Direct evidence that saquinavir is transported by multidrug resistance-associated protein (MRP1) and canalicular multispecific organic anion transporter (MRP2).

To determine if saquinavir mesylate (saquinavir) is a substrate of human multidrug resistance-associated protein 1 (hMRP1 [ABCC1]) or hMRP2 (cMOAT, or ABCC2), MDCKII cells that overexpress either hMRP1 (MDCKII-MRP1) or hMRP2 (MDCKII-MRP2) were used to investigate saquinavir's cytotoxicity and transport in comparison with those of control MDCKII wild-type (MDCKII/wt) cells. Cytotoxicity was assessed with the mitochondrial marker MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium], and saquinavir transport was measured directly through the cell monolayers. GF120918 (an inhibitor of P glycoprotein, but not of the MRP family) and MK-571 (an MRP family inhibitor) were used to delineate the specific contributions of these transporters to saquinavir cytotoxicity and transport. In the presence of GF120918 and increasing saquinavir concentrations, the MDCKII-MRP1 (50% lethal dose [LD(50)] = 10.5 micro M) and MDCKII-MRP2 (LD(50) = 27.1 micro M) cell lines exhibited statistically greater viability than the MDCKII/wt cells (LD(50) = 7.8 micro M). Saquinavir efflux was directional, not saturable, and was inhibited by MK-571 (35 and 75 micro M) in all cell lines. The ratios of saquinavir (3 micro M) basolateral to apical permeability (i.e., efflux ratios) for the MDCKII/wt, MDCKII-MRP1, and MDCKII-MRP2 cell monolayers were 2.6, 1.8, and 6.8, respectively. The MDCKII-MRP1 cells have a significantly reduced saquinavir efflux ratio relative to MDCKII/wt cells, due to basolaterally directed transport by hMRP1 competing with endogenous, apically directed canine MRP2. The MDCKII-MRP2 cells have a significantly increased saquinavir efflux ratio relative to MDCKII/wt cells, due to the additive effects of the apically directed transport by hMRP2 and endogenous MRP2. Collectively, the cytotoxicity and transport results provide direct evidence that saquinavir is transported by MRP1 and MRP2.