Saquinavir mesylate (SQV) is the first-in-class and prototypical HIV protease inhibitor (PI) used in the treatment of HIV infection. SQV undergoes extensive hepatic metabolism and intestinal and bile secretion, and has poor and variable oral bioavailability. In previous studies, our group and others have described the interactions between SQV and absorptive and secretory efflux transporters such as MRP1, MRP2, and P-gp. However, the potential role of absorptive influx transporters such as OATP-A (SLC21A3) has not yet been reported for SQV. In the study presented here, the role of OATP-A in the influx transport of SQV was studied using a hepatic cell model, Hep G2, and Xenopus laevis oocytes overexpressing human OATP-A. In Hep G2 cells, SQV transport was found to be (i) concentration-dependent and saturable, (ii) temperature-sensitive, and (iii) proton (pH)- and sodium-independent. While GF120918, a specific inhibitor of P-gp, and MK571, a MRP transporter family inhibitor, significantly enhanced SQV uptake, estrone 3-sulfate, a substrate of OATP-A, significantly inhibited SQV uptake by Hep G2 cells. The observation that inhibitors of P-gp, MRP, or OATP-A have opposite effects on SQV uptake in polarized Hep G2 cells is consistent with their functions as hepatic efflux or influx transporters. In X. laevis oocytes into which OATP-A cRNA had been injected, the level of uptake of SQV was significantly greater than the level of uptake by oocytes into which water had been injected and was concentration-dependent and saturable (Km = 36.4+/-21.8 microM). This is the first report showing that SQV influx transport is directly facilitated by OATP-A. Given the wide body distribution of OATP-A, the current results suggest a potentially important role for OATP-A in the absorption and disposition of SQV in vivo. The data also suggest that in human hepatocytes basolaterally located OATP-A (influx transporter) may act in concert with apically located P-gp and/or MRP2 (efflux transporters) for the vectorial transport and excretion of SQV into bile.