Organs-on-Chips as Bridges for Predictive Toxicology M. Shane Hutson, 1–3 Peter G. Alexander, 1,4 Vanessa Allwardt, 1,2 David M. Aronoff, 1,5,6 Kaylon L. Bruner-Tran, 1,7 David E. Cliffel, 1,2,8 Jeffrey M. Davidson, 1,6,9 Albert Gough, 1,10 Dmitry A. Markov, 1,2,11,12 Lisa J. McCawley, 1,2,11,12 Jennifer R. McKenzie, 1,8 John A. McLean, 1,2,8 Kevin G. Osteen, 1,6,7,9 Virginia Pensabene, 1,2,11 Philip C. Samson, 1–3 Nina K. Senutovitch, 1,10,13 Stacy D. Sherrod, 1,8 Matthew S. Shotwell, 1,2,14 D. Lansing Taylor, 1,10,13 Lauren M. Tetz, 1,5 Rocky S. Tuan, 1,4,15–18 Lawrence A. Vernetti, 1,10 and John P. Wikswo 1–3,11,19 Abstract The next generation of chemical toxicity testing will use organs-on-chips (OoCs)—3D cultures of heterotypic cells with appropriate extracellular matrices to better approximate the in vivo cellular microenvironment. Researchers are already working to validate whether OoCs are predictive of toxicity in humans. Here, we review two other key aspects of how OoCs may advance predictive toxicology—each taking advantage of OoCs as sys- tems of intermediate complexity that remain experimentally accessible. First, the intermediate complexity of OoCs will help elucidate the scale(s) of organismal complexity that currently confound computational predic- tions of in vivo toxicity from in vitro data sets. Identifying the strongest confounding factors will help researchers improve the computational models underlying such predictions. Second, the experimental accessibility of OoCs will allow researchers to analyze chemical-exposure responses in OoCs using an array of high-content read- outs—from fluorescent biosensors that report dynamic changes in specific cell signaling pathways to unbiased searches over broader biochemical space using technologies like ion mobility-mass spectrometry. Such high- content information on OoC responses will help determine the details of adverse outcome pathways. We note these possible uses of OoCs so that researchers and engineers can consider them in the design of next-generation OoC control, perfusion, and analysis platforms. Key words: computational model, environmental, fetal membrane, limb development, liver, mammary gland, organotypic culture model, toxicant. R egulatory agencies face a daunting task in deter- mining whether any of the tens of thousands of chemi- cals in common use pose a risk to human health and the environment. The prevailing environmental toxicology para- digm of the past 40 years has relied on a gold standard— guideline animal studies—that are expensive, pose troubling ethics questions because of the intentional exposure of large numbers of animals, and are difficult to relate directly to human health hazards because of interspecies discordance and testing at concentrations well above human exposure levels. Based on recommendations of the National Research Council, 1 several U.S. agencies (EPA, FDA, and NIH through NCATS and NIEHS) have pushed forward on new in vitro testing paradigms—for example, ToxCast and 1 Vanderbilt-Pittsburgh Resource for Organotypic Models for Predictive Toxicology, Vanderbilt University, Nashville, Tennessee, and University of Pittsburgh, Pittsburgh, Pennsylvania. 2 Vanderbilt Institute for Integrative Biosystems Research & Education; 3 Department of Physics & Astronomy; 8 Department of Chemistry; 11 Department of Biomedical Engineering; and 14 Department of Biostatistics; Vanderbilt University, Nashville, Tennessee. 4 Department of Orthopaedic Surgery, 15 Department of Bioengineering, 16 Center for Cellular and Molecular Engineering, 17 Center for Military Medicine Research, and 18 McGowan Institute for Regenerative Medicine, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania. 5 Division of Infectious Diseases, Department of Internal Medicine; 6 Department of Pathology, Microbiology and Immunology; 7 Department of Obstetrics & Gynecology; 12 Department of Cancer Biology; and 19 Department of Molecular Physiology & Biophysics; Vanderbilt University Medical Center, Nashville, Tennessee. 9 Research Service, Department of Veterans Affairs Tennessee Valley Healthcare System, Nashville, Tennessee. 10 Department of Computational & Systems Biology and 13 University of Pittsburgh Drug Discovery Institute, University of Pittsburgh, Pittsburgh, Pennsylvania. APPLIED IN VITRO TOXICOLOGY Volume 2, Number 2, 2016 ª Mary Ann Liebert, Inc. DOI: 10.1089/aivt.2016.0003 97