs Machine Learning Malignant - descriptor Malignant Neoplasms Manuals Mediator of activation protein Medical Medicine Membrane Metabolic Methods Modality Modeling Molecular Molecular Target Monitor Mus Nuclear Antigens Nuclear Protein Nuclear Proteins Numbers Operative Surgical Procedures Optics Outcome PTEN gene Paraffin Embedding Pathologist Pathology Pathway interactions Patient Care Patient Focused Care Patient Selection Patients Penetration Pennsylvania Pharmaceutical Preparations Pharmacologic Substance Phase Phosphoproteins Pilot Projects Play Polymers Population Predisposition Preparation Procedures Process Proteins Proteomics Proto-Oncogene Proteins c-akt Protocols documentation Range Reaction Reading Reagent Receptor Activation Reporting Research Research Design Resources Risk Roche brand of trastuzumab Role Sampling Scanning Score Shapes Side Signal Pathway Signal Transduction Signaling Protein Site Slide Source Specificity Specimen Staining method Stains Standards of Weights and Measures System Techniques Technology Testing Tetradecanoylphorbol Acetate Thick Thyroid Gland Time Tissue Fixation Tissue Microarray Tissue Sample Tissues Toxic effect Training Transillumination Translating Treatment outcome Tumor Tissue Ultrasonography United States Food and Drug Administration Universities Validation Variant Vascular Endothelial Growth Factor Receptor Vascular Endothelial Growth Factor Receptor-2 Vascular Endothelial Growth Factors Weight Western Blotting Work angiogenesis base cancer cell commercialization day design drug development follow-up image processing imaging Segmentation improved instrumentation interest lapatinib malignant breast neoplasm melanoma member method development microwave electromagnetic radiation molecular pathology mouse model nanoparticle new technology novel outcome forecast prospective protein expression protein folding research study response sample fixation statistics subcutaneous tissue culture tissue processing tissue/cell culture tool tumor validation studies

项目摘要

DESCRIPTION (provided by applicant): This is a Fast-Track application to provide reliable, cellularly resolved molecular pathway assessment in cancer biopsies to assist pharmaceutical drug development and provision of patient-specific prognosis and therapy guidance ("personalized medicine"). The organizing theme is that the appropriate unit of analysis should be the individual cell as opposed to averaged tumor extracts. To this end, novel technologies will be coupled with careful methods-development. Spectral imaging and advanced image analysis tools will permit multi-target immunohistochemical (IHC) and/or immunofluorescence (IF) detection, at the cellular and subcellular level in intact tissue sections. CRi-developed image processing and machine-learning tools provide automation and sophisticated quantitation options. Multiplexed staining protocols will yield independent, potentially stoichiometric labeling with combined IHC and IF. The sensitivity of all potential markers to variations in tissue handling will be carefully assessed; some may be robust and suitable for archival tissue studies, others will be too labile. Ultrasound-assisted fixation will be tested for its ability to preserve such labile epitopes for use in prospectively acquired tissues. Four or more pathway-related proteins will be detected in tissue sections, on a cell-by-cell basis, even if co-localized and with spectrally overlapping labels. The coordinated subcellular location of the pathway molecules will be also tracked, with simultaneous assessment of cell-surface receptors (e.g., EGFR, VEGF, Her2-neu), downstream signaling proteins and phosphoproteins (e.g., pAKT, pERK), nuclear proteins (e.g., ER, Ki67), and novel players such as protein-folding mediators (e.g. BIP1). The project will combine optimized tissue protocols, multiplexed IHC/IF reagent kits, and unique machine- learning image analysis that can be used to automate region-detection and label-quantitation. All these depend on CRi's multispectral imaging approaches for assessing multiple analytes on a cell-by-cell and cell- compartment basis in tissue sections. Our collaborators will provide small-animal tumor models for early methods development, multiplexed immunohistochemical labeling of pathway proteins in clinical cancer biopsies, access to archived and prospectively acquired tissues from pathway-targeting clinical drug trials, highly informative archival tissue microarrays, access to validated, activation-specific antibodies, ultra-fast tissue fixation, and biostatistics support. The "deliverable" will be a suite of products suitable for clinical use that can provide much-needed valid information on single-cell-based pathway status in an intact tissue context to support pharmaceutical drug development efforts and provide molecularly focused patient care. Significance and lay narrative: The ability to quantitatively evaluate multiple molecular targets and pathways on a cell-by-cell basis, in a single preparation of clinical tissue, is missing from the current toolbox of personalized medicine, which lacks good means of matching novel drugs and drug candidates to specific patients. Conventional molecular pathology methods are typically limited to a single immunohistochemical (IHC) test on a given tissue section or to expensive and time-consuming proteomics or expression-array approaches (which cannot directly report out pathway activation status in cancer cell populations and subpopulations). Multiplexed IHC (including immuno-fluorescence) combined with optimized sample handling protocols to retain pathway proteins and advanced image analysis will enable the unambiguous detection of active signaling pathways, benefiting pharmaceutical research in the selection of patients for better targeted trials and in the monitoring of response, and clinical practice for diagnosis, therapy selection, and monitoring response (i.e., theranostics).

描述（由申请人提供）：这是一个快速通道应用程序，可在癌症活检中提供可靠的细胞解析分子途径评估，以协助药物开发和提供患者特异性预后和治疗指导（“个性化医疗”）。组织主题是，适当的分析单位应该是单个细胞，而不是平均肿瘤提取物。为此，新技术将与仔细的方法开发相结合。光谱成像和先进的图像分析工具将允许在完整组织切片的细胞和亚细胞水平上进行多目标免疫组织化学 (IHC) 和/或免疫荧光 (IF) 检测。 CRi 开发的图像处理和机器学习工具提供自动化和复杂的定量选项。多重染色方案将结合 IHC 和 IF 产生独立的、潜在化学计量的标记。将仔细评估所有潜在标记物对组织处理变化的敏感性；有些可能很稳健，适合档案组织研究，有些则太不稳定。将测试超声辅助固定保留此类不稳定表位以用于预期获得的组织的能力。即使共定位且具有光谱重叠的标签，也会在组织切片中逐个细胞地检测到四种或更多途径相关蛋白。还将跟踪途径分子的协调亚细胞位置，同时评估细胞表面受体（例如 EGFR、VEGF、Her2-neu）、下游信号蛋白和磷蛋白（例如 pAKT、pERK）、核蛋白（例如、ER、Ki67），以及蛋白质折叠介体（例如 BIP1）等新型参与者。该项目将结合优化的组织方案、多重 IHC/IF 试剂盒和独特的机器学习图像分析，可用于自动化区域检测和标签定量。所有这些都依赖于 CRi 的多光谱成像方法，用于在组织切片中逐个细胞和细胞区室评估多种分析物。我们的合作者将为早期方法开发提供小动物肿瘤模型，在临床癌症活检中对通路蛋白进行多重免疫组织化学标记，从通路靶向临床药物试验中获取存档和前瞻性获得的组织，信息丰富的存档组织微阵列，获取经过验证的、激活特异性抗体、超快速组织固定和生物统计学支持。 “交付品”将是一套适合临床使用的产品，可以提供急需的有关完整组织环境中基于单细胞的通路状态的有效信息，以支持药物开发工作并提供以分子为中心的患者护理。意义和通俗叙述：当前的个性化医疗工具箱缺乏匹配新药的良好方法，在临床组织的单一制备中逐个细胞地定量评估多个分子靶点和途径的能力以及针对特定患者的候选药物。传统的分子病理学方法通常仅限于对给定组织切片进行单一免疫组织化学 (IHC) 测试，或昂贵且耗时的蛋白质组学或表达阵列方法（无法直接报告癌细胞群和亚群中的通路激活状态）。多重 IHC（包括免疫荧光）与优化的样品处理方案相结合，以保留通路蛋白和先进的图像分析，将能够明确检测活性信号通路，从而有利于药物研究，选择患者进行更有针对性的试验，并监测反应，以及诊断、治疗选择和监测反应（即治疗诊断）的临床实践。