Abstract
Cytochrome P450 (CYP) induction is often considered a liability in drug development. Using calibration curve-based approaches, we assessed the induction parameters R3, relative induction score (RIS), Cmax/EC50 and AUC/F2, derived from concentration-response curves of CYP3A4 mRNA and enzyme activity data in vitro, as predictors of CYP3A4 induction potential in vivo. Plated cryopreserved human hepatocytes from three donors were treated with 20 test compounds, including several clinical inducers and non-inducers of CYP3A4. After the two day treatment, CYP3A4 mRNA levels and testosterone 6β-hydroxylase activity were determined by RT-PCR and LC-MS/MS analysis, respectively. Our results demonstrated a strong and predictive relationship between the extent of midazolam AUC change in human and the various parameters calculated from both CYP3A4 mRNA and enzyme activity. The relationships exhibited with non-midazolam in vivo probes, in aggregate, were unsatisfactory. In general, the models yielded better fits when unbound rather than total plasma Cmax was used to calculate the induction parameters, as evidenced by higher R2 and lower RMSE and GMFE. With midazolam, the R3 cut-off value of 0.9, as suggested by FDA guidance, effectively categorized strong inducers, but was less effective in classifying mid-range or weak inducers. This study supports the use of calibration curves generated from in vitro mRNA induction response curves to predict CYP3A4 induction potential in human. With the caveat that most compounds evaluated here were not strong inhibitors of enzyme activity, testosterone 6β-hydroxylase activity was also demonstrated to be a strong predictor of CYP3A4 induction potential in this assay model.
- The American Society for Pharmacology and Experimental Therapeutics