Toxicology in the 21st Century Program (Tox21) - Systems Toxicology

21 世纪毒理学计划 (Tox21) - 系统毒理学

• 批准号: 10261234
• 负责人: Menghang Xia
• 金额: $ 29.55万
• 依托单位: NATIONAL CENTER FOR ADVANCING TRANSLATIONAL SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10261234
• 关键词: 3-Dimensional; Acetylcholine; Ache; Advocate; Anabolism; Animal Testing; Biochemical Pathway; Bioenergetics; Biological; Biological Assay; Biomedical Engineering; Biotechnology; CREB1 gene; CYP2B6 gene; CYP2C9 gene; CYP2D6 gene; CYP3A4 gene; Caenorhabditis elegans; Cardiac Myocytes; Cell Line; Cell model; Cell physiology; Cells; Cellular Assay; Chemical Warfare Agents; Chemicals; Cholinesterases; Collaborations; Collection; Consumption; Cosmetics; Cytidine Diphosphate Diglycerides; Development; Drug toxicity; Elements; End Point Assay; Epithelial; Epithelium; European Union; Evaluation; Exposure to; Genetic; Genomics; Gluconeogenesis; Goals; Guidelines; Health; Human; Immune response; In Vitro; Laws; Lead; Leadership; Libraries; Liver Microsomes; Malignant Neoplasms; Manuscripts; Mass Spectrum Analysis; Measures; Membrane Potentials; Metabolism; Methods; Mission; Mitochondria; Modeling; National Institute of Environmental Health Sciences; National Toxicology Program; Nematoda; Neurotransmitters; Nuclear Receptors; Organelles; Oxygen Consumption; Pathway interactions; Pattern; Pesticides; Pharmaceutical Preparations; Phase; Phosphatidylglycerols; Phytochemical; Pilot Projects; Preparation; Proteins; Proteomics; Publications; Publishing; Research Personnel; Safety; Signal Transduction; Skin; Specificity; Structure; System; TP53 gene; Technology; Testing; Thick; Tight Junctions; Time; Tissues; Topical application; Toxicology; Translational Research; Triglycerides; United States Environmental Protection Agency; United States Food and Drug Administration; United States National Institutes of Health; Validation; Work; base; bioinformatics tool; biological systems; bioprinting; computational toxicology; consumer product; cytokine; design; drug metabolism; environmental chemical; environmental toxicology; follow-up; frontier; high throughput screening; in vivo; inhibitor/antagonist; irritation; lipid biosynthesis; lipid metabolism; mitochondrial dysfunction; mitochondrial membrane; neurotoxicity; parkin gene/protein; predictive modeling; programs; protein expression; response; robotic system; screening; three dimensional cell culture; toxicant; two-dimensional

The Tox21 programs federal partners include the Environmental Protection Agency (EPA), the Food and Drug Administration (FDA) and NIH, with leadership from NCATS and the National Toxicology Program (NTP) at the National Institute of Environmental Health Sciences (NIEHS). These agencies work together to advance in vitro toxicological testing. The Tox21 Program is comprised of three NCATS teams: Systems Toxicology, Genomic Toxicology, and Computational Toxicology. The Systems Toxicology team has identified, developed, optimized, and/or screened more than 10 assays. Highlights range from performing 6 online screenings, including MSTI/electrophile assay, p53 assay with or without liver microsomes, CREB, CYP2C9 and CYP2D6 against the Tox21 10K compound library on the Tox21 robotic system. Mitochondria are essential cellular organelles that participate in important cellular processes, including bioenergetics, metabolism, and signaling. As part of the Tox21 effort in the phase II of U.S. Tox21 program, the Systems Toxicology team has identified a group of mitochondria toxicants from previous screen against the Tox21 10K compound collection using a panel of assays including mitochondrial membrane potential (MMP), ROS formation, p53, Nrf2/ARE, mitochondrial oxygen consumption, cellular Parkin translocation, and larval development and ATP status in the nematode C. elegans. To further study the mechanism of compound action, we have performed a global proteomic profiling of several lesser-known mitochondria toxicants identified from our previous study (Xia et al., 2018) in human AC16 human cardiomyocytes. After expose to these mitochondria toxicants, the expression level of a group of proteins has been significantly changed in several lipid metabolism related pathways including CDP-diacylglycerol, triacylglycerol and phosphatidylglycerol biosynthesis using Mass spectrometry (MS)-based omics technology and bioinformatics tools. These protein expression changes involved in metabolism and redirection of energy usage were also related to mitochondria dysfunction. This pilot study will help to advance drug/toxicity target validation in translational sciences such as summarized a general pattern of protein changes indicating mitochondrial dysfunction. The manuscript describing this study has been accepted for publication in Frontiers in Genetics. Assessing irritation and sensitization potential is a key element in the safety evaluation of topical drugs and other consumer products such as cosmetics. The use of advanced cellular models as alternatives to animal testing for both products and ingredients in consumer products is already mandated by law in the European Union (EU). To evaluate the compounds for their irritation and sensitization potential, we tested about 500 topically applied compounds by using two-dimensional (2D) and three-dimensional (3D) culture of skin cells as an alternative method. The assay endpoints in reconstructed human epithelial (RhE) and full-thickness skin (FTS) include viability; TEER, a measure of the tight junctions found in skin; and cytokine secretions to assess irritation and sensitization of topical compounds. This study represents the first steps in advocating bio-engineered skin models to replace current animal tests. The findings from this study have been published in Frontiers of Bioengineering and Biotechnology. To profile compounds for their sensitization potential, we have used KeratinoSens assay suggested by the OECD test guideline to screen the Tox21 10K compound library in a qHTS platform. After the primary screening, we identified a group of actives and will further test them for their sensitization potential using a panel of the follow-up assays including use of the 3D-bioprinted tissues. AChE is the primary cholinesterase in the body that metabolizes a key neurotransmitter, acetylcholine. Inhibition of AChE activity can lead to neurotoxicity and known inhibitors include organophosphorus pesticides, chemical warfare agents, drugs, and various phytochemicals. In collaboration with the CFSAN/FDA, NTP and EPA, the Systems Toxicology team has identified a group of AChE inhibitors from the primary screening and performed a group of follow-up assays to further study the mechanism of compound action. The manuscript summarized the experimental findings has been submitted for the publication. PXR is an important nuclear receptor that regulates drug metabolism; it has also recently been shown to have an impact on gluconeogenesis, cancer, lipogenesis, and the immune response. To profile the compounds that activate PXR, we have screened the hPXR-luc cell line. Four structural clusters were identified to highly activate PXR, while 20 compounds were selected for further evaluation based on potency, efficacy, structural clustering, and novelty. These chosen compounds were then treated in HepaRG cells to analyze the induction of CYP3A4 and CYP2B6. Eleven compounds significantly induced CYP3A4 and were analyzed for their PXR activity. These compounds were further tested in a HepaRG-PXR-KO cell line to confirm the involvement of PXR. The manuscript is currently under preparation.

Tox21 计划的联邦合作伙伴包括环境保护局 (EPA)、食品和药物管理局 (FDA) 和 NIH，并由 NCATS 和国家环境健康科学研究所 (NIEHS) 的国家毒理学计划 (NTP) 领导。这些机构共同努力推进体外毒理学测试。 Tox21 计划由三个 NCATS 团队组成：系统毒理学、基因组毒理学和计算毒理学。 系统毒理学团队已鉴定、开发、优化和/或筛选了 10 多种检测方法。亮点包括在 Tox21 机器人系统上针对 Tox21 10K 化合物库进行 6 项在线筛选，包括 MSTI/亲电子试剂测定、使用或不使用肝微粒体的 p53 测定、CREB、CYP2C9 和 CYP2D6。 线粒体是重要的细胞器，参与重要的细胞过程，包括生物能、代谢和信号传导。作为美国 Tox21 计划第二阶段 Tox21 工作的一部分，系统毒理学团队使用一系列测定（包括线粒体膜电位 (MMP)、ROS 形成），从之前针对 Tox21 10K 化合物集合的筛选中鉴定出了一组线粒体毒物、p53、Nrf2/ARE、线粒体耗氧量、细胞 Parkin 易位以及线虫线虫中的幼虫发育和 ATP 状态。为了进一步研究复合作用机制，我们对人类 AC16 人心肌细胞中从我们之前的研究 (Xia et al., 2018) 中鉴定出的几种鲜为人知的线粒体毒物进行了全局蛋白质组学分析。暴露于这些线粒体毒物后，使用基于质谱（MS）的组学技术和生物信息学工具，一些脂质代谢相关途径中的一组蛋白质的表达水平发生了显着变化，包括CDP-二酰基甘油、三酰基甘油和磷脂酰甘油生物合成。这些参与新陈代谢和能量使用重定向的蛋白质表达变化也与线粒体功能障碍有关。这项试点研究将有助于推进转化科学中的药物/毒性目标验证，例如总结表明线粒体功能障碍的蛋白质变化的一般模式。描述这项研究的手稿已被《遗传学前沿》接受发表。 评估刺激和致敏潜力是外用药物和化妆品等其他消费品安全性评估的关键要素。欧盟 (EU) 法律已强制要求使用先进的细胞模型来替代动物测试的产品和消费品成分。为了评估这些化合物的刺激性和致敏潜力，我们使用皮肤细胞的二维 (2D) 和三维 (3D) 培养作为替代方法，测试了约 500 种局部应用的化合物。重建人上皮 (RhE) 和全层皮肤 (FTS) 的检测终点包括活力； TEER，皮肤紧密连接的衡量标准；和细胞因子分泌物以评估局部化合物的刺激和致敏作用。这项研究代表了倡导生物工程皮肤模型取代当前动物测试的第一步。这项研究的结果发表在《生物工程和生物技术前沿》上。为了分析化合物的致敏潜力，我们使用 OECD 测试指南建议的 KeratinoSens 测定法在 qHTS 平台中筛选 Tox21 10K 化合物库。初步筛选后，我们确定了一组活性物质，并将使用一系列后续测定（包括使用 3D 生物打印组织）进一步测试它们的致敏潜力。 AChE 是体内主要的胆碱酯酶，负责代谢关键的神经递质乙酰胆碱。抑制 AChE 活性可导致神经毒性，已知的抑制剂包括有机磷农药、化学战剂、药物和各种植物化学物质。系统毒理学团队与 CFSAN/FDA、NTP 和 EPA 合作，从初步筛选中鉴定出一组 AChE 抑制剂，并进行了一组后续测定，以进一步研究复合作用机制。该手稿总结了实验结果，已提交出版。 PXR是调节药物代谢的重要核受体；最近还被证明对糖异生、癌症、脂肪生成和免疫反应有影响。为了分析激活 PXR 的化合物，我们筛选了 hPXR-luc 细胞系。确定了 4 个结构簇可高度激活 PXR，同时选择 20 种化合物根据效力、功效、结构簇和新颖性进行进一步评估。然后将这些选定的化合物在 HepaRG 细胞中进行处理，以分析 CYP3A4 和 CYP2B6 的诱导作用。十一种化合物显着诱导 CYP3A4，并分析了它们的 PXR 活性。这些化合物在 HepaRG-PXR-KO 细胞系中进行了进一步测试，以确认 PXR 的参与。目前手稿正在准备中。