The antiretroviral agent efavirenz enhances the systemic clearance of coadministered drugs that are cytochrome P450 (CYP) 3A4 substrates. The mechanism of the apparent increase in CYP3A4 activity by efavirenz and the magnitude of change relative to other known inducers are not known. The authors tested the hypothesis that increased enzymatic activity by efavirenz entails CYP3A4 induction and activation of the human pregnane X receptor (hPXR), a key transcriptional regulator of CYP3A4. Employing primary cultures of human hepatocytes, they compared the CYP3A4 inductive effects of efavirenz (1-10 microM) to rifampin (10 microM) and phenobarbital (2 mM). A cell-based reporter assay was employed to assess hPXR activation. The authors observed that efavirenz caused a concentration-dependent CYP3A4 induction and hPXR activation. Based on the CYP3A4 activity assay, the average magnitude of induction by efavirenz (5-10 microM) was approximately 3- to 4-fold. In comparison, phenobarbital (2 mM) and rifampin (10 microM) caused a 5- and 6-fold induction, respectively.

ABSTRACT:Although many of the clinically significant drug interactions of the anti-human immunodeficiency virus (HIV) protease inhibitors (PIs) can be explained by their propensity to inactivate CYP3A enzymes, paradoxically these drugs cause (or lack) interactions with CYP3A substrates that cannot be explained by this mechanism (e.g., alprazolam). To better understand these paradoxical interactions (or lack thereof), we determined the cytochromes P450 and transporters induced by various concentrations (0-25 M) of two PIs, ritonavir and nelfinavir, and rifampin (positive control) in primary human hepatocytes. At 10 M, ritonavir and nelfinavir suppressed CYP3A4 activity but induced its transcripts and protein expression (19-and 12-and 12-and 6-fold, respectively; a >2-fold change over control was interpreted as induction). At 10 M, rifampin induced CYP3A4 transcripts, CYP3A protein, and activity by 23-, 12-, and 13-fold, respectively. The induction by rifampin of CYP3A activity was significantly correlated with its induction of CYP3A4 transcripts (r ‫؍‬ 0.96, p < 0.05) and CYP3A protein (r ‫؍‬ 0.89, p < 0.05). All three drugs (10 M) induced CYP2B6 activity by 2-to 4-fold, CYP2C8 and 2C9 activity by 2-to 4-fold and the transcripts of CYP2B6, 2C8, and 2C9 by >3-, 5-, and 3-fold, respectively. CYP2C19 and 1A2 activity and transcripts were modestly induced (2-fold), whereas, as expected, CYP2D6 was not induced by any of the drugs. Of the transporters studied, protease inhibitors moderately induced multidrug resistance 1 (ABCB1) and multidrug resistance-associated protein (ABCC1) transcripts but had no or minimal effect on the transcripts of breast cancer resistance protein (ABCG2), organic anion-transporting peptide (OATP) 1B1 (SLCO1B1), or OATP1B3 (SLCO1B3). On the basis of these data, we concluded that many of the paradoxical drug interactions (or lack thereof) with the PIs are metabolism-rather than transporter-based and are due to induction of CYP2B6 and 2C enzymes.

The Innovation and Quality Induction Working Group presents an assessment of best practice for data interpretation of in vitro induction, specifically, response thresholds, variability, application of controls, and translation to clinical risk assessment with focus on CYP3A4 mRNA. Single concentration control data and Emax/EC data for prototypical CYP3A4 inducers were compiled from many human hepatocyte donors in different laboratories. Clinical CYP3A induction and in vitro data were gathered for 51 compounds, 16 of which were proprietary. A large degree of variability was observed in both the clinical and in vitro induction responses; however, analysis confirmed in vitro data are able to predict clinical induction risk. Following extensive examination of this large data set, the following recommendations are proposed. a) Cytochrome P450 induction should continue to be evaluated in three separate human donors in vitro. b) In light of empirically divergent responses in rifampicin control and most test inducers, normalization of data to percent positive control appears to be of limited benefit. c) With concentration dependence, 2-fold induction is an acceptable threshold for positive identification of in vitro CYP3A4 mRNA induction. d) To reduce the risk of false positives, in the absence of a concentration-dependent response, induction ≥ 2-fold should be observed in more than one donor to classify a compound as an in vitro inducer. e) If qualifying a compound as negative for CYP3A4 mRNA induction, the magnitude of maximal rifampicin response in that donor should be ≥ 10-fold. f) Inclusion of a negative control adds no value beyond that of the vehicle control.