Microemulsions Improve Topical Protoporphyrin IX (PpIX) Delivery for Photodynamic Therapy of Skin Cancer

Abstract We report here microemulsions (MEs) for topical delivery of protoporphyrin IX (PpIX) for Photodynamic Therapy (PDT) of skin cancers. Selected MEs consisting of Oil/Water (O/W) bicontinuous (BC) and Water/Oil (W/O) preparations were characterized as to pH, nanometric size, zeta potential, drug content, and viscosity. Sustained in vitro PpIX release was achieved from MEs 2A (O/W), 10B (BC) and 16B (W/O) through an artificial membrane for up to 24 h, characterizing MEs as drug delivery systems. None of these MEs showed permeation through the skin, demonstrating the required topical effect. After 4 h, in vitro retention of PpIX in the stratum corneum (SC) was higher from both ME 10B and control (PpIX at 60 µg/mL in PEG 300). However, in the Epidermis + Dermis ([Ep + D]), retention from ME 10B and ME 16B was ~40 times higher compared to control. Confocal Laser Scanning Microscopy (CLSM) showed higher fluorescence intensity in the SC for both control and ME 10B, whereas ME 10B fluorescence was higher in [Ep+D]. The results indicate that ME 10B is suitable for PpIX encapsulation, showing good characteristics and a localized effect for a potential delivery system for PDT-associated treatments of skin cancers.

5-Aminolaevulinic acid-mediated photodynamic therapy in multidrug resistant leukemia cells.

To verify if photodynamic therapy (PDT) could overcome multidrug resistance (MDR) when it it applied to eradicate minimal residual disease in patients with leukemia, we investigated the fluorescence kinetics of 5-aminolaevulinic acid (ALA)-induced protoporphyrin IX (PpIX) and the effect of subsequent photodynamic therapy on MDR leukemia cells, which express P-glycoprotein (P-gp), as well as on their parent cells. Evaluation of PpIX accumulation by flow cytometry showed that PpIX accumulated at higher levels in mdr-1 gene-transduced MDR cells (NB4/MDR) and at lower levels in doxorubicin-induced MDR cells (NOMO-1/ADR) than in their parent cells. A P-gp inhibitor could not increase PpIX accumulation. Measurement of extracellular PpIX concentration by fluorescence spectrometry showed that P-gp did not mediate the fluorescence kinetics of ALA-induced PpIX production. Assessment of ferrochelatase activity using high-performance liquid chromatography indicated that PpIX accumulation in drug-induced MDR cells was probably regulated by this enzyme. Assessment of phototoxicity of PDT using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that PDT was effective in NB4, NB4/MDR, NOMO-1 and NOMO-1/ADR cells, which accumulated high levels of PpIX, but not effective in K562 and K562/ADR cell lines, which accumulated relatively low levels of PpIX. These findings demonstrate that P-gp does not mediate the ALA-fluorescence kinetics, and multidrug resistant leukemia cells do not have cross-resistance to ALA-PDT.