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Pregnane X receptor and constitutive androstane receptor activation by non-nucleoside HIV-1 reverse transcriptase inhibitors Sharma, Devinder


Non-nucleoside reverse transcriptase inhibitors (NNRTIs) are important anti-retroviral drugs indicated in combination therapy of human immunodeficiency virus-1 (HIV-1) infection. NNRTI therapy is associated with pharmacokinetic drug interactions, the underlying mechanisms of which are poorly understood. The present study investigated the effects of NNRTIs on the activity of pregnane X receptor (PXR) and constitutive androstane receptor (CAR), key transcriptional factors regulating the expression of various drug-metabolizing enzymes and transporters. The experimental approaches included cell-based luciferase reporter gene assays, in vitro competitive ligand binding assay, nuclear translocation analysis by confocal imaging, coactivator recruitment assays, and target gene expression determination in primary human hepatocytes. Rilpivirine, etravirine, and efavirenz, but not nevirapine or delavirdine, were identified as agonists of PXR and inducers of cytochrome P450 3A4 (CYP3A4), a target gene of PXR. By comparison, rilpivirine, etravirine, and efavirenz, but not nevirapine or delavirdine, were indirect activators of the wild-type isoform of CAR and inducers of cytochrome P450 2B6, a target gene of CAR. Among the NNRTIs investigated, only efavirenz activated the SV23 and SV24 splice variants of CAR, indicating that NNRTIs activated CAR in a drug-specific and isoform-selective manner. To further understand PXR regulation by rilpivirine, the role of microRNA in rilpivirine activation of PXR was investigated. MicroRNAs are small non-coding RNAs that are post-transcriptional regulators causing mRNA degradation or translational repression by binding to complementary regions in target mRNA. Bioinformatic analysis (www.microRNA.org) predicted sequence complementarity between hsa-miR-18a-5p and PXR. Reporter gene assays revealed a functional hsa-miR-18a-5p microRNA recognition element in the 3′-untranslated region of PXR. In cell-based assays, over-expression of hsa-miR-18a-5p by transfecting LS180 human colon adenocarcinoma cells with a mimic of hsa-miR-18a-5p decreased PXR mRNA and protein expression. Rilpivirine and rifampin did not affect PXR expression, but it decreased endogenous expression of hsa-miR-18a-5p in LS180 cells. In contrast, over-expression of hsa-miR-18a-5p decreased PXR mRNA expression and CYP3A4 inducibility by rilpivirine and rifampin. These data suggest that hsa-miR-18a-5p regulates PXR and contributes to drug activation of PXR. Overall, the present study shows activation of PXR and CAR by select NNRTIs and provides mechanistic understanding of NNRTI-mediated drug-drug interactions.

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