VALIDATION AND QUANTIFICATION OF FFPE ANTIGEN RETRIEVAL BY PROTEOME ANALYSIS

通过蛋白质组分析验证和定量 FFPE 抗原回收

• 批准号: 7677468
• 负责人: Satya Prakash Saxena
• 金额: $ 75万
• 依托单位: CALIBRANT BIOSYSTEMS, INC.
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7677468
• 关键词: Actins; Antibodies; Antigens; Archives; Area; Back; Biliary; Bioinformatics; Biological; Biological Assay; Biological Markers; Biological Markers; Biological Products; Biopsy; Blood Cells; Blood capillaries; Breast; Breast Carcinoma; Buffers; California; Carcinoma; Cataloging; Catalogs; Cells; Clinical; Clinical Trials; Collagen; Control Groups; Coupling; Critical Pathways; DNA; Databases; Detection; Development; Diagnosis; Diagnostic; Digestion; Disease; Disease Outcome; Drug Approval Processes; Duct (organ) structure; Ductal Epithelium; E-Cadherin; Electrospray Ionization; Epithelial Cells; Epitopes; Evaluation; Fixatives; Formalin; Foundations; Functional disorder; Furuncles; Generations; Growth; Guidelines; Hepatocyte; Histopathology; Human; Human Resources; Immunohistochemistry; In Situ Hybridization; Individual; Industry; Inflammatory; Investigation; Ions; Isoelectric Focusing; Ki-67 Antigen; Knowledge; Kupffer Cells; Label; Lasers; Lead; Length; Libraries; Liquid Chromatography; Liver; Lymphoplasmacytic Infiltrate; Malignant Neoplasms; Mammary Gland Parenchyma; Maps; Mass Fragmentography; Measurement; Measures; Medical; Methodology; Microdissection; Modeling; Molecular Analysis; Molecular Profiling; Monitor; Morphology; Mucinous; Myosin Heavy Chains; Nature; Noninfiltrating Intraductal Carcinoma; Normal tissue morphology; Outcome; Paraffin; Paraffin Embedding; Patients; Peptide Library; Peptides; Phase; Phase I Clinical Trials; Ploidies; Population; Population Control; Preclinical Testing; Process; Progress Reports; Proteins; Proteome; Proteomics; RNA; Reaction; Recording of previous events; Reporting; Reproducibility; Research; Resources; Retrieval; Running; Sampling; Smooth Muscle; Solutions; Specimen; Staining method; Stains; Stromal Cells; Structure; System; TP53 gene; Techniques; Technology; Therapeutic; Tissue Banking; Tissue Banks; Tissue Fixation; Tissues; Trypsin; Two-Dimensional Polyacrylamide Gel Electrophoresis; Universities; Validation; Variant; anticancer research; base; calponin; capillary; cell type; clinical Diagnosis; comparative; cost; crosslink; expectation; human disease; improved; insight; laser capture microdissection; link protein; macromolecule; medical schools; mucoid; neoplastic cell; novel; outcome forecast; prevent; product development; prognostic; protein expression; public health relevance; response; technology development; therapy outcome; tissue fixing; tissue processing; tool; tumor; tumor eradication; validation studies

DESCRIPTION (provided by applicant): Because of the long history of the use of formalin as the standard fixative for tissue processing in histopathology, there are a large number of archival formalin-fixed and paraffin-embedded (FFPE) tissue banks worldwide. These FFPE tissue collections, with attached clinical and outcome information, present invaluable resources for conducting retrospective protein biomarker investigations. In addition to sample amount constraints imposed by current proteome techniques including two-dimensional polyacrylamide gel electrophoresis and multidimensional liquid chromatography system, the lack of optimized methodologies for retrieving proteins from FFPE tissues further restricts the ability to perform the molecular analysis of archival tissues. By collaborating with Drs. Shan-Rong Shi and Clive R. Taylor from the University of Southern California (USC) Keck School of Medicine during the R41 Phase I studies, the combination of antigen retrieval (AR) with Gemini proteomic technologies not only accomplished the rigorous evaluation of the quality and the reproducibility of proteins retrieved from FFPE tissues for the optimization of AR methodology, but also demonstrated significant opportunities in the pursuit of biomarker discovery using archived FFPE tissue collections. The proposed synergistic efforts between Calibrant and the USC team during the R42 Phase II project aim to generate proteotypic peptide libraries among model and tumor FFPE tissues. These proteotypic peptide libraries represent the first step toward globally cataloging antigens retrievable from FFPE tissues and presenting the available epitope database for subsequent immunohistochemistry (IHC) antibody development. Besides providing guidelines for practitioners of IHC to select the optimized AR condition/antibody combination, the comparative proteomic and validation studies involving the use of proteotypic peptide libraries and associated antibodies will provide further enhancements in the reproducibility and the sensitivity of quantitative IHC measurements. The demand for quantitative IHC continues to escalate due to the widespread utilization of IHC in clinical diagnosis/prognosis and translational cancer research. Furthermore, the greatest expectations for targeted proteomics research using enriched and selected cells from high quality specimens reside in the identification of diagnostic, prognostic, and predictive biological markers in the clinical setting and during preclinical testing and clinical trials, as well as the discovery and validation of new protein targets in the biopharmaceutical industry. The Critical Path Opportunity Report released by FDA in March 2006 not only serves as the first specific blueprint for the Critical Path Initiative, an effort to streamline the drug-approval process by applying new strategies and technologies, but also highlights biomarker development as one of the "most important areas for improving medical product development." Public Health Relevance Statement: By joining Calibrant's unique tissue proteome capabilities with the expertise of Dr. Clive R. Taylor at the University of Southern California in antigen retrieval-immunohistochemistry, the proposed research not only aims to evaluate and optimize quantitative immunohistochemistry measurements through the creation of proteotypic peptide libraries, but also focuses on further development and demonstration of a novel biomarker discovery paradigm for enabling comprehensive and comparative proteomic analysis of archived formalin-fixed and paraffin-embedded tissue collections in support of cancer research, diagnosis, and treatment.

描述（由申请人提供）：由于福尔马林作为组织病理学中组织处理的标准固定剂的使用有着悠久的历史，因此全世界有大量的档案福尔马林固定石蜡包埋（FFPE）组织库。这些 FFPE 组织集合以及附加的临床和结果信息为进行回顾性蛋白质生物标志物研究提供了宝贵的资源。除了当前蛋白质组技术（包括二维聚丙烯酰胺凝胶电泳和多维液相色谱系统）所施加的样品量限制之外，缺乏从 FFPE 组织中检索蛋白质的优化方法进一步限制了对档案组织进行分子分析的能力。通过与博士合作。南加州大学 (USC) Keck 医学院的 Shan-Rong Shi 和 Clive R. Taylor 在 R41 I 期研究中，抗原修复 (AR) 与 Gemini 蛋白质组技术的结合不仅完成了严格的质量评估以及从 FFPE 组织中检索蛋白质以优化 AR 方法的可重复性，而且还证明了使用存档的 FFPE 组织集合追求生物标志物发现的重大机会。 Calibrant 和 USC 团队在 R42 II 期项目期间提出的协同努力旨在在模型和肿瘤 FFPE 组织中生成蛋白肽库。这些蛋白型肽库代表了对可从 FFPE 组织中检索的抗原进行全球编目并为后续免疫组织化学 (IHC) 抗体开发提供可用表位数据库的第一步。除了为 IHC 从业者选择优化的 AR 条件/抗体组合提供指南外，涉及使用蛋白肽库和相关抗体的比较蛋白质组学和验证研究将进一步增强定量 IHC 测量的再现性和灵敏度。由于 IHC 在临床诊断/预后和转化癌症研究中的广泛应用，对定量 IHC 的需求不断升级。此外，使用来自高质量样本的富集和精选细胞进行靶向蛋白质组学研究的最大期望在于在临床环境中以及临床前测试和临床试验期间鉴定诊断、预后和预测生物标志物，以及发现和验证生物制药行业的新蛋白质靶标。 FDA 于 2006 年 3 月发布的《关键路径机会报告》不仅是关键路径计划的第一个具体蓝图，旨在通过应用新的策略和技术来简化药物审批流程，而且还强调生物标志物开发是关键路径计划的关键路径之一。 “改善医疗产品开发的最重要领域。” 公共卫生相关性声明：通过将 Calibrant 独特的组织蛋白质组能力与南加州大学 Clive R. Taylor 博士在抗原修复-免疫组织化学方面的专业知识相结合，拟议的研究不仅旨在通过创建来评估和优化定量免疫组织化学测量蛋白质型肽库，但也侧重于进一步开发和演示一种新的生物标志物发现范式，以实现对存档的福尔马林固定和支持癌症研究、诊断和治疗的石蜡包埋组织集合。