In Silico Comparison of Rifampicin and 25-desacetyl Rifampicin-Induced PXR-Mediated CYP450 Transcriptional Response...

The pregnane X receptor (PXR) regulates the expression of cytochrome P450 (CYP) enzymes and plays a crucial role in the metabolism of various drugs. Rifampicin (RIF) is a PXR ligand that forms the primary metabolite, 25-desacetyl rifampicin (25-DRIF), which retains the antimicrobial activity of the original drug. In this study, we quantified PXR activation and its associated effects on CYP3A4, CYP2C9, and CYP2B6 enzymes in response to 25-DRIF treatment by combining mathematical modeling with long-term mRNA expression analysis of these enzymes in 3D primary human hepatocyte (3D PHH) spheroids. Our estimates suggest that 25-DRIF activates PXR at a rate 20 times lower than RIF. The PXR-dependent rate constant for CYP3A4 transcription was estimated to be higher in 3D PHHs treated with 25-DRIF than in those treated with RIF and also higher than that for CYP2B6 transcription in 3D PHHs treated with 25-DRIF. The rate constants driving PXR-dependent transcription of CYP2C9 were comparable in RIF- and 25-DRIF-treated 3D PHHs. These results demonstrate the ligand-specific nature of PXR activation and suggest that the transcription of PXR-controlled CYP enzymes is ligand- and CYP-specific in 3D PHHs. Finally, we showed that the half-maximal effective concentration ( ${\text{EC}}_{50}$ ) evaluated from our mathematical predictions was time-dependent, which was further validated by CYP3A4 gene reporter assays that measured RIF-induced PXR activity.