Intracellular Tenofovir-Diphosphate and Emtricitabine-Triphosphate in Dried Blood Spots following Directly Observed Therapy

Abstract

Studies of daily emtricitabine-tenofovir disoproxil fumarate (FTC-TDF) for HIV preexposure prophylaxis (PrEP) in men who have sex with men (MSM) modeled intracellular tenofovir-diphosphate (TFV-DP) in dried blood spots (DBS) to assess adherence and corresponding PrEP outcomes. We conducted a prospective, randomized, crossover pharmacokinetic study of TFV-DP in DBS during 33%, 67%, or 100% of daily dosing under directly observed therapy (DOT). Participants were assigned to two 12-week dosing regimens, separated by a 12-week washout. Forty-eight adults (25 women) from Denver and San Francisco were included. TFV-DP exhibited a median half-life of 17 days, reaching steady state in 8 weeks. TFV-DP was dose proportional with mean (SD) steady-state concentrations of 530 (159), 997 (267), and 1,605 (405) fmol/punch for the 33%, 67%, and 100% arms, respectively. Prior work in MSM demonstrated clinically meaningful TFV-DP thresholds of 350, 700, and 1,250 fmol/punch, which were estimated 25th percentiles for 2, 4, and 7 doses/week. In the present study, corresponding TFV-DP was within 3% of the prior estimates, and subgroups by site, race, and sex were within 14% of prior estimates, although males had 17.6% (95% confidence intervals [CIs], 6.5, 27.4%) lower TFV-DP than females. The thresholds of 350, 700, and 1,250 fmol/punch were achieved by 75% of men taking ≥1.2, 3.2, and 6 doses/week and 75% of women taking ≥0.6, 2.0, and 5.3 doses/week, indicating that lower dosing reached these thresholds for both sexes. In conclusion, TFV-DP arising from DOT was similar to previous estimates and is useful for interpreting PrEP adherence and study outcomes. (This study has been registered at ClinicalTrials.gov under identifier NCT02022657.).

Adherence Measurements in HIV: New Advancements in Pharmacologic Methods and Real-Time Monitoring

“…Plasma and urine drug and metabolite concentrations provide proof of drug ingestion and can be predictive of virologic failure in diverse settings[22, 23]. However, their utility is limited by the short half-lives (~10–20 hrs) that most antiretrovirals have in these matrices (Table 1), and by the fact that single-dose concentrations usually match steady-state concentrations[24]. Due to this limitation, plasma and urine drug concentrations are usually only able to detect drug intake within a short period of time (days to one week).…”

“…This preference for DBS reflects various advantages over traditional plasma blood sampling; it can be collected in small quantities from venipuncture or needlestick, does not require special sample processing and is associated with a substantial cost benefit[29]. Likewise, DBS samples can be stored for relatively long periods of time after collection and can be shipped without special biohazard precautions[24, 30]. These advantages have led to the use of DBS sampling for analysis beyond the neonatal period, including rapid diagnostics of vector-borne diseases[31], identification of disease-associated genetic mutations[32], diagnosis of HIV infection, and quantification of HIV VL[33–36], among others.…”

“…Parallel to the quantification of antiretroviral parent drug concentrations in DBS, this matrix has also been evaluated for the measurement of intracellular concentrations of antiretroviral anabolites in red blood cells (RBCs), given the abundance of RBCs in DBS[24, 29, 50–52]. This area of research has focused on TFV and emtricitabine (FTC), because these nucleoside analogues are also phosphorylated inside of RBCs in a similar fashion to their metabolism in peripheral blood mononuclear cells (PBMCs), where they are phosphorylated to their pharmacologically-active moieties TFV diphosphate (TFV-DP) and FTC triphosphate (FTC-TP), respectively[29].…”

“…This area of research has focused on TFV and emtricitabine (FTC), because these nucleoside analogues are also phosphorylated inside of RBCs in a similar fashion to their metabolism in peripheral blood mononuclear cells (PBMCs), where they are phosphorylated to their pharmacologically-active moieties TFV diphosphate (TFV-DP) and FTC triphosphate (FTC-TP), respectively[29]. Once phosphorylated, TFV-DP, it is trapped and accumulates 25-fold from first dose to steady state in RBCs, with an intracellular half-life of 17 days[24, 29]. These unique pharmacologic characteristics were used in pharmacokinetic modeling analyses to develop a TFV-DP gradient that can be used to estimate TDF-FTC dosing over a preceding period of approximately 8 weeks[29].…”

“…These unique pharmacologic characteristics were used in pharmacokinetic modeling analyses to develop a TFV-DP gradient that can be used to estimate TDF-FTC dosing over a preceding period of approximately 8 weeks[29]. These findings were recently confirmed using directly-observed dosing in healthy volunteers, in whom the TFV-DP concentration benchmarks for daily TDF-FTC dosing were established[24]. This methodology has also been extended recently to quantify TFV-DP arising from tenofovir alafenamide (TAF)[52], with an ongoing study to establish the drug concentrations associated with different adherence benchmarks in healthy volunteers (NCT02962739).…”