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: 7503795
• 负责人: Satya Prakash Saxena
• 金额: $ 74.84万
• 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/7503795
• 关键词: Actins; Analysis of Variance; Antibodies; Antigens; Archives; Area; Back; Biliary; Bioinformatics; Biological; Biological Assay; Biological Markers; Biological Products; Biopsy; Blood Cells; Blood capillaries; Breast; Breast Carcinoma; Buffers; California; Carcinoma; Cataloging; Catalogs; Cells; Clinical; Clinical Trials; Collagen; Condition; Control Groups; Coupling; Critical Pathways; DNA; Databases; Detection; Development; Development, Other; Diagnosis; Diagnostic; Digestion; Disease; Disease Outcome; Drug Approval Processes; Duct (organ) structure; Ductal Epithelium; E-Cadherin; Electrospray Ionization; Epithelial Cells; Epitopes; Evaluation; Facility Construction Funding Category; 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; Neuro-Oncological Ventral Antigen 2; Noninfiltrating Intraductal Carcinoma; Normal tissue morphology; Numbers; Outcome; PCNA gene; Paraffin; Paraffin Embedding; Patients; Peptide Library; Peptides; Phase; Phase I Clinical Trials; Ploidies; Population; Population Control; Preclinical Testing; Process; Progress Reports; Proteins; Proteome; Proteomics; Public Health; RNA; Range; Reaction; Recording of previous events; Reporting; Reproducibility; Research; Resources; Retrieval; Running; Sampling; Smooth Muscle; Solutions; Specimen; Staining method; Stains; Standards of Weights and Measures; Stromal Cells; Structure; System; TP53 gene; Techniques; Technology; Therapeutic; Tissue Banks; Tissue Fixation; Tissues; Trypsin; Two-Dimensional Polyacrylamide Gel Electrophoresis; United States Food and Drug Administration; 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; prognostic; protein expression; 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（由申请方提供）：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历史悠久，因此全球有大量福尔马林固定和石蜡包埋（FFPE）存档组织库。这些FFPE组织收集，随附临床和结果信息，为进行回顾性蛋白质生物标志物研究提供了宝贵的资源。除了由当前蛋白质组技术（包括二维聚丙烯酰胺凝胶电泳和多维液相色谱系统）施加的样品量限制之外，缺乏用于从FFPE组织中检索蛋白质的优化方法进一步限制了对存档组织进行分子分析的能力。通过与Shan-Rong Shi和克莱夫R.在R41 I期研究期间，抗原修复（AR）与Gemini蛋白质组学技术的结合不仅完成了对从FFPE组织中提取的蛋白质的质量和再现性的严格评估，以优化AR方法，而且还证明了在使用存档的FFPE组织收集来寻求生物标志物发现方面的重要机会。在R42 II期项目期间，Calibrant和USC团队之间的协同努力旨在在模型和肿瘤FFPE组织中生成蛋白质型肽库。这些蛋白质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随后的免疫组织化学（IHC）抗体开发提供可用表位数据库的第一步。除了为IHC从业者提供选择最佳AR条件/抗体组合的指南外，涉及使用蛋白质型肽文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测量的重现性和灵敏度。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的需求持续升级。此外，使用从高质量标本中富集和选择的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在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，以及在生物制药行业中发现和验证新的蛋白质靶点。FDA于2006年3月发布的《关键路径机会报告》不仅是关键路径倡议的第一个具体蓝图，该倡议旨在通过应用新的战略和技术来简化药物批准过程，而且还强调生物标志物开发是改善医疗产品开发的最重要领域之一。" 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克莱夫R.泰勒在南加州大学的抗原检索-免疫组织化学，拟议的研究不仅旨在通过建立蛋白质型肽库来评估和优化定量免疫组织化学测量，而且还专注于进一步开发和展示一种新的生物标志物发现范例，用于对存档的福尔马林固定和石蜡固定的蛋白质组进行全面和比较性的蛋白质组学分析，支持癌症研究、诊断和治疗的嵌入式组织收集。