Background:  Phosphonomethoxy-2’-fluoro-2’,3’-dideoxydidehydroadenosine (GS9148) is a nucleotide reverse transcriptase inhibitor (NRTI) with a favorable in vitro profile including activity against drug resistant HIV strains. Amidate prodrug technology previously explored with tenofovir (TFV) was applied to GS9148 to maximize the intracellular accumulation of its active diphosphate (DP) metabolite. This study explored whether the oral administration of GS9148 amidate (ethylalaninyl phenyl derivative) can deliver GS9148-DP into peripheral blood mononuclear cells (PBMC) at levels sufficient for a clinical antiviral effect.

Methods:  Pharmacokinetics were determined in vivo in beagle dogs with plasma and PBMC samples analyzed by LC/MS/MS. The relative intracellular potency of GS9148 was compared to TFV by determining the concentrations of GS9148-DP and TFV-DP sufficient for >90% inhibition of virus replication in vitro in HIV-infected PBMC. Using these data together, the intracellular levels of GS9148-DP required for a clinical effect were extrapolated from known intracellular concentrations of TFV-DP in patients.

Results:  While delivering similar amounts of GS9148 into plasma, a single dose of GS9148 prodrug administered orally to dogs (3 mg/kg) resulted in >90-fold higher intracellular concentrations of GS9148 and GS9148-DP compared to intravenous administration of GS9148 itself (0.5 mg/kg). In the oral arm, GS9148 prodrug showed a bioavailability of >20% with intracellular levels of GS9148-DP reaching 9.0±2.3 µM. The levels of GS9148-DP persisted in PBMC for >24 hours. Drug metabolism screening in vitro indicated that the projected exposure in humans would be at least as high as that observed in dogs. In addition, metabolism and HIV-1 inhibition experiments in activated PBMC demonstrated the intracellular equipotency of GS9148-DP and TFV-DP. Patients treated with clinical doses of TFV had median concentration of 0.5 µM TFV-DP in PBMC, suggesting that ≥0.5 µM intracellular concentrations of GS9148-DP should result in a clinically significant antiviral response.

Conclusions:  Oral administration of GS9148 amidate prodrug to dogs resulted in a substantial accumulation of GS9148-DP in PBMC. Clinically effective concentrations of GS9148-DP in patient PBMC are expected to be readily achieved following once daily oral administration of the GS9148 prodrug at doses <2 mg/kg.