Interestingly, although β-estradiol modestly inhibits HIV-1 replication in PBL, it increases HIV-1 replication in the presence of a fixed amount of D4T, and this increase is specifically dependent on the anti-retroviral effect of the drug. Thus β-estradiol seems to decrease the efficacy of D4T against HIV-1 infection of PBL. The data suggests that the magnitude of the effect on D4T efficacy is such that approximately at least a two fold increase in the concentration of D4T would be necessary to overcome the effects of the hormone.
β-estradiol increased the amount of HIV-1 replication in the presence of D4T from a baseline of 33% (of VA, +/- SE = 5.4) to 74% (+/- SE = 5.4), (Tables &, Figure & Figure ) whereas progesterone had little or no effect on viral replication in the presence (or absence) of D4T (Figure ). The concentrations of D4T used here for viral inhibition are within range of levels typically used for tissue culture work [7
] and have not been reported to cause significant cytotoxicity. Nevertheless, we did test to see if the combinations of drugs and hormones studied caused detectable non-specific cytotoxicity in PBL. Even excessively high concentrations of D4T (1 uM) caused no cytotoxicity in the presence or absence of β-estradiol, as measured by trypan blue exclusion. β-estradiol alone also caused no cytotoxicity (Table ).
The mechanism by which β-estradiol promotes HIV replication in the presence of D4T remains unknown. However, we have observed β-estradiol has no (or minimal) effect on HIV replication in the presence of the protease inhibitor, Saquinavir (unpublished observations). The finding that in the absence of D4T β-estradiol inhibits HIV replication, whereas in the presence of D4T it enhances HIV replication, strongly suggests that the mechanism of the enhancement is D4T-specific. In confirmation of this, we determined that β-estradiol has no effect on HIV replication in the presence of D4T when the HIV is resistant to D4T. Thus, the observed enhancement is most likely on the anti-retroviral efficacy of D4T. This is consistent with β-estradiol inhibiting the concentration or activity of the cellular enzymes used to phosphorylate D4T to its active form, D4T-TP, but does not rule out a mechanism involving changes in drug influx or efflux. Experiments to address these issues are currently ongoing in our laboratory.
The inhibition of antiretroviral drug efficacy by estrogen may have implications for anti-HIV-1 drug therapies. The studies presented here put forth the novel concept that at any given plasma concentration of drug, the final efficacy may be significantly affected by the hormone status of the patient. Most likely, β-estradiol acts by modifying intracellular levels of the active form of D4T through mechanisms which may include controlling drug influx or efflux or, more likely, controlling the phosphorylation steps which lead to the D4T-TTP active form of D4T [11
]. Thus, monitoring of β-estradiol levels, which vary during pregnancy, menstrual cycling and with hormone replacement therapy and birth control, or monitoring intracellular levels of active drug may provide significant added benefit over monitoring plasma levels of drug alone.