Human Pluripotent Stem Cell Test for Assessing the Potential Teratogen Risk

In order to reduce the reliance on animal studies, both the pharmaceutical and chemical industries have been investigating new human cell-based in vitro assays capable of predicting a chemical’s potential harm to humans. Here we describe a human pluripotent stem cell test (hPST) capable of identifying compounds that pose teratogenic risk to humans such as thalidomide. Given the complexity of human development and the broad spectrum of compounds correctly classified based on their known in vivo teratogenic risk, the hPST is a remarkably simple assay. Following 3 days of directing the differentiation of a monolayer of human pluripotent stem cells toward the mesendoderm fate, the nuclear localization of a single transcription factor, SOX17, is quantitated. The reduction of SOX17 in the presence of test compounds is used to stratify the compounds’ teratogenic risk. However, as is the case with many human pluripotent stem cell experimental systems, careful optimization of the differentiation conditions is required in order to obtain reproducible results. Therefore, in addition to outlining the methodology of the hPST, we describe in detail various techniques we have used to optimize the assay.