Melanosomal targeting via caveolin-1 dependent endocytosis mediates ZN(II) phthalocyanine phototoxic action in melanoma cells.

Melanosomes have been considered crucial targets in melanoma treatments. In this study we explored the role of melanosomes in photodynamic therapy (PDT), employing the synthetic Zn(II) phthalocyanine Pc13, a potent photosensitizer that promotes melanoma cell death after irradiation. Phototoxic action is mediated by reactive oxygen species increase. The internalization mechanism of Pc13 and its consequent subcellular localization were evaluated in melanotic B16-F0 cells. Pharmacological inhibitors of dynamin or caveolae, but not of clathrin, decreased Pc13 cellular uptake and phototoxicity. Similar results were obtained when cells over-expressed dominant negative mutants of dynamin-2 and caveolin-1, indicating that Pc13 is internalized by caveolae-mediated endocytosis. Confocal microscopy analysis revealed that Pc13 targets melanosomes and damage of these structures after irradiation was demonstrated by transmission electron microscopy. Treatment of pigmented B16-F0 and WM35 melanoma cells with the melanin synthesis inhibitor phenylthiourea for 48 h led to cell depigmentation and enhanced cell death after irradiation, whereas a 3-h period of inhibition did not modify melanin content but produced a marked reduction of Pc13 phototoxicity, together with a decrease of oxidative melanin synthesis intermediates. In contrast, the effect of Pc13 in amelanotic A375 cells was not altered by phenylthiourea treatment. These results provide evidence that melanosomes have a dual role in the efficacy of PDT. While melanin antagonizes the phototoxic action of Pc13, the release of cytotoxic synthetic intermediates to cytosol after irradiation and melanosome damage is conducive to the phototoxic response. Based on these findings, we demonstrate that melanosome-targeted PDT could be an effective approach for melanoma treatment.

Action of Metarhizium brunneum (Hypocreales: Clavicipitaceae) Against Organophosphate- and Pyrethroid-Resistant Aedes aegypti (Diptera: Culicidae) and the Synergistic Effects of Phenylthiourea.

Dengue, yellow fever, Zika, and chikungunya arboviruses are endemic in tropical countries and are transmitted by Aedes aegypti. Resistant populations of this mosquito against chemical insecticides are spreading worldwide. This study aimed to evaluate the biological effects of exposure of pesticide-sensitive Ae. aegypti larvae (Rockefeller) to conidia of the entomopathogen, Metarhizium brunneum, laboratory strains ARSEF 4556 and V275, and any synergistic activity of phenylthiourea (PTU). In addition, to investigate the nature of any cross-resistance mechanisms, these M. brunneum strains were tested against the Rockefeller larvae and two temephos- and deltamethrin-resistant wild mosquito populations from Rio de Janeiro. Treatment of Rockefeller larvae with 106 conidia/ml of ARSEF 4556 and V275 fungal strains resulted in significant decreased survival rates to 40 and 53.33%, respectively (P < 0.0001), compared with untreated controls. In contrast, exposure to 104 or 105 conidia/ml showed no such significant survival differences. However, the addition of PTU to the conidia in the bioassays significantly increased mortalities in all groups and induced a molt block. Experiments also showed no differences in Ae. aegypti mortalities between the fungal treated, wild pesticide-resistant populations and the Rockefeller sensitive strain. The results show the efficacy of M. brunneum in controlling Ae. aegypti larvae and the synergistic role of PTU in this process. Importantly, there was no indication of any cross-resistance mechanisms between Ae. aegypti sensitive or resistant to pesticides following treatment with the fungi. These results further support using M. brunneum as an alternative biological control agent against mosquito populations resistant to chemical insecticides.

Recycling and refurbishing old antitubercular drugs: the encouraging case of inhibitors of mycolic acid biosynthesis.

One of the first approaches undertaken in the quest for antitubercular compounds was that of understanding the mechanism of action of old drugs and proposing chemical modifications or other strategies to improve their activity, generally lost to the mechanisms of resistance developed by Mycobacterium tuberculosis. A leading case was the work carried out on a set of compounds with proven activity on the essential pathway of the synthesis of mycolic acids. As a result, different solutions were presented, improving the activity of those inhibitors or producing novel compounds acting on the same molecular target(s), but avoiding the most common resistance strategies developed by the tubercle bacilli. This review focuses on the activity of those compounds, developed following the completion of the studies on several of the classic antitubercular drugs.