Implementation of phosphoprotein preservation technology for cancer biospecimens

癌症生物样本磷蛋白保存技术的实现

• 批准号: 8311650
• 负责人: Lance Allen Liotta
• 金额: $ 30.95万
• 依托单位: GEORGE MASON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-15 至 2014-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8311650
• 关键词: Address; Adoption; Advanced Development; Antigens; Architecture; Archives; Biological Markers; Biological Preservation; Biopsy Specimen; Blinded; Breast; Buffers; Cancer Histology; Cancer Patient; Cell Count; Cell membrane; Cells; Chemistry; Clinical; Clinical Research; Collagen Type IV; Collection; Colon; Community Hospitals; Cytokeratin; DNA; Diagnosis; Diagnostic; ERBB2 gene; Endometrium; Epidermal Growth Factor Receptor; Epitopes; Equipment; Estrogen Receptors; Female; Fixatives; Formalin; Freezing; Funding; Generations; Goals; Guidelines; Health Care Costs; Hematoxylin and Eosin Staining Method; Histocompatibility Testing; Histology; Human; Immunohistochemistry; Informed Consent; International; Ischemia; Knowledge; Liver parenchyma; Malignant Neoplasms; Measurement; Measures; Molecular; Molecular Profiling; Molecular Target; Morphology; Mucous Membrane; Mus; Nuclear; One-Step dentin bonding system; Operative Surgical Procedures; Organ; Paraffin; Paraffin Embedding; Pathologist; Pathway interactions; Phase; Phosphoproteins; Phosphotransferases; Process; Progesterone Receptors; Prostate; Protein Kinase; Protein Microchips; Proteins; RNA; Research; Shipping; Ships; Side; Signal Pathway; Specimen; Staining method; Stains; Structure of parenchyma of lung; Technology; Testing; Therapeutic; Time; Tissue Preservation; Tissue Procurements; Tissues; United States National Institutes of Health; Validation; cost; diagnostic accuracy; inhibitor/antagonist; innovation; kinase inhibitor; laser capture microdissection; lymph nodes; male; novel; sample fixation; tissue fixing; tissue processing; tumor

DESCRIPTION (provided by applicant): An urgent clinical goal is to identify molecular networks associated with subpopulations of cancer patients who may respond individually to molecular targeted inhibitors. Current molecular targeted therapeutics is directed at protein kinases and/or their phosphorylated substrates. Therefore, measurement of this new class of phosphoprotein signal pathway epitopes in tumor biopsy samples is crucial for individualizing molecular targeted therapies. Phosphoprotein antigen epitopes are not adequately preserved by formalin fixation and paraffin embedding, and freezing of tissue is very expensive and compromises diagnostic accuracy. We propose the advanced development and clinical validation of an innovative and transformative technology for preserving tissue phosphoproteins and diagnostic histomorphology for clinical cancer molecular profiling. Applying knowledge gained under an NIH R21 funded study, we created a novel tissue preservation chemistry that stabilizes all classes of phosphoproteins, is compatible with paraffin embedding, while maintaining complete diagnostic histomorphology, and fully preserving critical diagnostic immunohistology (IHC) antigens including Estrogen Receptor, Progesterone Receptor, HER2, and Ki-67. These IHC antigens are not preserved by special research fixatives used for tissue RNA preservation. Our new non-formalin tissue preservative, termed Biomarker and Histology Preservative (BHP) can be seamlessly introduced into the current community hospital clinical diagnostic workflow with no additional steps or equipment. At the time of procurement, tissue can be immersed directly in the new fixative and processed into a paraffin block for routine diagnosis, obviating the need for costly freezing during shipping or storage. BHP offers the potential for substantial improvements over conventional formalin fixation. In the present application we propose the blinded clinical validation of our novel preservation chemistry in community hospital settings, utilizing a team of international pathologists for validation. The goal of the project is one-step paraffin block stabilization of all classes of cellular phosphoproteins, diagnostic histomorphology, and diagnostic immunohistochemistry antigens, while at the same time maintaining full diagnostic morphology equivalent or superior to standard formalin fixation. We will collect fresh surgical tissue, under informed consent, covering a broad variety of organs and cancer histology to develop an archive of 150 cases of matched paraffin and frozen specimens. We will measure 100 validated phosphoprotein epitopes spanning membrane, cytoplasmic and nuclear compartments from extracted paraffin sections using Reverse Phase Protein Microarray (RPMA) and Laser Capture Microdissection (LCM) technology. Following objective independent validation by diagnostic pathologists, this transformative technology will be ready for widespread clinical and research use. Adoption of the technology would mean that only one diagnostic paraffin block could be used for all classes of molecular profiling rather than the current requirement for multiple blocks. This would increase diagnostic accuracy while substantially reducing costs.

描述（由申请人提供）：一个迫切的临床目标是确定与可能对分子靶向抑制剂有反应的癌症患者亚群相关的分子网络。目前的分子靶向治疗是针对蛋白激酶和/或其磷酸化底物。因此，在肿瘤活检样本中测量这类新的磷酸化蛋白信号通路表位对于个体化分子靶向治疗至关重要。磷酸蛋白抗原表位不能通过福尔马林固定和石蜡包埋得到充分保存，组织冷冻非常昂贵，而且会影响诊断的准确性。我们建议先进的开发和临床验证的创新和变革性技术保存组织磷酸化蛋白和诊断组织形态学为临床癌症分子谱。应用在NIH R21资助研究下获得的知识，我们创造了一种新的组织保存化学，稳定所有类别的磷蛋白，与石蜡包埋兼容，同时保持完整的诊断组织形态学，并充分保存关键的诊断免疫组织学（IHC）抗原，包括雌激素受体，孕激素受体，HER2和Ki-67。这些免疫组化抗原不被用于组织RNA保存的特殊研究固定剂保存。我们的新型非福尔马林组织防腐剂，称为生物标志物和组织学防腐剂（BHP），可以无缝地引入到当前的社区医院临床诊断工作流程中，不需要额外的步骤或设备。在采购时，组织可以直接浸泡在新的固定液中，并加工成石蜡块进行常规诊断，避免了运输或储存期间昂贵的冷冻。与传统的福尔马林固定相比，必和必拓提供了大幅改进的潜力。在目前的应用中，我们建议在社区医院环境中对我们的新型保存化学进行盲法临床验证，利用国际病理学家团队进行验证。该项目的目标是一步石蜡块稳定所有类别的细胞磷酸化蛋白、诊断组织形态学和诊断免疫组织化学抗原，同时保持完全诊断形态学等同于或优于标准福尔马林固定。我们将在知情同意的情况下收集新鲜的手术组织，涵盖各种器官和癌症组织学，以建立150例匹配的石蜡和冷冻标本的档案。我们将使用反相蛋白微阵列（RPMA）和激光捕获显微解剖（LCM）技术，从提取的石蜡切片中测量100个经过验证的跨越膜、细胞质和核区室的磷酸化蛋白表位。经过诊断病理学家的客观独立验证后，这种变革性技术将为广泛的临床和研究应用做好准备。采用该技术意味着只需一个诊断石蜡块就可以用于所有类型的分子分析，而不是目前需要多个块。这将提高诊断的准确性，同时大大降低成本。