Omics-Based Assays for Screening Chemical Toxicity Using Stem Cells

项目摘要/摘要 化学毒性的测试和建模发生了范式转变。目前的毒理学要求人类细胞-- 基于(与基于动物)、基于证据和基于Omics的分析。间充质干细胞(MSC)的作用 在各种组织的维护和修复中起重要作用，包括骨、软骨和肌肉。MSCs 也在循环血液中发现，并参与伤口愈合过程。因此，MSC可以说是 化学毒理学中生理上最相关的成体干细胞之一。回应RFA-ES-13- 003，Neomeics Inc.建议开发基于Omics的高通量检测方法，以评估 化学毒物对人骨髓间充质干细胞增殖分化的影响。核心技术平台将是 基于Neomeics的突破性硅微流控芯片，多喷口发射极阵列芯片(MEA芯片)， 这使得纳米流液相色谱-电喷雾电离质谱仪(NanoLC-ESI/MS)- 基于多类别分析物(多肽、多肽、 蛋白质和代谢物)在OMICS水平上，从少量样品。该项目还将需要 我们在人类间充质干细胞的表征及其机制研究方面的丰富经验的优势 对遗传毒性应激的反应，如电离辐射和化学毒物，如过氧化氢。 在此第一阶段项目中，我们将演示基于OMICS和 利用人骨髓间充质干细胞进行高含量化学毒性筛选。在目标1中，我们将开发表型分析来 对基于OMICS的重点后续检测的条件进行优先排序。在目标2中，我们将表演高- MSC化学处理前后的蛋白质组学和代谢组学分析 用于定量测定的96孔或384孔板上每个孔的联合培养上清液 高通量筛选(QHTS)。我们将进行生物信息学分析并描绘出 对MSC的化学毒性负有责任的网络。如果成功，Neomeics的MEA芯片可能成为 使用干细胞的基于Omics的化学毒理学通用平台。

Project Summary/Abstract There is a paradigm shift in testing and modeling of chemical toxicity. Current toxicology calls for human-cell- based (vs. animal-based), evidence-based, and Omics-based assays. Mesenchymal stem cells (MSC) play important roles in the maintenance and repair of various tissues, including bone, cartilage, and muscle. MSCs are also found in circulating blood and participate in the wound-healing process. Therefore, MSC is arguably one of the most physiologically relevant adult stem cells for chemical toxicology. In response to RFA-ES-13- 003, Newomics Inc. proposes to develop high-throughput Omics-based assays to evaluate the effects of chemical toxicants on proliferation and differentiation of human MSC. The core technology platform will be based on Newomics' breakthrough silicon-microfluidic-chip, the multinozzle emitter array chip (MEA chip), which enables nanoflow liquid chromatography-electrospray ionization mass spectrometry (nanoLC-ESI/MS)- based, high-sensitivity, high-throughput, and multiplex measurements of multiclass analytes (peptides, proteins, and metabolites) at the Omics level, from small volumes of samples. The project will also take advantage of our extensive experience in the characterization of human MSC and mechanistic studies of their responses to genotoxic stresses such as ionizing radiation and chemical toxicants such as hydrogen peroxide. In this Phase I project, we will demonstrate a robust and universal two-stage workflow for Omics-based and high-content screening of chemical toxicity using human MSC. In Aim 1, we will develop phenotypic assays to prioritize the conditions for the focused follow-up Omics-based assays. In Aim 2, we will perform high- throughput proteomic and metabolomic analyses of MSC before and after chemical treatments, and the associated medium supernatants from each well on the 96-well or 384-well plates used for the quantitative high-throughput screening (qHTS). We will perform bioinformatics analysis and delineate the pathways and networks responsible for the chemical toxicity to MSC. If successful, Newomics' MEA chip may become a universal platform for Omics-based chemical toxicology using stem cells.