Planejamento, síntese e avaliação farmacológica de novos candidatos a protótipos de fármacos anti-leishmaniose

Abstract

Leishmaniasis is a neglected tropical disease present in more than 90 countries and caused by different species of the genus Leishmania. In particular, L. amazonensis is associated with cutaneous forms, including severe diffuse manifestations. Current treatment involves drugs such as pentavalent antimonials, amphotericin B, and miltefosine (MIL), but these present limitations such as toxicity, high cost, and increasing resistance. MIL, a synthetic phospholipid with surfactant properties, affects the parasite's plasma membrane dynamics through osmotic stress, linking ionic leakage to the production of reactive oxygen species (ROS) and membrane rigidity. Similar effects have been observed for zwitterionic analog compounds, such as N-hexadecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate (HPS), which reinforces the interest in developing new analogs with potential anti-leishmanial activity. In this context, this study aimed to plan and synthesize novel anti-leishmaniasis drug prototype candidates structurally derived from HPS and 2,6-di-tert-butylphenol (BHT), as well as to evaluate their activity against L. amazonensis and investigate their mechanisms of action through Electron Paramagnetic Resonance (EPR) spectroscopy. Compound (5) was obtained through a Mannich reaction between BHT, formaldehyde, and dimethylamine in an acidic medium. In the following step, compound (5) was reacted with 1,3-propanesultone in acetonitrile, resulting in the final compound (3). The antiproliferative activity against L. amazonensis promastigotes was using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide colorimetric assay, while mechanistic studies were performed on stationary-phase promastigotes treated with MIL and compounds (3) and (5), labeled with the 5-DSA spin probe and analyzed by EPR to assess alterations in plasma membrane molecular dynamics. Compounds (3) and (5) were obtained through two synthetic steps, and their structures were elucidated by Nuclear Magnetic Resonance spectroscopy. Antiproliferative assays demonstrated that although compounds (3) and (5) showed inhibitory effects on L. amazonensis promastigotes, their IC50 values were higher than those of MIL, indicating lower in vitro potency. Conversely, EPR spectra showed that MIL and compounds (3) and (5) promoted an increase in the 2A// parameter, suggesting a mechanism of action associated with plasma membrane interaction, fluctuations in cellular and mitochondrial electrical potentials, ROS generation, and membrane rigidity resulting from oxidative stress. It is concluded that the planning and synthesis of new structural derivatives of compounds (3) and (5) with increased lipophilicity represent a promising strategy to enhance interaction with the lipid bilayer, contributing to the development of more potent and selective anti-leishmanial drug prototypes.