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A Novel High Throughput Tumor Spheroid Microtechnology

一种新型高通量肿瘤球体显微技术

基本信息

批准号：
8738627
负责人：
Gary D Luker
金额：
$ 24.57万
依托单位：
UNIVERSITY OF AKRON
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-20 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8738627
关键词：
Academia Address Adherent Culture Biological Assay Breast Cancer Cell Cancer Biology Cell Aggregation Cell Culture System Cell Density Cell Survival Cells Cellular Spheroids Clinical Complex Consumption Culture Media Development Dextrans Diffusion Drops Drug Compounding Drug Formulations Drug Targeting Ensure Environment Equipment Exhibits Gene Expression Generations Goals Growth Image Immersion Investigative Technique Industry Kinetics Laboratories Liquid substance Mainstreaming Maintenance Malignant Neoplasms Malignant Stromal Cell Metabolic Methods Modeling Molecular Molecular Profiling Morphology Nutrient Osmolalities Oxygen Pharmaceutical Preparations Phase Physiological Physiology Polyethylene Glycols Polymers Preclinical Drug Evaluation Process Reader Reagent Research Personnel Robotics Solid Neoplasm Stromal Cells Stromal Neoplasm Surface System Techniques Technology Testing Time Validation anticancer research aqueous cancer cell cancer therapy chemotherapy clinical efficacy cost effective cost effectiveness dextran drug development drug discovery drug testing evaporation high standard high throughput screening improved in vivo instrument nanolitre neoplastic cell novel physiologic model public health relevance response screening technology validation tumor tumor microenvironment two-dimensional

项目摘要

DESCRIPTION (provided by applicant): Two-dimensional (2D) cultures of cancer cells are routinely used in drug discovery for screening and initial characterization of the efficacy of librry of potential drug compounds. Despite their simplicity and compatibility with high throughput screening instruments, 2D cell assays often fail to predict the efficacy of compounds in vivo, making drug development and discovery an extremely costly process. Disparity between 2D cultures and the complex 3D environment of cancer cells in vivo is the major shortcoming of monolayer culture systems. Development of novel chemotherapeutics requires compound screening against malignant tumor cells in a setting that resembles the 3D tumor environment. Cancer cell spheroids (CCS) are 3D clusters of malignant cells that are regarded as physiologic models of solid tumors; they possess similar metabolic and proliferative gradients to avascular tumors and exhibit the clinical expression profiles of solid tumors. Despite the inherent power of CCS to predict clinical efficacy of drugs, incorporation of CCS into the mainstream drug development process is severely hindered by complex and expensive methodological requirements for the formation and maintenance of CCS. We overcome the limitations of existing techniques by developing a technological platform to generate spheroids of consistent size in standard 384-microwell plates using an aqueous two- phase system (ATPS). A drop of the denser aqueous phase containing cancer cells is robotically dispensed into each well containing the second, immersion phase. The drop confines cells and remains immiscible from the immersion phase to facilitate aggregation of cells into a compact CCS of well-defined size. Importantly the overlay of culture media provides nutrients and minimizes the well-known problem of evaporation and changes in osmolality of media as in other assays. The entire process of generating 384 spheroids is done robotically and in a single step. The unprecedented ease of formation and maintenance of CCS and full compatibility with standard equipment in high throughput screening laboratories makes this microtechnology readily available to the researchers in academia and industry. We anticipate that this microtechnology will make drug testing and screening with 3D tumor models a routine laboratory technique prior to expensive and tedious in vivo analyses. In addition, it will dramatically improve testing throughput and cost-effectiveness (increasing numbers of tested compounds and reduced reagent consumption) and efficiency (reducing hands-on time) to expedite drug discovery. We will accomplish our goals through two specific aims: (i) Generation of cancer cell spheroids with aqueous biphasic systems; (ii) Initial validation of ATPS spheroids for compound testing.

描述（由申请人提供）：癌细胞的二维（2D）培养通常用于药物发现，用于筛选和潜在药物化合物库功效的初步表征。尽管2D细胞分析简单且与高通量筛选仪器兼容，但它们往往无法预测化合物在体内的功效，这使得药物开发和发现成为一个极其昂贵的过程。二维培养与癌细胞在体内复杂的三维环境之间的差异是单层培养系统的主要缺点。新型化疗药物的开发需要在类似于3D肿瘤环境的环境中对恶性肿瘤细胞进行复合筛选。癌细胞球体（CCS）是恶性细胞的三维集群，被认为是实体瘤的生理模型；它们具有与无血管肿瘤相似的代谢和增殖梯度，并表现出实体肿瘤的临床表达谱。尽管CCS具有预测药物临床疗效的内在力量，但CCS形成和维持的复杂和昂贵的方法要求严重阻碍了将CCS纳入主流药物开发过程。我们通过开发一种技术平台，克服了现有技术的局限性，使用水两相系统（ATPS）在标准384微孔板上生成尺寸一致的球体。将含有癌细胞的较浓的水相滴入每一个含有第二浸入相的孔中。从浸泡阶段开始，液滴限制细胞并保持不混溶，以促进细胞聚集成尺寸明确的紧凑CCS。重要的是，培养基的覆盖层提供了营养，并最大限度地减少了众所周知的蒸发问题和其他检测中培养基渗透压的变化。产生384个球体的整个过程是机器人完成的，而且是一步完成的。CCS的形成和维护前所未有的容易，并且与高通量筛选实验室的标准设备完全兼容，使得学术界和工业界的研究人员随时可以使用这种微技术。我们预计，这种微技术将使药物测试和筛选与3D肿瘤模型的常规实验室技术之前昂贵和繁琐的体内分析。此外，它将显著提高测试吞吐量和成本效益（增加测试化合物的数量和减少试剂消耗）和效率（减少动手时间），以加快药物发现。我们将通过两个具体目标来实现我们的目标：(i)用水相双相系统产生癌细胞球体；（ii）用于化合物测试的ATPS球体的初步验证。