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

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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.