The constitutive androstane receptor is a novel therapeutic target facilitating cyclophosphamide-based treatment of hematopoietic malignancies

Abstract

Key Points• The constitutive androstane receptor as a novel molecular target can facilitate cyclophosphamide-based chemotherapy.Cyclophosphamide (CPA) is one of the most widely used chemotherapeutic prodrugs that undergoes hepatic bioactivation mediated predominantly by cytochrome P450 (CYP) 2B6. Given that the CYP2B6 gene is primarily regulated by the constitutive androstane receptor (CAR, NR1I3), we hypothesize that selective activation of CAR can enhance systemic exposure of the pharmacologically active 4-hydroxycyclophosamide (4-OH-CPA), with improved efficacy of CPA-based chemotherapy. In this study, we have developed a unique human primary hepatocyte (HPH)-leukemia cell coculture model; the chemotherapeutic effects of CPA on leukemia cells can be directly investigated in vitro in a cellular environment where hepatic metabolism was well maintained. Our results demonstrated that activation of CAR preferentially induces the expression of CYP2B6 over CYP3A4 in HPHs, although endogenous expression of these enzymes in leukemia cells remains negligible. Importantly, coadministration of CPA with a human CAR activator led to significantly enhanced cytotoxicity in leukemia cells by inducing the apoptosis pathways, without concomitant increase in the off-target hepatotoxicity. Associated with the enhanced antitumor activity, a time and concentration-dependent increase in 4-OH-CPA formation was observed in the coculture system. Together, our findings offer proof of concept that CAR as a novel molecular target can facilitate CPA-based chemotherapy by selectively promoting its bioactivation. (Blood. 2013;121(2):329-338) IntroductionHematopoietic malignancies are a group of heterogeneous disorders associated with considerably variable prognoses, depending in large part on the specific diagnosis and available treatment options. Cyclophosphamide (CPA), an alkylating prodrug, has been widely used in combination with other antineoplastics in the treatment of various cancers, including hematologic neoplasms, such as lymphoma and leukemia. [1][2][3][4] Nevertheless, a significant number of patients succumb to their disease despite standard therapeutic regimens. 5 The need for further optimization of current treatment paradigms is evident. Given that CPA is a prodrug requiring metabolic activation by hepatic drug-metabolizing enzymes, CPAbased chemotherapy could be improved by selectively enhancing the metabolic conversion of CPA to the pharmacologically active metabolite but not to the nontherapeutic byproducts.On administration, CPA undergoes hepatic oxidation to form the therapeutically active intermediate metabolite, 4-hydroxycyclophosphamide (4-OH-CPA), primarily mediated by cytochrome P450 (CYP) 2B6 and to a lesser extent by CYP3A4 and CYP2C9. 6,7 4-OH-CPA is further tautomerized to aldophosphamide, followed by spontaneous ␤-elimination to release the phosphoramide mustard that exerts chemotherapeutic effects by attacking specific nucleophilic groups of DNA molecules in target cancer cells. 8 Alternatively, CPA is subjec...

Targeting xenobiotic receptors PXR and CAR in human diseases

“…Wang et al showed that selective activation of CAR and subsequent induction of CYP2B6 in human primary hepatocytes enhanced the bioactivation of chemotherapeutic prodrug cyclophosphamide. Therefore activator of human CAR might improve the efficacy of cyclophosphamide-based chemotherapy [52]. …”

Small-molecule modulators of the constitutive androstane receptor

“…As a prodrug, CPA is converted in the body to its active form mainly by CYP2B6. Because CYP2B6 is a major target of CAR, in hepatocyte co-cultures with a leukemia cell line, CAR activation (with agonist CITCO) can enhance CPA-based antitumor activity and apoptosis [120] by increasing the availability of the active form. Recently, the common chemotherapeutic paclitaxel was used to determine that CAR agonists modestly increase paclitaxel-induced tumor depletion in 5 human lung cancer cell lines [121].…”

The Role of Bile Salt Export Pump Gene Repression in Drug-Induced Cholestatic Liver Toxicity

“…Human primary hepatocytes seeded 7.5 Â 10 4 cells/well in 96-well plates were cultured for 48 hours before treatment with BSEP inhibitors at previously indicated concentrations. A typical 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay was carried out as described previously (Wang et al, 2013). Cell viability was expressed as percent of vehicle control (0.1% DMSO).…”