Intracellular carbovir triphosphate levels in patients taking abacavir once a day

Relatively long intracellular half-lives of active triphosphate metabolites support the once daily dosing of nucleoside analogues such as didanosine and lamivudine [1–4]. The guanosine analogue abacavir is phosphorylated intracellularly into carbovir triphosphate [5]. It is not known whether daily abacavir doses of 600 mg produce levels of intracellular carbovir triphosphate similar to those achieved with the standard abacavir dose of 300 mg twice a day. We examined the intracellular levels of the active nucleotide carbovir triphosphate in HIV-positive patients taking abacavir 600 mg once a day, with the aim of determining whether this drug has the potential to be dosed on a once daily basis. Five HIV-positive adults (four men, one woman) taking abacavir 600 mg once a day for 5–17 months as a component of multiple drug rescue therapy including two or three other nucleosides, one non-nucleoside, and one or two protease inhibitors consented to participate in this pharmacokinetic study. Blood samples were drawn into cell preparation tubes (Vacutainer CPT, Becton Dickinson, Franklin Lakes, NJ, USA) at time 0 (24 h after the previous abacavir dose) and 1, 12, 14–16, 18, 20, 22, and 24 h after an observed 600 mg abacavir dose. Peripheral blood mononuclear cells were isolated and phosphates extracted using 60% methanol. The methanol extracts were then dried and stored at −70°C. After a second extraction with perchloric acid, endogenous deoxyguanosine triphosphate (dGTP) levels were determined by a DNA polymerase assay. For the measurement of carbovir triphosphate, reaction mixtures included 3H-dGTP, template primer and reverse transcriptase in a total volume of 50 μl. Radioactivity bound to the primer was separated by spotting onto filter papers and washing, and was then determined by liquid scintillation counting [6]. Carbovir triphosphate and dGTP concentrations are shown in Table 1. The half-life of carbovir triphosphate was estimated to be greater than 12 h (data analysed by topfit, Gustav Fischer Verlag, Stuttgart, Germany). An interesting observation was that the female patient's carbovir triphosphate levels were consistently higher (by two to eightfold) than those of the four men.Table 1: Carbovir triphosphate and deoxyguanosine triphosphate concentrations. A previous study using the same assay [7] demonstrated a mean carbovir triphosphate concentration of 100 fmoles/106 cells (200 nmol/l) 12 h after a 300 mg abacavir dose. In our study, one patient had intracellular carbovir triphosphate levels of 80, 107 and 62 fmoles/106 cells at 20, 22 and 24 h, respectively. Otherwise, in these five patients intracellular carbovir triphosphate levels greater than 100 fmoles/106 cells (greater than 200 nmol/l) were sustained over the entire 24 h period after a single 600 mg abacavir dose. These preliminary results of 24 h carbovir triphosphate concentrations support the use of once daily abacavir. Further clinical studies are needed before this dose can be generally recommended. Marianne Harrisa David Backb Sean Kewnc Salima Juthaa Raymond Marinac Julio S. G. Montanera