Proteomic Predictors of Clinical Outcome of Targeted Therapies in Prostate Cancer

缺乏生物标志物来评估哪些患者可能对途径靶向治疗有反应，这造成了 对应用集成技术进行发现和翻译验证的深刻需求 这样的生物标志物。蛋白质组学方法和计算生物信息学处理的最新进展 使整合蛋白质组学技术应用于发现生物标志物成为可能。如果体外使用 纳米传感器装置和体内纳米颗粒成像方法， 癌症治疗开发和使用的工具。我们建议通过整合两个 定向和两种全面的蛋白质组学技术：a）细胞内信号传导蛋白芯片， 定量已知的细胞内信号传导蛋白的定向分析方法; B）活细胞捕获 传感器阵列，其代表用于细胞表面和分泌物的定向分析的纳米传感器方法， 蛋白质组; c）基于生物素捕获的细胞表面分析方法， 鉴定和定量细胞表面蛋白质丰度的方法; d）固相萃取 糖蛋白（SPEG）谱，这是一种用于研究分泌蛋白的综合分析方法 还有血生物标志物的发现最初将是疾病-（前列腺癌）和途径-（人类表皮 生长因子受体（Her）-激酶轴）的重点，并将提供一个基础，我们的CCNE-TR中心将使用 生产、评估和验证纳米传感器和基于纳米颗粒的成像。我们将首先定义细胞表面 雄激素非依赖性前列腺癌的分泌蛋白质组，并确定这些蛋白质中的蛋白质 和细胞内蛋白质组，它们指示对Her激酶轴的扰动。我们将分析 这些亚蛋白质组在雄激素非依赖性前列腺癌的原代培养模型中， Her激酶靶向治疗剂2C 4（结合Her-2上的表位的人源化单克隆抗体 阻止配体介导的Her-2异二聚化）和吉非替尼（一种小分子抑制剂， 表皮生长因子受体上的ATP结合位点）。我们还将用配体刺激 靶向该轴的每个受体二聚体伴侣。轴反应信息蛋白将是 通过与基因数据库的比较，评估其Her-kinase和前列腺癌特异性， 研究者组可获得的不同组织来源的其他细胞系中的蛋白质表达。接下来我们将 将我们的体外结果与人类异种移植模型的蛋白质表达模式的信息相结合， 鉴定具有预测和/或监测反应效用的一组标记物。血的银行， 由UCLA前列腺孢子开发的用于该项目的活组织样本代表了不同的前体细胞。 和治疗后时间点以及雄激素非依赖性异种移植模型的多样性集合 其特征在于对Her激酶靶向治疗剂的敏感性。最后，我们将验证 在特定目的1和2中发现的生物标志物，用于离体纳米传感器装置和体内纳米传感器装置 通过产生蛋白候选物的亲和试剂进行纳米颗粒成像;我们还将验证小组的 通过使用前列腺的孢子组织和血清样品指导人类治疗干预的能力 用Her-kinase定向疗法治疗的癌症患者。

The lack of biomarkers for assessment of which patients may respond to pathway-targeted therapy creates a profound need for the application of integrated technologies for the discovery and translational validation of such biomarkers. Recent advances in proteomic methods and computational bioinformatics processing have enabled application of integrated proteomic technologies to the discovery of biomarkers. If used by ex vivo nano-sensor devices and in vivo nanoparticle imaging methods such biomarkers may provide effective new tools to cancer therapy development and use. We propose to discover candidate markers by integrating two directed and two comprehensive proteomic technologies: a) intracellular-signaling protein chips consisting of a directed analysis method which quantitates known, intracellular signaling proteins; b) living-cell capture sensor arrays which represent a nanosensor approach for directed analysis of the cell-surface and secretory proteomes; c) biotin-capture-based cell-surface profiling methods consisting of a comprehensive analysis method that identifies and quantitates the abundance of cell-surface proteins; d) solid-phase extraction of glycoprotein (SPEG) profiling, which is a comprehensive analysis method for the study of secreted proteins and blood. Biomarker discovery will initially be disease- (prostate cancer) and pathway- (human epidermal growth factor receptor (Her)-kinase axis) focused and will provide a foundation our CCNE-TR Center will use to produce, evaluate and validate nanosensors and nanoparticle-based imaging. We will first define the cell-surface and secretory proteomes of androgen-independent prostate cancer and identify proteins within these and the intracellular proteome that are indicative of the perturbations to the Her-kinase axis. We will analyze these sub-proteomes in primary culture models of androgen-independent prostate cancer by treatment with the Her-kinase targeted therapeutics, 2C4 (a humanized monoclonal antibody that binds epitopes on Her-2 that prevent ligand-mediated Her-2 heterodimerization) and geftinib (a small molecule inhibitor that competes for the ATP binding site on epidermal growth factor receptor). We will additionally stimulate with ligands targeting each of the receptor-dimer partners of this axis. Axis-response-informative proteins will be evaluated for their Her-kinase and prostate cancer specificity, by comparison with a database of gene and protein expression in other cell lines of different tissue origin available to the investigator group. Next we will integrate our in vitro results with information of the protein expression patterns of human xenograft models to identify a panel of markers with utility for predicting and/or for monitoring response. The bank of blood and viable tissue samples developed by the UCLA Prostate SPORE for use in this project represent different pre and post-treatment time points and a diverse collection of androgen-independent xenograft models characterized for sensitivity to the Her-kinase-targeted therapeutics. Lastly we will validate the utility of the biomarkers discovered in Specific Aims 1 and 2 for use with ex vivo nanosensor devices and in vivo nanoparticle imaging by generating affinity reagents for protein candidates; We will also validate the panel's ability to guiding human therapeutic intervention by using SPORE tissue and serum samples of prostate cancer patients treated with Her-kinase-directed therapies.