Chloroquine (CQ) has previously been shown to exert post-intergrational antiretroviral activity in vitro by affecting newly produced viral envelope glycoproteins (Savarino et al., 2001). Clinical trials are ongoing to characterise the potential use of CQ in vitro.

The antiretroviral activity of CQ, mefloquine (MQ), mepacrine (MC), amadiaquine and halofantrine against HIV_IIIB were determined in U937 cells. Isobolograms were then constructed for CQ and MQ in combination with SQV. CQ and MQ were then tested for their ability to inhibit recombinant HIV protease in a fluorescence based cell free assay. CQ and MQ were also investigated for their ability to alter intracellular concentrations of ³H-SQV in U937 cells, MT4/DOX500 (BCRP-overexpressing), CEM_VBL (P-glycoprotein-overexpressing) and CEM_E1000 (MRP1-overexpressing) cells. Expression of drug transporters P-glycoprotein, Multidrug resistance protein 1 (MRP1) and breast cancer resistance protein (BCRP) was assessed by Western blot analysis and flow cytometry. Finally, the effect of MQ (50mg kg^-1; i.v.) on SQV (10mg kg^-1; i.p.) tissue distribution was determined in male CD1 mice.

A limited selectivity (EC50 against virus / IC50 against cells) was observed for CQ (1.5), mefloquine (MQ; 2.5) and mepacrine (2.3). However, when combined with saquinavir (SQV), antagonism was observed with CQ and synergy observed with MQ. Neither CQ nor MQ altered SQV ability to inhibit protease in the cell-free substrate assay. MQ significantly enhanced the accumulation of SQV (accumulation ratio 26.5 ± 3.1 versus 51.9 ± 11.3; n = 4; p < 0.0001) but assays failed to localise the known SQV transporters P-glycoprotein and MRP1, to U937 cells. Further analysis using flow cytometry showed expression of BCRP, which was significantly down regulated after incubation with MQ (Fluorescence of 3.7 ± 0.2 versus 1.5 ± 1.9; n = 4; p < 0.005). The MT4/DOX500 cell line, which overexpresses BCRP, accumulates significantly lower SQV than the parental cell line (p < 0.01). Finally, a significantly higher penetration of SQV to brain, testis, liver and kidney were seen in CD-1 mice pre-administered with MQ as compared to those pre-administered with vehicle (brain/blood ratio of 0.02 ± 0.01 versus 0.07 ± 0.04; n = 9; p < 0.001).

MQ synergy with SQV in this in vitro system was due to a dual effect on the BCRP protein; direct inhibition of BCRP at the cell membrane coupled with a delayed down-regulation of expression of this transporter. This inhibition is likely to impact on the enhanced sanctuary site penetration in mice. Given the toxicity profile of MQ in man it is important to consider analogues for further investigation.

Savarino, A., et al. (2001). AIDS, 15, 2221-9.