Kleinstreuer Nicole C, Ceger Patricia, Watt Eric D, Martin Matthew, Houck Keith, Browne Patience, Thomas Russell S, Casey Warren M, Dix David J, Allen David, Sakamuru Srilatha, Xia Menghang, Huang Ruili, Judson Richard
NIH/NIEHS/DNTP/The NTP Interagency Center for the Evaluation of Alternative Toxicological Methods , Research Triangle Park, North Carolina 27713, United States.
Integrated Laboratory Systems, Inc. , Research Triangle Park, North Carolina 27560, United States.
Chem Res Toxicol. 2017 Apr 17;30(4):946-964. doi: 10.1021/acs.chemrestox.6b00347. Epub 2016 Dec 9.
Testing thousands of chemicals to identify potential androgen receptor (AR) agonists or antagonists would cost millions of dollars and take decades to complete using current validated methods. High-throughput in vitro screening (HTS) and computational toxicology approaches can more rapidly and inexpensively identify potential androgen-active chemicals. We integrated 11 HTS ToxCast/Tox21 in vitro assays into a computational network model to distinguish true AR pathway activity from technology-specific assay interference. The in vitro HTS assays probed perturbations of the AR pathway at multiple points (receptor binding, coregulator recruitment, gene transcription, and protein production) and multiple cell types. Confirmatory in vitro antagonist assay data and cytotoxicity information were used as additional flags for potential nonspecific activity. Validating such alternative testing strategies requires high-quality reference data. We compiled 158 putative androgen-active and -inactive chemicals from a combination of international test method validation efforts and semiautomated systematic literature reviews. Detailed in vitro assay information and results were compiled into a single database using a standardized ontology. Reference chemical concentrations that activated or inhibited AR pathway activity were identified to establish a range of potencies with reproducible reference chemical results. Comparison with existing Tier 1 AR binding data from the U.S. EPA Endocrine Disruptor Screening Program revealed that the model identified binders at relevant test concentrations (<100 μM) and was more sensitive to antagonist activity. The AR pathway model based on the ToxCast/Tox21 assays had balanced accuracies of 95.2% for agonist (n = 29) and 97.5% for antagonist (n = 28) reference chemicals. Out of 1855 chemicals screened in the AR pathway model, 220 chemicals demonstrated AR agonist or antagonist activity and an additional 174 chemicals were predicted to have potential weak AR pathway activity.
使用当前经过验证的方法来测试数千种化学物质以鉴定潜在的雄激素受体（AR）激动剂或拮抗剂，将耗资数百万美元且需要数十年才能完成。高通量体外筛选（HTS）和计算毒理学方法能够更快速且低成本地鉴定潜在的雄激素活性化学物质。我们将11种HTS ToxCast/Tox21体外试验整合到一个计算网络模型中，以区分真正的AR途径活性与技术特异性试验干扰。体外HTS试验在多个点（受体结合、共调节因子募集、基因转录和蛋白质产生）以及多种细胞类型中探究AR途径的扰动。确证性体外拮抗剂试验数据和细胞毒性信息被用作潜在非特异性活性的额外标志。验证此类替代测试策略需要高质量的参考数据。我们通过国际测试方法验证工作和半自动系统文献综述相结合的方式，汇编了158种假定的雄激素活性和非活性化学物质。详细的体外试验信息和结果使用标准化本体被汇编到一个单一数据库中。确定激活或抑制AR途径活性的参考化学物质浓度，以建立具有可重复参考化学物质结果的一系列效价。与来自美国环保署内分泌干扰物筛选计划的现有一级AR结合数据进行比较，结果表明该模型在相关测试浓度（<100μM）下鉴定出了结合剂，并且对拮抗剂活性更敏感。基于ToxCast/Tox21试验的AR途径模型对激动剂（n = 29）参考化学物质的平衡准确率为95.2%，对拮抗剂（n = 28）参考化学物质的平衡准确率为97.5%。在AR途径模型中筛选的1855种化学物质中，有220种化学物质表现出AR激动剂或拮抗剂活性，另外有174种化学物质预计具有潜在的弱AR途径活性。
