Purpose: Antigen presenting cells, such as macrophages and dendritic cells, are pivotal in the innate-to-adaptive immune response transition and therefore they are perfect candidates to target either at the site of injection or at draining lymph nodes in vaccine design. Furthermore, as interaction between cell and antigen ultimately requires a physical encounter, vaccine biodistribution is an important factor to consider. The aim of this work was to compare the uptake of three cationic liposomal delivery systems by macrophage in vitro and in vivo biodistribution within mice.
Methods: Cationic liposomes, composed of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) and 1,2-stearoyl-3-trimethylammonium-propane (DOTAP), 3ß-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol hydrochloride (DC-Chol) or dimethyldioctadecylammonium at 1:1 mole ratio (DOPE:DOTAP, DOPE:DC-Chol) were formulated in the NanoassemblrTM platform, dialyzed against TRIS buffer and subsequently characterized in terms of size, polydispersity and zeta potential by dynamic light scattering. Bone marrow cells obtained from male BALB/c mice (6-8 weeks) were differentiated into bone marrow-derived macrophages (bmMΦs) by culturing them GM-CSF+M-CSF respectively. In vitro cell cytotoxicity and cell uptake were studied F4/80+ bmMΦs by Alamar Blue assay and flow cytometry. Finally, liposomes were radiolabelled with 3H-Cholesterol and the biodistribution was studied in mice upon intramuscular injection.
Results: DOPE:DOTAP, DOPE:DC-Chol and DOPE:DDA liposomes had a hydrodynamic size below 100 nm and low PDI (<0.25). In vitro cytotoxicity was not observed in any of the three formulations showed (>90% cell survival) at concentrations up to 10 µg/mL liposomes. Importantly, macrophage viability was almost 100% after 24 h with no difference among formulations. Regarding the in vivo biodistribution, a high percentage of the injected dose (%ID) remained at the site of injection as previously reported for cationic liposomes . More specifically, the %ID of DOPE:DC-Chol was slightly higher (10 – 30%) than that of DOPE:DDA and DOPE:DOTAP liposomes. The movement of these liposomes from the injection site translated into higher doses (2 – 4-fold) of the DDA and DOTAP liposomes draining to the popliteal (PLN) and inguinal lymph nodes (ILN).
Conclusion: Previous studies have shown that the physico-chemical attributes of liposomes, such as size, surface charge, lipid choice, hydrophobicity/hydrophilicity and degree of PEGylation have a great impact on the biodistribution profile of liposome-based delivery systems. More importantly, these studies have shown the impact of biodistribution on vaccine efficacy. From this point of view, we have shown that cationic lipid selection can have a notable impact on the pharmacokinetic profile of liposomal vaccine adjuvants