ANALYSIS OF DRUG RESISTANCE BY FLOW MICROFLUOROCYTOMETRY

流式微荧光细胞术耐药性分析

• 批准号: 3874461
• 负责人: J B TREPEL
• 金额: --
• 依托单位: DIVISION OF CANCER TREATMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3874461
• 关键词: doxorubicin; drug resistance; drug screening /evaluation; flow cytometry; gene expression; glycoproteins; human tissue; neoplastic cell; oncogenes; tissue /cell culture

Since the discovery of the importance of the mdr-1 gene product in multidrug resistance, most of the studies of the multidrug resistance phenotype have been performed at the nucleic acid level. We have established assays to examine drug resistance at the protein level by using flow cytometry to look at intact single cells. There are significant advantages to this approach, especially in probing the role of P-glycoprotein expression in innate and acquired multidrug resistance in clinical specimens. The assays we have developed allow us to measure P-glycoprotein expression and adriamycin content in each of thousands of cells, within hours of receiving a specimen. We have developed a two-parameter technique that allows us to correlate expression and adriamycin content in individual cells in a tumor specimen or cell line. Using this technique, we have been able to identify a subpopulation of innately resistant cells (high P-glycoprotein expression, low Adr content) in a CML-blast crisis cell line that was not detectable by previously available single parameter techniques. conversely, we were able to detect a drugsensitive subpopulation in a CMLblast crisis cell lines selected for Adr resistance and in MCF-7 cells transfected with the mdr-1 gene. In addition, we are working on flow cytometric analysis of other measures of drug resistance, including single cell detection of dihydrofolate reductase by fluoresceinated methotrexate and on the use of flow cytometry for the rapid screening of resistance reversal agents. We will use the techniques outlined to study drug resistance and its reversal in a variety of tumors. A project utilizing the unique multiparamter capabilities of flow analysis has been initiated to study the relationship of drug resistance to N-myc expression.

自从发现MDR-1基因产物在 多药耐药，多药耐药的研究最多 已经在核酸水平上进行了表型。我们有 建立了在蛋白质水平上检测耐药性的方法，方法是使用 流式细胞术来观察完整的单个细胞。有重要的 这种方法的优势，特别是在探索 P-糖蛋白在人卵巢癌先天和获得性多药耐药中的表达 临床标本。我们开发的分析方法使我们能够测量 P-糖蛋白表达和阿霉素含量在上千例 细胞，在收到样本后的几个小时内。我们已经开发出一种 双参数技术，允许我们将表达式和 肿瘤标本或细胞系中单个细胞中的阿霉素含量。 使用这项技术，我们已经能够识别出 天生耐药细胞(高P-糖蛋白表达，低ADR含量) 在先前不能检测到的CML-BLAST危机细胞系中 可用的单参数技术。相反，我们能够检测到一个 CML危机细胞系中药物敏感亚群的选择 MDR-1基因转导的MCF-7细胞对ADR耐药。 此外，我们正在进行其他措施的流式细胞仪分析。 抗药性，包括单细胞检测二氢叶酸 荧光甲氨蝶呤还原酶及其在流式细胞术中的应用 用于快速筛选抗药性逆转剂。我们将使用 概述了研究多种抗药性及其逆转的技术 肿瘤的症状。一个利用独特的多参数功能的项目 流动分析已经被用来研究药物之间的关系 对N-myc表达耐药。