Amphotericin B (AmB) is a high-molecular weight poorly soluble drug with antiparasitic activity (1, 2), which possesses a very low oral and topical bioavailability (1, 2). Transferosomes (TFs), which are ultradeformable vesicles consisting on lipids, an edge activator and a low amount of ethanol (<10%)(3), could be a suitable formulation to deliver AmB through the skin (4, 5).
TFs were obtained by thin film hydration method. A 3-level Box-Behnken design of experiments was used to optimise the formulation. Optimised AmB-TFs were characterised in terms of particle size, zeta-potential, drug loading and morphology. The permeability of the AmB-TFs was tested in vitro using Franz cells across different membranes (6) but also in vivo in CD-1 mice. The toxicological profile of the formulation was tested by performing histological studies of the mouse skin after a 6 h exposure and haemolytic studies using human RBCs (7). In vitro and in vivo antiparasitic efficacy was also assessed (8-10) in different species of Leishmania spp.
The optimised formulation according to the Box-Behnken design consisted of 14:86 w:w edge activator : lipids and a drug loading of 0.086%. The formulation exhibited a good physicochemical stability for 6 months under desiccated conditions. AmB-TFs illustrated a flux of 41.18 ± 1.39 µg/cm2/h across Strat-M® membrane and the diffusion fitted well the Korsmeyer-Peppas model. Upon histological evaluation minor swelling indicative of inflammation or epithelial hyperplasia were appreciated. Haemolytic studies demonstrated that AmB-TFs possessed a 10-fold higher HC50 compared to equivalent AmB concentrations dissolved in DMSO. AmB-TFs showed excellent in vitro activity against against L. amazonensis and L. brazilensis promastigotes was in the micromolar range with a selectivity index above 5 in both cases. In vivo studies demonstrated a good permeation of the drug after topical application on healthy mouse skin which increased over exposure time and was accumulated within the epidermis and dermis. The amount of AmB recovered from the epidermis was well above the IC50 and likely to be efficacious in eliminating microorganisms form the skin. This was confirmed by in vivo efficacy studies in which parasite load was decreased in a 98.24 ± 1.54%.
AmB-TFs enabled the permeation of AmB after topical administration that allowed therapeutically relevant amounts to be uptaken and accumulated within the dermis where parasites accumulate. The low toxicity of the formulation and minimal if any changes in the skin morphology allows for safe and effective non-invasive formulations for the treatment of these fungal and parasitic infections.
