High-Throughput Spheroid Screening Platform

高通量球体筛选平台

• 批准号: 10013252
• 负责人: James P Freyer
• 金额: $ 49.97万
• 依托单位: BENNUBIO INC
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-09 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10013252
• 关键词: 3-Dimensional; Achievement; Acoustics; Address; Algorithms; Apoptosis; Back; Biological Assay; Biological Models; Caliber; Cell Communication; Cells; Cellular Spheroids; Coculture Techniques; Complex; Computer software; Coupled; Data Analyses; Detection; Development; Drug Screening; Ensure; Excision; Exposure to; Flow Cytometry; Fluorescence; Health; Human; Image; Incubated; Lasers; Liquid substance; Location; Measurement; Measures; Methods; Microspheres; Modeling; Necrosis Induction; Optics; Penetration; Performance; Pharmaceutical Preparations; Pharmacologic Substance; Research Personnel; Resolution; Running; Sampling; Scanning; Series; Signal Transduction; Stream; Stress; Structure; Surface; Suspension Culture; Suspensions; System; Technology; Tissue Model; Tissues; Tumor Tissue; Variant; analytical method; base; cell type; commercialization; data acquisition; design; high throughput screening; improved; innovation; instrument; nephelometry; novel; particle; response; screening; software systems; tool; tumor; uptake

PROJECT SUMMARY This proposal will develop the High-Throughput Spheroid Screening Platform (HTSSP) to create an entirely new tool to study cell-cell interactions and microenvironmental stresses that occur in tissues and tumors. Such interactions are critical in the pharmaceutical response of cells within tissues, but current methods do not allow high throughput screening (HTS) of cellular responses in a tissue-like microenvironment. The HTSSP is being developed to enable the use of the powerful multicellular spheroid 3D tissue/tumor model system, which has many potential applications in 3D drug screening, in true high-throughput screening (HTS) applications. To create the HTSSP, we are addressing critical limitations in large particle analysis via flow cytometry (Aim 1), continuous sample handling of large particles such as spheroids (Aim 2), improving analytical methods to obtain spatial/structural information from spheroids in analysis (Aim 3), and demonstrating key applications of the HTSSP in real world assays (Aim 4). To improve flow cytometry analysis, we will optimize our current high throughput parallel flow cytometer (the 10xFC) by modifying the flow cells and the optics for ideal use with particles between 100 and 1000 microns in diameter. This flow cytometer will be integrated with an innovative suspension culture and sampling fluidic system to create the hardware of the HTSSP. These fluidics will maintain multiple samples of spheroids in stirred suspension chambers during incubation or exposure to drugs, then use the same chambers for sampling and resampling for analysis by our flow cytometer. The final synergistic technical achievement will be the development of improved data acquisition systems that enable the rapid extraction of 3D location information from probes within a spheroid. When combined with the hardware, this will synergistically complete the HTSSP, which will be used for key high-throughput pharmaceutical screening applications, including: apoptosis/necrosis induction, fraction of each cell type in a co-culture, fluorescent drug uptake. Taken as a whole, the HTSSP will represent a significant technological advance in pharmaceutical screening as it will provide the first truly high throughput approach to the study of pharmaceutical interactions in a 3D tissue model.

项目概要 该提案将开发高通量球体筛选平台（HTSSP），以创建一个完全 研究细胞间相互作用以及组织和肿瘤中发生的微环境应激的新工具。这样的 相互作用对于组织内细胞的药物反应至关重要，但目前的方法不允许 组织样微环境中细胞反应的高通量筛选（HTS）。 HTSSP 正在 开发用于使用强大的多细胞球体 3D 组织/肿瘤模型系统，该系统具有 3D 药物筛选和真正的高通量筛选 (HTS) 应用中的许多潜在应用。到 创建 HTSSP，我们正在通过流式细胞术解决大颗粒分析的关键限制（目标 1）， 对球体等大颗粒进行连续样品处理（目标 2），改进分析方法以获得 分析中来自球体的空间/结构信息（目标 3），并演示该方法的关键应用 现实世界检测中的 HTSSP（目标 4）。为了改进流式细胞仪分析，我们将优化我们目前的高 通过修改流通池和光学器件以实现理想的使用吞吐量并行流式细胞仪（10xFC） 直径在 100 至 1000 微米之间的颗粒。该流式细胞仪将与创新的 悬浮培养和采样流体系统，用于创建 HTSSP 的硬件。这些流体将维持 在孵育或接触药物期间将多个球体样品放入搅拌的悬浮液室中，然后使用 用于采样和重新采样的相同室，以便通过我们的流式细胞仪进行分析。最后的协同作用 技术成就将是开发改进的数据采集系统，使快速 从球体内的探针提取 3D 位置信息。当与硬件结合时，这将 协同完成HTSSP，用于关键高通量药物筛选 应用，包括：细胞凋亡/坏死诱导、共培养中每种细胞类型的分数、荧光药物 吸收。总体而言，HTSSP 将代表制药领域的重大技术进步 筛选，因为它将为研究药物相互作用提供第一个真正高通量的方法 3D 组织模型。