Tens of thousands of chemicals are currently in commerce, and hundreds more are introduced every year. Because current chemical testing is resource intensive, only a small fraction of chemicals have been adequately evaluated for potential human health effects. New technologies and computational tools have shown promise for closing this knowledge gap. In the U.S. EPA’s ToxCast effort, the use of ~700 high-throughput in vitro assays has broadly characterized the biological activity and potential mechanisms of ~1,800 chemicals. Coupling the high-throughput in vitro assays with additional in vitro pharmacokinetic assays and in vitro-to-in vivo extrapolation modeling allows conversion of in vitro bioactive concentrations to estimates to an administered dose (mg/kg/day). High throughput exposure models are generating exposure estimates based on key aspects of chemical production, fate, transport, and personal use. The path for incorporating new approach methods and technologies for prioritization and assessment of chemical alternatives poses multiple scientific challenges. These challenges include sufficient coverage of toxicological mechanisms to meaningfully interpret negative test results, development of increasingly relevant test systems, computational modeling to integrate experimental data, characterizing uncertainty, and efficient validation of the test systems and computational models. The presentation will cover progress at the U.S. EPA in the development and application of these technologies and approaches in evaluating alternatives and systematically addressing each of these challenges. This abstract does not necessarily reflect U.S. EPA policy.