<i>In vitro-in vivo</i> extrapolation of intrinsic clearance based on data from human hepatocyte spheroids – a case study with the low-clearance drug xevinapant

Xevinapant, a drug previously under development for head and neck cancer, was investigated to determine its intrinsic clearance (CL_int) <i>in vitro</i> and assess the involvement of CYP enzymes in its metabolism. Extensive <i>in vitro</i> studies showed very low turnover, necessitating advanced methodologies to accurately measure CL_int values.Two novel in vitro assays, using a modified suspension hepatocyte protocol or 3D hepatocyte spheroid cultures, allowed measuring hepatic turnover of xevinapant with very low CL_int values of 1.3 or 0.3 µL/min/10⁶ cells, respectively. Reaction phenotyping using specific inhibitors, enabled by enhanced detection capabilities in spheroids, identified primarily CYP3A (> 50%) as contributor to the hepatic metabolism of xevinapant.<i>In vitro</i>-<i>in vivo</i> extrapolation, in combination with data from the human mass balance study, revealed that hepatic metabolism, primarily mediated by CYP3A, accounts for approximately 30% of the overall clearance. The remaining clearance is renal (∼20%) and metabolic, but potentially extrahepatic (∼50%).This research highlights the importance and benefits of utilising advanced <i>in vitro</i> techniques, such as human hepatocyte spheroids, to accurately characterise drug metabolism, providing valuable insights for pharmacokinetic evaluations and, ultimately, supporting the development of a PBPK model and understanding the DDI risk of xevinapant.