Background:  Drug-drug interactions remain an important cause of morbidity and mortality associated with HIV drug therapy. Recent studies have shown that activation of certain regulatory proteins, called adopted orphan nuclear receptors--such as the pregnane X receptor (PXR), constitutive androstane receptor (CAR), and farnesoid X receptor (FXR)--result in the induction of many CYP enzymes and transporters. Indeed some protease inhibitors (PI) are known to be ligands for PXR. However, the potential clinical relevance of such findings in the setting of multiple drug combinations remains to be defined.

Methods:  A cell-based reporter assay was used to identify interactions of nuclear receptors with PI, either as single agents or combinations. HepG2 cells were  transfected with reporter gene constructs containing human CYP3A4 promoter (for PXR- and CAR-dependent activation) or BSEP promoter (for FXR-dependent activation). In addition, primary human hepatocytes were used as a model of the in vivo inductive response.

Results:  In single-agent studies, we observed that most PI are potent PXR agonists:  amprenavir (EC₅₀: 5.2 mM), lopinavir (EC₅₀: 1.5 mM), nelfinavir (EC₅₀: 6.1 mM), ritonavir (EC₅₀: 0.5 mM). Interestingly, when such PI were combined with rifampin, a prototypical PXR agonist, the extent of maximal CYP3A4 transcriptional activation was less than that with rifampin alone, suggesting that the PI tested are partial agonists of the receptor. Similar findings were obtained using Kaletra (ritonavir+lopinavir) combination as compared with ritonavir alone. Interestingly, the 3-PI combination (lopinavir+ritonavir+ amprenavir) appeared to have a synergistic effect on PXR.  Among the PI tested, nelfinavir and ritonavir appeared to interact with CAR but only nelfinavir activated FXR. Preliminary studies using primary human hepatocytes indicated that selected combination of PI attenuate the CYP3A inductive response, consistent with the in vitro reporter assays.

Conclusions:  Our studies indicate that many PI are in fact partial agonists of PXR, and that certain combinations attenuate the inductive response. In  addition, nelfinavir, but not other PI, is an activator of FXR, the nuclear receptors critical for cholesterol metabolism and bile acid homeostasis. Moreover, our data suggest that in vitro nuclear receptors activation studies may be able to better predict the in vivo inductive response of combination PI therapies and optimal dosing regimens.

Keywords: protease inhibitors; nuclear receptors; drug interactions