Quantitative high-throughput screening (qHTS) evaluates the pharmacology of drug and investigational agent libraries for potential therapeutic uses, toxicological risk assessment, and increasingly for academic chemical tool discovery. Phenotypic HTS assays aim to interrogate molecular pathways and networks, often relying on cell culture systems, historically with less emphasis on multicellular organisms. C. elegans has served as a powerful eukaryotic model organism for human biology and disease by virtue of genetic conservation and experimental tractability. Here we describe a paradigm to enable C. elegans in qHTS using 384-well microtiter plate laser scanning cytometry. GFP-expressing organisms are used to reveal phenotype-modifying structure-activity relationships to guide subsequent life stages and proteomic analysis. E. coli bacterial ghosts, a non-replicating nutrient source, allow compound exposures over 7-days spanning two life cycles to mitigate complications from bacterial overgrowth. We demonstrate the method with a library composed of anti-infective agents, or molecules of general toxicological concern. Each was tested in 7-point titration to assess the feasibility of nematode-based in vivo qHTS, and examples of follow-up strategies were provided to study organism-based chemotype selectivity and subsequent network perturbations having a physiological impact. We anticipate a broader application of this qHTS-coupled proteomics approach will enable the analysis of C. elegans orthologous transgenic phenotypes of human pathologies to facilitate drug and probe profiling from high-impact chemical libraries for a range of therapeutic indications and study of potential toxicological signatures.

中文翻译：

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用于药物发现和比较毒理学的体内定量高通量筛选

定量高通量筛选 (qHTS) 评估药物和研究试剂库的药理学，以用于潜在的治疗用途、毒理学风险评估，并越来越多地用于学术化学工具的发现。表型 HTS 测定旨在询问分子途径和网络，通常依赖于细胞培养系统，历史上不太重视多细胞生物。C. elegans凭借遗传保守性和实验易处理性，已成为人类生物学和疾病的强大真核模型生物。在这里，我们描述了一种使C. elegans成为可能的范例在 qHTS 中使用 384 孔微量滴定板激光扫描细胞仪。表达 GFP 的生物体用于揭示改变表型的结构-活性关系，以指导随后的生命阶段和蛋白质组学分析。大肠杆菌细菌幽灵是一种非复制性营养源，允许在跨越两个生命周期的 7 天内接触化合物，以减轻细菌过度生长引起的并发症。我们使用由抗感染剂或一般毒理学关注的分子组成的库来演示该方法。每个都在 7 点滴定中进行测试，以评估基于线虫的体内可行性qHTS 和后续策略示例用于研究基于生物体的化学型选择性和随后的具有生理影响的网络扰动。我们预计这种 qHTS 耦合蛋白质组学方法的更广泛应用将能够分析秀丽隐杆线虫的人类病理学直系同源转基因表型，以促进从高影响化学库中进行药物和探针分析，以用于一系列治疗适应症和潜在毒理学特征的研究.