Background: Rilpivirine (RPV) is mainly metabolised by cytochrome P450 (CYP) enzymes but less is known about the importance of transporters in its disposition. Many enzymes and transporters are transcriptionally regulated by the pregnane-X-receptor (PXR; NR1I2). In this study, we sought to determine associations between genetic variants within the genes coding for these proteins and RPV plasma concentrations. Methods: Blood samples were collected 12 hours post dose (C12) from 150 patients receiving RPV-containing regimens recruited in Turin (Italy) and London (UK). Plasma concentrations of RPV were analysed by validated LC-MS/MS methods. Genotyping was conducted by real-time PCR-based allelic discrimination using standard methods for the following polymorphisms: CYP3A4*22 (rs35599367), CYP2C19*2 (rs4244285), CYP2C19*17 (rs12248560), NR1I2 (rs2472677), SLC22A1 (rs628031, rs72552763, rs622342 and rs683369), SLCO2B1 (rs1077858, rs12422149, rs35199625, rs2712807 and rs2851069).