The Effect of Cyclophosphamide with and without Dexamethasone on Cytochrome P450 3A4 and 2B6 in Human Hepatocytes

Abstract

ABSTRACT:The purpose of this study was to characterize the concentrationresponse effects of cyclophosphamide (CPA) with and without dexamethasone (DEX; 10 M) on the expression of CYP3A4 and CYP2B6 in cultured human hepatocytes at concentrations representative of standard-and high-dose CPA therapy (25 to 750 M). CPA produced concentration-dependent increases in CYP3A4 and CYP2B6 activity and immunoreactive protein that peaked at 250 and 125 M, respectively, and declined thereafter. The inductive effect of CPA alone and in combination with DEX was greater in magnitude for CYP2B6 compared with CYP3A4. To further examine the inductive effect of CPA on CYP3A4, CPA (250 M) and DEX (10 M) alone and in combination were examined in 10 hepatocyte preparations. The combination of CPA and DEX yielded higher rates of 6␤-hydroxytestosterone formation than either agent alone. However, the effect was less than additive in human hepatocyte cultures with relatively high baseline CYP3A4 activity and additive or synergistic in human hepatocyte cultures with relatively low baseline CYP3A4 activity. Induction index was highly correlated with CYP3A4 baseline activity for both CPA (r 2 ‫؍‬ 0.75) and CPA plus DEX (r 2 ‫؍‬ 0.85). To investigate the potential mechanism for CPA-induced increases in CYP3A4 activity, the ability of CPA alone and in combination with DEX to activate pregnane X receptor (PXR) was explored using transient transfection assays. CPA produced a dose-dependent increase in PXR activation that was maximal at the highest CPA concentration studied (500 M). The addition of DEX to CPA resulted in a minor increase in PXR activation compared with CPA alone. These results indicate that CPA alone and in combination with DEX differentially induces the expression of CYP3A4 and 2B in a concentration-dependent manner, which may be mediated partially through activation of PXR. The impact of these effects on the efficacy and toxicity of CPA therapy warrants further investigation.

Nuclear receptor mediated induction of cytochrome P450 3A4 by anticancer drugs: a key role for the pregnane X receptor

“…Remarkably, ifosfamide had a greater capacity to induce CYP3A4 compared to cyclophosphamide, which is an isomer of ifosfamide. Lindley et al [22] showed that cyclophosphamide is able to activate PXR at a concentration >100 M in vitro, which is conWrmed by our results. Clinical proof for our observations that the other anticancer drugs, paclitaxel, docetaxel, Xutamide and erlotinib, have the potential to cause pharmacokinetic drug-drug interaction is not yet known and should be further investigated.…”

“…Paclitaxel [10], cyclophosphamide [22] and tamoxifen [11] were already known to induce CYP3A4 expression via activation of PXR and our data conWrm these results. However, in contrast to other studies [10,25] we observed a small, but signiWcant CYP3A4 reporter activity after treatment with docetaxel.…”

Relative Activation of Human Pregnane X Receptor versus Constitutive Androstane Receptor Defines Distinct Classes of CYP2B6 and CYP3A4 Inducers

“…Compounds that achieved significantly greater than 30% of RIF-normalized hPXR activation of CYP3A4 reporter gene expression at one or more concentrations were classified as moderate to strong activators of hPXR, whereas drugs exhibiting less than 30% were classified as weak or nonactivators of hPXR. Consistent with previous reports, ART, CPA, LOV, MET, MEV, RU486, NIC, NIF, OMP, RES, SIM, and TGZ were classified as moderate to strong hPXR activators (fold activation Ͼ 30% of RIF response) (Drocourt et Kocarek et al, 2002;Lindley et al, 2002;Raucy, 2002;Burk et al, 2005). To our knowledge, this is the first demonstration of hPXR activation by CPZ.…”

“…By combining results from the hPXR-and hCAR3-based reporter gene assays, these inducers were classified as hPXR, hCAR, or hPXR/hCAR dual activators. Our results demonstrate that CMZ, EFV, and NVP induce CYP2B6 and CYP3A4 preferentially through hCAR and that hCAR3 represents a sensitive tool for in vitro prediction of chemical-mediated human CAR activation.CYP3A4 and CYP2B6 are induced at the mRNA, protein, and activity levels by the same compounds, including rifampin, phenobarbital, clotrimazole, cyclophosphamide, calcium channel antagonists, HMG-CoA reductase inhibitors, and thiazolidinediones (Drocourt et al, 2001;Kocarek et al, 2002;Lindley et al, 2002;Sahi et al, 2003;Faucette et al, 2004). Coinduction of these enzymes is mediated through transcriptional activation of the corresponding genes by the nuclear receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR), which are capable of binding to the same response elements in the promoter regions of the CYP3A4 and CYP2B6 genes (Goodwin et al…”

“…CYP3A4 and CYP2B6 are induced at the mRNA, protein, and activity levels by the same compounds, including rifampin, phenobarbital, clotrimazole, cyclophosphamide, calcium channel antagonists, HMG-CoA reductase inhibitors, and thiazolidinediones (Drocourt et al, 2001;Kocarek et al, 2002;Lindley et al, 2002;Sahi et al, 2003;Faucette et al, 2004). Coinduction of these enzymes is mediated through transcriptional activation of the corresponding genes by the nuclear receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR), which are capable of binding to the same response elements in the promoter regions of the CYP3A4 and CYP2B6 genes (Goodwin et al, 1999(Goodwin et al, , 2001Sueyoshi et al, 1999;Wang et al, 2003).…”