Application of a Novel Nanotechnology for Molecular Profiling of Tumor Cellular E

描述(申请人提供)：我们对1)肿瘤遗传和细胞异质性，2)最近对癌症干细胞、内皮祖细胞和肌上皮细胞等的描述，以及3)即使在组织学上相似的肿瘤，对治疗的反应也不同；提出了关于不同肿瘤细胞亚群对肿瘤生物学行为的相对贡献的基本问题。然而，允许对离散的肿瘤细胞成分进行前瞻性表征并从肿瘤中回收选定的存活细胞的技术尚未开发出来。我们开发了一种新的纳米技术，由一系列微制造的SU8聚合物元件组成，允许分离和回收单个贴壁细胞。这种先进的纳米技术与多色免疫荧光和先进的共聚焦显微镜相结合，使我们能够提出将该技术应用于同时识别、恢复和评估代表单个肿瘤内各种细胞成分的可存活的原代贴壁细胞群体中选定的分子图谱。这些研究的假设是，托盘阵列纳米技术将允许识别、计数和恢复以下单个细胞肿瘤元素：癌症干细胞、内皮祖细胞、肌上皮细胞、上皮细胞和炎性细胞，导致单个肿瘤内这些细胞亚群的分子特征，并将通过追求以下特定目标进行测试：目标1.优化托盘阵列，以最大限度地捕获、检测和恢复肿瘤细胞元素。目的#2将托盘阵列应用于原发性乳腺肿瘤的细针抽吸(FNA)标本，从离散的肿瘤细胞成分中鉴定和分离单个细胞。目的#3建立回收单个稀有细胞的分子分析方法，如单细胞RT-PCR。这些研究代表了生物医学工程、先进激光光学、细胞生物学、免疫学和临床肿瘤学的融合，并将推动未来的研究，以解决基本的生物学和临床问题。

DESCRIPTION (provided by applicant): Our increasing appreciation of 1) tumor genetic and cellular heterogeneity, 2) the recent descriptions of cancer stem, endothelial progenitor, & myoepithelial cells, among others, and 3) disparate responses to treatment even for histologically similar tumors; raise fundamental questions as to the relative contributions of various tumor cellular subsets to the biologic behavior of a tumor. However, technologies permitting the prospective characterization of discrete tumor cellular elements and recovery of selected viable cells from a tumor have yet to be developed. We have developed a novel nanotechnology consisting of an array of microfabricated SU8 polymer elements that permits the isolation and recovery of individual adherent cells. This advanced nanotechnology combined with multicolor immunofluorescence and advanced confocal microscopy enables us to propose application of this technology to the simultaneous identification, recovery, and evaluation of selected molecular profiles from viable primary adherent cell populations representing the various cellular elements within individual tumors. The HYPOTHESIS for these studies is that the pallet array nanotechnology will permit identification, enumeration, and recovery of the following individual cellular tumor elements: cancer stem, endothelial progenitor, myoepithelial, epithelial, and inflammatory cells leading to the molecular characterization of these cellular subsets within individual tumors and will be tested by pursuing the following Specific Aims: AIM #1. Refine the pallet array for maximum cell capture, detection, and recovery of tumor cellular elements. AIM #2 Apply pallet array to Fine Needle Aspirate (FNA) samples of primary breast tumors to identify and isolate individual cells from discrete tumor cellular elements. AIM #3 Establish feasibility of molecular analysis of recovered individual rare cells, e.g. single cell RT-PCR. These studies represent a convergence of biomedical engineering, advanced laser optics, cell biology, immunology, and clinical oncology and will drive future studies to address fundamental biological and clinical questions.