Project 1: Identification and Validation of Panel of Early Cell Surface Gene Targets

摘要-这项研究属于肿瘤异质性基因组学及其对肿瘤的影响。 向EAC进展，作为RFA-CA-16-006中定义的可能区域之一。我们之前已经 利用基因表达谱鉴定EGFR和ErbB2作为有希望的细胞表面靶点 用于EAC成像的频率基因扩增，并开发出针对这些EAC的特异性多肽 目标。我们现在的目标是确定BE向EAC发展过程中出现的早期目标。我们希望找到更好的 变异性和较低的表达水平对于较早的目标和较晚的目标，从而准备了更多 严谨的方法。我们将使用Affymetrix阵列和RNAseq分析来识别早期基因靶点 在细胞表面过度表达，可以显影用于成像。我们将利用这些复杂的基因组 提供全面分析基因改变、突变和DNA拷贝数变异的工具 人类食道样本，以确定一组有希望的候选者。我们将严格验证 应用定量RT-PCR和免疫组织化学技术研究日本血吸虫组织芯片的候选靶点 一个独立的样本队列。这种方法需要大量唯一的HGD 可在我们的大型组织生物库中获得n=780个人的食道标本，包括n=560个EAC， 从头收集(没有新辅助化疗或放射治疗)。对于每个经过验证的早期目标，我们将 转染端粒酶永生化的BE细胞，使其在细胞表面过表达以用于发育 项目2中的单体和二聚体多肽。 这些目标的成功完成将导致一组早期基因的识别和验证 在BE向EAC发展过程中出现的靶点，将在项目2中用于开发新的多肽成像 探员们。

Abstract – This research project falls under genomics of tumor heterogeneity and their impact on the progression toward EAC as one of the possible areas defined in RFA-CA-16-006. We have previously identified EGFR and ErbB2 using gene expression profiles as promising cell surface targets based on high- frequency gene amplification for imaging of EAC, and have developed peptides that are specific for these targets. We now aim to identify early targets that arise in progression of BE to EAC. We expect to find greater variability and lower levels of expression for early compared with late targets, thus have prepared a more rigorous approach. We will use Affymetrix arrays and RNAseq analysis to identify early gene targets that are overexpressed on the cell surface and can be developed for imaging. We will use these sophisticated genomic tools to provide a comprehensive analysis of gene alterations, mutations, and DNA copy number variations in human esophagus specimens to identify a panel of promising candidates. We will rigorously validate candidate targets using quantitative RT-PCR and immunohistochemistry on tissue microarrays prepared from an independent cohort of specimens. This approach requires a large number of HGD that are uniquely available in our large tissue biorepository of n = 780 human esophageal specimens, including n = 560 EAC, collected de novo (no neo-adjuvant chemo or radiation therapy). For each early target validated, we will transfect telomerase-immortalized BE cells to overexpress the target on the cell surface for use in developing the monomer and dimer peptides in Project 2. Successful completion of these aims will result in identification and validation of a panel of early gene targets that arise in progression of BE to EAC that will be used in Project 2 to develop novel peptide imaging agents.