Blood-brain barrier (BBB) cell assays make it possible to investigate whether a compound is likely to compromise its functionality, thereby probably resulting in neurotoxicity and to predict CNS distribution. Recent advances in in vitro modeling of the blood-brain barrier have shown significant progress in replicating its complex physiological properties. Researchers have achieved more accurate mimicry of the blood-brain barrier structure and function from human stem cell source using advanced cell culture techniques, such as co-culturing multiple cell types, incorporating three dimensional cultures, and utilizing microfluidic systems. Several protocols to obtain human brain-like endothelial cells (BLECs) from induced pluripotent stem cells (iPSCs) have been reported. Within the framework of the European MSCA-ITN in3 project, we explored the possibility to use an iPSC-derived BBB model to assess the effects of repeated dose treatment with chemicals. Our first objective was to evaluate different published protocols to differentiate iPSCs into BBB like endothelial cells regarding their expression of endothelial markers, formation of tight barrier and the presence of functional efflux pumps. After some protocol optimizations, the iPSCs derived BBB cells were found to exhibit important BBB characteristics up to 15 days after the end of the differentiation and could be used to assess the effects of repeated dose treatment, using Cyclosporine A (CsA) as a model compound. Although iPSCs derived BBB cells were still undergoing transcriptional changes over time, a targeted transcriptome analysis (TempO-Seq) indicated a time and concentration dependent activation of ATF4, XBP1, Nrf2 and p53 stress response pathways under CsA treatment. Taken together, these results demonstrate that this iPSC-derived BBB model and iPSC-derived models in general hold great potential to study the effects of repeated dose exposure with chemicals allowing personalized and patient-specific studies in the future.