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Application of Targeted Functional Assays to Assess a Putative Vascular Disruption Developmental Toxicity Pathway Informed By ToxCast High-Throughput Screening Data
Citation:
ELLIS-HUTCHINGS, R. G., R. S. SETTIVARI, A. A. MCCOY, N. KLEINSTREUER, V. A. MARSHALL, T. B. KNUDSEN, AND E. W. CARNEY. Application of Targeted Functional Assays to Assess a Putative Vascular Disruption Developmental Toxicity Pathway Informed By ToxCast High-Throughput Screening Data. Presented at The Society of Toxicology (SOT) 51st Annual Meeting and ToxExpo 2012, Moscone Convention Center, San Francisco, CA, March 11 - 15, 2012.
Impact/Purpose:
Chemical perturbation of vascular development is a putative toxicity pathway which may result in developmental toxicity. EPA’s high-throughput screening (HTS) ToxCast program contains assays which measure cellular signals and biological processes critical for blood vessel development. By testing the Phase-1ToxCast chemicals in these assays, and comparing the results to prenatal DT study summary information derived from ToxRefDB, a vascular disruption signature was identified.
Description:
Chemical perturbation of vascular development is a putative toxicity pathway which may result in developmental toxicity. EPA’s high-throughput screening (HTS) ToxCast program contains assays which measure cellular signals and biological processes critical for blood vessel development. By testing the Phase-1ToxCast chemicals in these assays, and comparing the results to prenatal DT study summary information derived from ToxRefDB, a vascular disruption signature was identified. This signature was correctly observed when the antiangiogenic thalidomide analogue, 5HPP-33, was tested in a ToxCast assay subset. There is utility in using targeted in vitro functional assays to explore the potential consequences of chemicals that test positive in the ToxCast program, both for chemicals without DT data (5HPP-33) or as an intermediate tier for DT data comparisons. Therefore, 5HPP-33 was tested in rat whole embryo culture (WEC) and in vitro rat aortic explant (AE) cultures. Mid-somite stage rat embryos were cultured in media containing 0, 1.6, 5, 15, 30 or 46 uM 5HPP-33 for 48 h followed by evaluation for developmental defects. To further confirm the direct effects of 5HPP-33 on angiogenesis, rat AE were cultured in media containing 0, 0.46, 4.6, 46, 93, or 247 uM 5HPP-33 for four days and the resulting inhibition of microvessel outgrowth was evaluated. In WEC, 5HPP-33 caused developmental defects and embryolethality at ≥ 15 uM. Consistent with an antiangiogenic mode of action in embryos, 5HPP-33 inhibited microvessel outgrowth in cultured AEs at ≥ 0.46 uM and completely abolished vessel outgrowth at 46 uM with a cell morphology similar to the outcome of cell-agent based in silico models informed by ToxCast data. Data from these targeted functional assays correlated with the in vitro HTS assay data for this vascular disrupting compound. This abstract does not necessarily reflect US EPA policy.