Breast Cancer Proteomic via Laser-Free Microdissection and Gemini Technologies

描述(由申请人提供)：生物相关蛋白质组学数据的生成需要由同质细胞群组成的样本，其中不同类型和/或发育阶段的不需要的细胞不会掩盖结果。对于组织活检的分析，问题更加复杂，因为通常存在许多不同的细胞类型，并且少量异常细胞可能位于未受影响的区域内或邻近区域。虽然激光捕获显微切割(LCM)等方法能够分离出同质的细胞亚群，但LCM获取的样本的蛋白质组学分析受到产生的样本量非常少的严重限制。为了避免已建立的蛋白质组技术在分析从显微解剖获取的组织标本中获得的蛋白质提取物方面的局限性，最近在CALIBRANT开发了一个有效的基于发现的蛋白质组平台。这个名为Gemini的蛋白质组平台将独特的多维分离系统与定制的后端生物信息学工具结合在一起，可以对从组织显微切割捕获的细胞中提取的微量蛋白质进行超灵敏分析。该项目还旨在采用我们的合作者、国家神经疾病和中风研究所(NINDS)的庄正平博士开创的一种新颖的、无激光的显微解剖技术，能够提供丰富、高质量和可重复的组织样本。通过将CALIBRANT从微小样本中进行蛋白质组分析的能力与庄博士(NINDS)在组织显微解剖和肿瘤病理学方面提供的技术和专业知识相结合，拟议中的研究代表着一项协同努力，旨在评估和验证一种新的生物标记物发现范式，使能够对癌细胞及其微环境进行蛋白质组分析，从而支持癌症研究、诊断和治疗。通过与耶鲁大学医学院的Fattaneh A.Tavassoli教授的合作，将应用由此产生的生物标记物发现平台在全球层面上研究与乳腺癌相关的分子机制，Tavassoli教授将应用30多年的乳腺癌病理学和生物学研究经验，并提供用于生物标记物发现的新鲜冷冻人类乳腺癌活检标本的集合。

DESCRIPTION (provided by applicant): The generation of biologically relevant proteomics data requires samples consisting of homogenous cell populations, in which no unwanted cells of different types and/or development stages obscure the results. The problem is compounded for the analysis of tissue biopsies, since many different cell types are typically present, and small numbers of abnormal cells may lie within or adjacent to unaffected areas. While methods such as laser capture microdissection (LCM) enable the isolation of homogeneous subpopulations of cells, proteomic analysis of LCM-procured specimens is severely constrained by the very low amounts of sample generated. To avoid the limitations of established proteome techniques for analyzing protein extracts obtained from microdissection-procured tissue specimens, an effective discovery-based proteome platform has recently been developed at Calibrant. This proteome platform, called Gemini, combines a unique multidimensional separation system with customized back-end bioinformatics tools, and allows ultrasensitive analysis of minute protein amounts extracted from cells captured by tissue microdissection. This project further aims to employ a novel, laser-free microdissection technique pioneered by our collaborator, Dr. Zhengping Zhuang at the National Institute of Neurological Disorders and Stroke (NINDS), capable of providing enriched, high quality, and reproducible tissue samples. By combining Calibrant's ability to perform proteomic profiling from minute samples with the technology and expertise offered by Dr. Zhuang (NINDS) in tissue microdissection and tumor pathology, the proposed research represents a synergistic effort toward the evaluation and validation of a novel biomarker discovery paradigm for enabling the proteome analysis of cancer cells and their micro-environment in support of cancer research, diagnosis, and treatment. Application of the resulting biomarker discovery platform for studying the molecular mechanisms associated with breast carcinoma at the global level will be realized through a collaboration with Professor Fattaneh A. Tavassoli (Yale University School of Medicine), who will apply more than 30 years of research experience in breast cancer pathology and biology and provide access to a collection of fresh frozen human breast cancer biopsies for biomarker discovery.