RAPID RARE CANCER CELL ISOLATION FOR MOLECULAR DIAGNOSIS

快速分离罕见癌细胞以进行分子诊断

• 批准号: 2862487
• 负责人: JEFFREY John CHALMERS
• 金额: $ 38.25万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-06 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2862487
• 关键词: cell line; circulating neoplastic cell; clinical research; diagnosis design /evaluation; flow cytometry; human subject; immunoconjugates; immunomagnetic separation; method development; molecular oncology; monoclonal antibody; neoplasm /cancer immunodiagnosis; rapid diagnosis

Certain tumors shed neoplastic cells into the blood before the primary tumor growth can be detected in the body. It has been shown recently that epithelial cells from the tumor can be detected in the blood at concentrations as low as 1 cell per ml of blood, and that the presence of rare cancer cells in blood has an important diagnostic value. The current screening methods are limited by relatively low speed of Fluorescent Activated Cell Scanning. (FACS),and complexity of preenrichment methods. The overall objective of this proposal is to develop a high-throughput, immunomagnetically based cell separating system to recover as many rare cancer cells in human blood as possible, for further molecular analysis (such as PCR, biological assays and other). The specific aims are as follows. First: the primary separation will be conduced using a novel continuos, flow-through immunomagnetic unit developed in our laboratories. Continuos units are intrinsically more efficient with respect to high throughput. We propose to develop experimental and theoretical basis for the next generation system which will sort cells at a rate of 10/7 cells/s, and allow a non- destructive screening of an entire volume of blood product used for cancer cell therapy ( typically 0.5 to 1.0 liters) in a short period of time (<1hour). Second: the continuos cell separation process can be staged, unlike the currently used batch systems. We propose to develop experimental and theoretical basis for the continuos staged separation process which will further increase the purity and decrease the volume of the cell product, and thus increase the rate of success of analysis downstream of the cell separation step. Third: the current cell labeling procedure may require modifications and optimization for the best performance in targeting and isolating rare cancer cells. We propose to screen available monoclonal antibodies and colloidal magnetic labels for the highest sensitivity and specificity in targeting rare cancer cells using unique Cell Tracking Velocimetry instrument developed in our laboratories. In summary, this proposal focuses on the "front-end" of cancer screening namely a high-throughoutput device to rapidly isolate and concentrate rare cancer cells form large numbers of cells. This proposal is responsive to PAR-98-067, Innovative Technologies for the Molecular analysis of Cancer, and we believe it addresses the second objective of the PAR, namely "novel technologies that will allow high- throughput analysis of genetic alterations, expression of genome products, and monitoring of signal transduction pathways to cancer."

某些肿瘤在体内检测到原发肿瘤生长之前就已将肿瘤细胞脱落到血液中。最近的研究表明，在血液中可以检测到来自肿瘤的上皮细胞，浓度低至每毫升血液1个细胞，并且血液中罕见癌细胞的存在具有重要的诊断价值。目前的筛选方法受限于相对较低的荧光活化细胞扫描速度。（FACS），以及预富集方法的复杂性。本提案的总体目标是开发一种高通量、基于免疫磁的细胞分离系统，以尽可能多地从人类血液中回收罕见的癌细胞，用于进一步的分子分析（如PCR、生物检测等）。具体目的如下。首先，初级分离将使用我们实验室开发的新型连续流免疫磁单元进行。Continuos单元在高吞吐量方面本质上更高效。我们建议为下一代系统开发实验和理论基础，该系统将以10/7个细胞/秒的速度分选细胞，并允许在短时间内（<1小时）对用于癌细胞治疗的整个体积的血液制品（通常为0.5至1.0升）进行非破坏性筛选。第二：连续细胞分离过程可以分阶段进行，不像目前使用的批处理系统。我们建议建立连续阶段分离工艺的实验和理论基础，进一步提高细胞产物的纯度，减少细胞产物的体积，从而提高细胞分离下游分析的成功率。第三，目前的细胞标记程序可能需要修改和优化，以获得最佳的靶向和分离罕见癌细胞的性能。我们建议使用我们实验室开发的独特的细胞跟踪测速仪筛选可用的单克隆抗体和胶体磁标记，以获得针对罕见癌细胞的最高灵敏度和特异性。综上所述，本提案关注的是癌症筛查的“前端”，即高通量输出装置，快速分离和浓缩罕见的癌细胞形成大量细胞。该提案是对PAR-98-067（癌症分子分析的创新技术）的响应，我们相信它解决了PAR的第二个目标，即“允许高通量分析基因改变的新技术，基因组产物的表达，以及监测癌症的信号转导途径。”