Comparative study of CYP450 gene regulation in human pluripotent stem cell-derived liver 2D cell and 3D organoid...

Human pluripotent stem cell (hPSC)-derived hepatic models, including 2D hepatocyte-like cells (2D HLCs) and hepatic organoids (HOs), are valuable in vitro models for evaluating the safety and efficacy of drugs. However, 2D HLCs show limited expression and activity of drug-metabolizing enzymes, particularly cytochrome P450 (CYP450), which are involved in detoxification, a major liver function. HOs have more mature properties than 2D HLCs, particularly enhanced CYP450 gene expression. However, the transcriptional regulatory mechanisms that correlate with CYP450 expression in HOs remain unclear. Epigenetic mechanisms, including DNA methylation and histone modification, are essential for controlling gene expression during stem cell differentiation. Here, we identified epigenetic states around transcriptional regulatory regions and compared them with those in primary human hepatocytes. We found that significantly higher CYP450 gene expression in hPSC-derived HOs than in 2D HLCs was strongly associated with decreased DNA methylation and increased enrichment of histone H3 lysine 27 acetylation in their transcriptional regulatory regions. Furthermore, because of the higher expression of nuclear receptor genes, especially constitutive androstane receptor and pregnane X receptor, HOs showed higher NR-mediated induction of CYP3A4, UGT1A1, and MDR1 than 2D HLCs. Therefore, these results suggest that mature epigenetic regulation may have an impact on drug metabolism and toxicity outcomes in hPSC-derived hepatic models and, hence, be used as an indicator of model maturation.