Previous work has demonstrated the existence of systemic interaction between tenofovir (TFV) disoproxil fumarate (TDF) and didanosine as well as between TDF and lopinavir-ritonavir (LPV/r). Here we investigated TDF interactions with the nucleoside reverse transcriptase inhibitors (NRTIs) lamivudine (3TC) and abacavir (ABC), comparing both the concentrations of nucleoside/nucleotide reverse transcriptase inhibitors in plasma and the intracellular concentrations of their triphosphate metabolites (NRTI-TP) for human immunodeficiency virus-infected patients receiving these NRTIs with TDF and after 4 weeks of TDF interruption. We also looked at interactions between TDF-ABC and LPV/r, comparing patients receiving or not receiving LPV/r. Blood samples were taken at baseline and at 1, 2, and 4 h after dosing. Liquid chromatography-tandem mass spectrometry was used to measure NRTIs and NRTI-TPs. Statistical analyses were performed on pharmacokinetic parameters: the area under the concentration-time curve from 0 to 4 h (AUC 0-4 ), the maximum concentration of the drug (C max ), and the residual concentration of the drug at the end of the dosing interval (C trough ) for plasma and the AUC 0-4 and C trough for intracellular data. Among the groups of patient discontinuing TDF, the very long intracellular half-life of elimination (150 h) of TFV-DP (the diphosphorylated metabolite of TFV, corresponding to a triphosphorylated species) was confirmed. Comparison between groups as well as the longitudinal study showed no significant systemic or intracellular interaction between TDF and ABC or 3TC. Significant differences were observed between patients receiving LVP/r and those receiving nevirapine. For ABC, plasma exposure was decreased (40%) under LVP/r, while, in contrast, plasma exposure to TFV was increased by 50% and the intracellular TFV-DP AUC 0-4 was increased by 59%. A trend for a gender effect was observed for TFV-DP at the intracellular level, with higher and C trough values for women.Highly active antiretroviral therapy regimens include at least three antiretroviral agents combining a nucleoside/nucleotide reverse transcriptase inhibitor [N(t)RTI] backbone with a protease inhibitor (PI) or a nonnucleoside reverse transcriptase inhibitor (NNRTI) (51). The issue of drug-drug interaction between NRTIs, including N(t)RTIs, and also between N(t)RTIs and PIs or NNRTIs needs to be addressed (37).All these drugs or prodrugs could interact at the systemic level (absorption, metabolism, distribution, or elimination) or at the cellular level through membrane transport, or through intracellular metabolism for the N(t)RTIs.Several in vitro and in vivo intracellular interactions have been reported between NRTIs sharing the same metabolic pathways for thymidine analogues (14, 17) and for cytidine analogues (24).More recently, a significant interaction between didanosine (ddI) and tenofovir (TFV) disoproxil fumarate (TDF) has been clinically demonstrated (6,23,29,32), showing a higher plasma exposure to ddI when it is combined with TD...