The development of a microscopy-based cell-array toxicity assay for quantifying C

用于定量 C 的基于显微镜的细胞阵列毒性测定的开发

• 批准号: 8625209
• 负责人: MATTHEW B FRANCIS
• 金额: $ 18.68万
• 依托单位: ADHEREN, INC.
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-23 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8625209
• 关键词: Acute Myelocytic Leukemia; Adhesions; Adoption; Annexins; Antibodies; Automation; Binding; Biological Assay; Biological Markers; Blood; Breast Cancer Cell; Cell Adhesion; Cell Death; Cell Line; Cell physiology; Cell surface; Cell-Matrix Junction; Cells; Colon Carcinoma; Complement; Complement-Dependent Cytotoxicity; DNA; DNA Microarray Chip; Data; Data Analyses; Development; Dose; Drug resistance; Effectiveness; Effector Cell; Evaluation; Fluorescence; Fluorescence Microscopy; Glass; Goals; Grant; HL60; Human; Image; Image Analysis; Imagery; In Vitro; Industry; Label; Life; Lymphoma; Measures; Methods; Microscopy; Morphology; Normal Cell; Patients; Pattern; Pharmaceutical Preparations; Pharmacologic Substance; Plastics; Polystyrenes; Population; Preclinical Drug Evaluation; Printing; Process; Production; Protocols documentation; Recovery; Research; Research Personnel; Resistance; Roche brand of trastuzumab; Screening for cancer; Solutions; Spottings; Surface; Techniques; Technology; Testing; Therapeutic antibodies; Time; Toxic effect; Whole Blood; antibody-dependent cell cytotoxicity; anticancer research; base; cancer cell; cancer therapy; cell growth; cell killing; cell type; cost; cytotoxicity; dosage; fluorescence imaging; fluorophore; improved; lithography; neoplastic cell; programs; public health relevance; radiochemical; scale up; screening; statistics; time use; tool; tumor

DESCRIPTION (provided by applicant): Live cell screening represents a major paradigm shift for the development of new pharmaceuticals and biologics. Despite the many automation and data analysis advances for this purpose, however, many challenges still hinder the implementation of this approach. As one set of examples, the most common cell cytotoxicity assays used to evaluate the efficacy of an antibody drug, the complement-dependent cytotoxicity (CDC) and antibody-dependent cellular cytotoxicity (ADCC) assays, only focus on end-point results. As a result, they yield little information about which cells are killed or resitant in a heterogeneous tumor population. In addition, none of the assay products offered today are compatible with the use of high content imaging to study the onset and rate of cytotoxicity, despite the rich quantity of data that this approach can provide. Adheren's new cell-array toxicity (CAT) assay platforms promise to advance this field by minimizing false positives, providing earlier cytotoxicity data, and allowing the visualization of cell death in real time. These producs are based on a core DNA-based cell attachment platform, which allows strong and programmable cell binding for a very wide range of cell types. The adhesion method allows both live and killed cells to be retained on the microarray, providing improved cytotoxicity statistics. This technology also allows the cell microarray to be washed without cell loss, and it enables the direct visualization of cell cytotoxicity over time using fluorescence imaging. This can providing unprecedented levels of real-time cell-specific data for the elucidation of complement- and effector cell-based cytotoxicity mechanisms. We propose to scale up and further develop these cell array-based ADCC and CDC microscopy assays by validating their effectiveness for a wide range of cell types and biological assays. In addition, the assay platform will be extended from currently the used glass bottom 96 well plates to include low-cost industry standard polystyrene 96 well plates.

描述(由申请人提供)：活细胞筛选代表了新药物和生物制品开发的重大范式转变。然而，尽管为此目的取得了许多自动化和数据分析方面的进展，但仍有许多挑战阻碍了这一方法的实施。作为一组例子，最常见的用于评价抗体药物疗效的细胞毒性试验，补体依赖性细胞毒性(CDC)和抗体依赖性细胞毒性(ADCC)试验，只关注终点结果。因此，他们几乎没有提供关于异质肿瘤群体中哪些细胞被杀死或耐药的信息。此外，目前提供的检测产品中没有一种与使用高含量成像来研究细胞毒性的开始和比率是兼容的，尽管这种方法可以提供丰富的数据。粘附素的新细胞阵列毒性 (CAT)分析平台承诺通过将假阳性降至最低，提供更早的细胞毒性数据，并允许实时显示细胞死亡，从而推动这一领域的发展。这些产品基于基于DNA的核心细胞附着平台，该平台允许对非常广泛的细胞类型进行强大的可编程细胞结合。粘附法允许活细胞和死亡细胞都保留在微阵列上，提供改进的细胞毒性统计数据。 这项技术还允许清洗细胞微阵列而不会丢失细胞，并能够使用荧光成像直接显示细胞随时间推移的细胞毒性。这可以为阐明补体和效应细胞的细胞毒性机制提供前所未有的实时细胞特异性数据。我们建议扩大和进一步发展这些基于细胞阵列的ADCC和CDC显微镜分析，通过验证它们对广泛的细胞类型和生物分析的有效性。此外，测试平台将从目前使用的玻璃底96孔板扩展到包括低成本的行业标准聚苯乙烯96孔板。