To determine if saquinavir mesylate (saquinavir) is a substrate of individual multidrug resistance-associated proteins 1 (hMRP1 [ABCC1]) or hMRP2 (cMOAT or ABCC2) MDCKII cells that overexpress possibly hMRP1 (MDCKII-MRP1) or hMRP2 (MDCKII-MRP2) were used to research saquinavir’s cytotoxicity and transportation in comparison to those of control MDCKII wild-type (MDCKII/wt) cells. dosage [LD50] = 10.5 μM) and MDCKII-MRP2 (LD50 = 27.1 μM) CCG-63802 cell lines exhibited statistically better viability compared to the MDCKII/wt cells (LD50 = 7.8 μM). Saquinavir efflux was directional not really saturable and was inhibited by MK-571 (35 and 75 μM) in every cell lines. The ratios of saquinavir (3 μM) basolateral to apical CCG-63802 permeability (i.e. efflux ratios) for the MDCKII/wt MDCKII-MRP1 and MDCKII-MRP2 cell monolayers had been 2.6 1.8 and 6.8 respectively. The MDCKII-MRP1 cells possess a significantly decreased saquinavir efflux proportion in accordance with MDCKII/wt cells because of basolaterally directed transportation by hMRP1 contending with endogenous apically directed canine MRP2. The MDCKII-MRP2 cells possess 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 CCG-63802 provide direct evidence that saquinavir is usually transported by MRP1 and MRP2. The oral bioavailabilities of the human CD97 immunodeficiency computer virus (HIV) protease inhibitors (saquinavir ritonavir indinavir nelfinavir and amprenavir) are low and/or variable with limited penetration into the central nervous system (CNS) (18). Saquinavir mesylate was the first drug approved in this class. The two marketed saquinavir capsule formulations have mean oral bioavailabilities that range from 4 to 16% and are highly variable as indicated by area under the concentration-time curve (AUC) coefficients of variation that are ≥30% (11). Saquinavir’s low and variable bioavailability is usually primarily attributed to metabolism by cytochrome P-450 3A4 (27). However there is increasing understanding CCG-63802 that membrane transporters contribute significantly to the biopharmaceutic characteristics of saquinavir and this entire class of medications. To connect bioavailability to molecular transportation features we like numerous others speculated an efflux (countertransport) system might donate to the reduced and adjustable bioavailability of saquinavir and various other HIV protease inhibitors. Saquinavir efflux (basolateral to apical [BL→AP] permeability > AP→BL permeability) as well as the inhibition of saquinavir efflux with verapamil hydrochloride a substrate for multiple transporters and a non-specific efflux inhibitor had been demonstrated using a Caco-2 cell model (3). In primary work we confirmed that saquinavir efflux from rat intestinal tissues is an energetic process (27). Nevertheless these research didn’t identify the transporter or transporters involved within these complex tissues particularly. Since Caco-2 cells and rat intestinal tissues are recognized to exhibit multiple transporters the noticed saquinavir transportation behavior could be linked to multiple transporters with multiple affinities. Latest studies show the fact that HIV protease inhibitors are substrates for the P glycoprotein (Pgp [ABCB1]) efflux pump and also have demonstrated decreased saquinavir cytotoxicity because of saquinavir transportation by Pgp (17). Saquinavir transportation by Pgp in addition has been correlated with minimal bioavailability and CNS penetration (18). Nevertheless saquinavir transportation by Pgp will not eliminate saquinavir being truly CCG-63802 a substrate for various other putative membrane transporters. To the end various CCG-63802 other investigators have confirmed that saquinavir inhibits multidrug resistance-associated proteins (MRP) family members (MRP1 and MRP2)-mediated transportation (15 20 24 and a fluorescent saquinavir derivative is certainly carried by MRP2 (14). Additionally intracellular saquinavir concentrations had been been shown to be low in MRP1-expressing individual lymphocytes in accordance with those in charge cells (15 16 To see whether unmodified saquinavir is certainly a substrate of individual MRP1 (hMRP1 [ABCC1]) or the individual canalicular multispecific organic anion transporter hMRP2 (cMOAT or ABCC2) MDCKII cells that overexpress either hMRP1 (MDCKII-MRP1) or hMRP2 (MDCKII-MRP2) had been used to research saquinavir’s cytotoxicity and transportation in comparison to those of control MDCKII wild-type (MDCKII/wt) cells (15 16 The outcomes of this study demonstrate saquinavir transport by hMRP1 and hMRP2 implicating these transporters in the reduced oral bioavailability and distribution of saquinavir. MATERIALS AND METHODS Chemicals. Saquinavir mesylate (hereafter “saquinavir”) and.