A Novel Anti-HIV Particle Provides Sustained Plasma and Cell Drug Levels in Lymph Nodes in Primates

Background: We have reported previously that an anti-HIV drug Indinavir bound to lipid nanoparticles (LNP) provide pH responsive release and extended plasma drug exposure and accumulation in lymph nodes in primates after subcutaneous dosing. The goal of this study is to develop and evaluate a HIV drug combination intended for a maximal suppression of residual virus. To do so we constructed an anti-HIV LNP containing two protease inhibitors [Lopinavir (LPV) and Ritonavir (RTV) and one nucleotide analogue reverse transcriptase inhibitor [Tenofovir (TNFV)] and compared the LNP versus free drug formulation for plasma time-course and intracellular drug levels in lymph nodes. Methodology: The LNPs composed of lipids (DSPC: mPEG2000-DSPE) (8:2 m/m); and anti-HIV drugs-LPV, RTV and RNFV (115:10:5:15 lipids:LPV:RTV:TNFV m/m/m/m) were produced by solvent evaporation, followed by rehydration inbuffered saline and homogenization with an Avestin Emulsiflex C5. Near-complete LNP incorporation of Lopinavir and Ritonavir and reproducible Tenofovir encapsulation (~5%) were verified. The anti-HIV LNP diameter was 50-80 nm. pH, osmolarity, and sterility were verified. Macaques (M. Nemestrina) (n=4/gp) received free or LNP drug formulation subcutaneously. Blood and peripheral blood mononuclear cells (PBMCs) were collected for 24 hours and at day-7. The inguinal lymph node was collected at 24 hours and day-7. Drug concentrations were determined by liquid chromatography tandem mass spectroscopy (LC-MS/MS-QTRAP). Results: Primates dosed subcutaneously with anti-HIV particles composed of LPV: RTV:TNFV at 25:14:17 mg/kg provided sustained plasma drug levels for more than 7 days. In contrast, animals treated with an identical dose in free and soluble form exhibited plasma and lymph node drug levels that decline rapidly below the detection limit by 24 hrs; and exhibited significant local skin reactions that persisted for more than 2 weeks, Animals that received LNP formation did not exhibit local reaction. Treatment with anti-HIV LNP provided significant cellular drug concentrations at 24 hrs. In addition, mononuclear cells dissociated from lymph nodes also contained significant levels of LPV, RTV, and TNFV in the cells even 7 days after a single-dose anti-HIV LNP administration. Conclusions: Anti-HIV lipid nanoparticles containing the three drugs, Lopinavir, Ritonavir and Tenofovir, demonstrated the ability of prolonged drug presence in plasma, PBMCs, and lymph node cells beyond levels achievable by free, non-particulate drug dosage form after a single subcutaneous administration in primates. Incorporation of the three drugs into LNP also reduced local side effects.