No Evidence for Induction of ABC Transporters in Peripheral Blood Mononuclear Cells in Humans after 14 Days of Efavirenz Treatment

Abstract

Intracellular concentrations of antiretroviral drugs in peripheral blood mononuclear cells (PBMCs) are an important determinant of therapeutic success. In vitro data indicate that efavirenz induces several ATP-binding cassette (ABC) transporters, and pharmacogenetic studies found an association between ABCB1(C3435T) and efavirenz exposure and between this polymorphism and improved virological outcomes. We therefore aimed to clarify whether efavirenz also induces ABC transporters in vivo in PBMCs and whether intracellular concentrations might be altered after induction. Twelve healthy individuals received multiple oral doses of efavirenz over 14 days (400 mg once daily). Blood samples were drawn on study days 1 (single dose) and 14 (multiple dose), and efavirenz concentrations were analyzed by liquid chromatography-tandem mass spectrometry. Expression of P glycoprotein (P-gp) and of the multidrug resistance-associated proteins 1 and 2 as well as P-gp activity was analyzed in PBMCs on day 1 and day 14 using real-time reverse transcription-PCR (RT-PCR) and rhodamine 123 efflux. Although a clear autoinduction could be confirmed by a significant decrease of efavirenz exposure from day 1 to day 14, efavirenz did not change expression of the ABC transporters or P-gp activity in PBMCs. Moreover, intracellular concentrations of efavirenz were 1.3-to 1.8-fold higher than the corresponding plasma concentrations, and the intracellular/plasma concentration ratio remained constant during the treatment and did not correlate with ABC transporter expression or function. In conclusion, our study confirmed that intracellular concentrations of efavirenz are independent from these efflux transporters and demonstrated for the first time that the transporters are not induced in PBMCs in vivo after 2 weeks of treatment with efavirenz.

Population Pharmacokinetic Model Linking Plasma and Peripheral Blood Mononuclear Cell Concentrations of Efavirenz and Its Metabolite, 8-Hydroxy-Efavirenz, in HIV Patients

“…This suggests that intracellular entry might not involve carrier-mediated transport mechanisms that are saturable in the concentrations observed. This observation may be strengthened by the fact that attempts to investigate whether EFV is a substrate of any of the influx or efflux transport processes have not led to any conclusive outcomes (26,27). Moreover, in the present study, covariate analysis between the population PK parameters and influx and efflux transporter systems did not reveal any significant impact of the transporters on the PK of EFV and 8OHEFV.…”

“…Isolation of the PBMCs from the plasma was done by centrifugation and subsequent cell counting, as described previously (26,35). Briefly, 8-ml venous blood samples were collected in BD Vacutainer CPT tubes, in duplicate.…”

Importance of Ethnicity, CYP2B6 and ABCB1 Genotype for Efavirenz Pharmacokinetics and Treatment Outcomes: A Parallel-Group Prospective Cohort Study in Two Sub-Saharan Africa Populations

“…Blood samples were centrifuged (1700 g for 20 min) and plasma and peripheral blood mononuclear cells (PBMC) were prepared as described by Burhenne et al [25] and stored at −80°C. Samples were sent on dry ice to the Department of Clinical Pharmacology and Pharmacoepidemiology, University of Heidelberg, Germany where plasma and intracellular efavirenz and 8-hydroxyefavirenz concentrations were determined by liquid chromatography-tandem mass spectrometry (LC/MS/MS) as described previously [6], [25]. Efavirenz and 8-hydroxyefavirenz were quantified using 13 C 6 -efavirenz and 2 H 4 -8-hydroxyefavirenz as internal standards and electrospray tandem mass spectrometry.…”

PharmGKB summary

“…PBMC mRNA expression of ABCB1 was significantly reduced at day 14 from day 1 of EFV treatment in healthy volunteers, though P-gp enzymatic activity in PBMCs was not altered by treatment [55, 56]. Polymorphisms in ABCB1 may influence the PK of drugs taken concomitantly with EFV, and thus may affect EFV levels via drug-drug interactions.…”