proximity assay)-based binding assay is prone to false positive results, while the
PXR reporter gene assay only represents the PXR-dependent induction
pathway, possibly ignoring other pathways of CYP3A4 induction that existin
vivo. Primary cultures of human hepatocytes provide an ideal screening system
in many respects. However, these assays are expensive and time consuming.
Availability, quality, and variability of donor variations also present potential
problems. Finally, induction may be masked by concomitant inhibition of
constitutive and induced CYP3A4 by the inducing agent. This ‘‘metabolic
masking effect’’ may generate false negative results (Luo et al., 2002, 2003,
2004). In spite of these potential drawbacks,in vitroand cell culture models
currently provide the best compromise between time- and cost-efficient
screening methods, and accurate prediction of induction potential in clinical
situations.
17.2.2.1 Primary Cultures of Human Hepatocytes Measurement of CYP3A4
induction in primary human hepatocyte cultures involves three steps: isolation
of hepatocytes from autopsy- or biopsy-derived human donor livers; treatment
with a test compound for three days following a 2-day recuperation culture
period; and measurement of the expression or activity of CYP3A4 (Fig. 17.1).
FIGURE 17.1 Isolation and culture of primary human hepatocytes for CYP induction
studies.
ASSESSMENTS 553