Antileishmanial Evaluation and Mechanism of Action of Benzothiazole Derivatives with Aromatic/Heteroaromatic Hydrazone Moiety

Introduction: Treatment of leishmaniasis is limited to a few drugs, with serious side effects and contraindications. Thus, the need for alternatives to treat this neglected infectious disease is undeniable. Methods: In this work, a series of benzothiazole derivatives was synthesized, and their in vitro effects against promastigotes and intracellular amastigotes of Leishmania spp. and their macrophage toxicity were evaluated. Peritoneal macrophages were successful obtained from BALB/c mice were used for intracellular amastigotes of Leishmania spp. assays and to assess their toxicity on mammalian cells. In vitro studies on the mechanism of action and drug combination assays were also performed. Results: Compound 2a exhibited remarkable activity against L. amazonensis and L. infantum, with IC50 values below 5 μM against amastigote forms (3.96 μM and 4.0 μM, respectively), surpassing the reference drug miltefosine, and had no cytotoxic effect on macrophages (CC50 > 150 μM). Discussion: The antileishmanial effect of compound 2a was associated with hyperpolarization of the mitochondrial membrane potential, increased ROS levels, and the formation of lipid bodies in treated promastigote forms of L. amazonensis, indicating mitochondrial dysfunction and oxidative stress. No disruption of the promastigotes' plasma membrane was observed after treatment with this compound, ruling out necrotic cell death. Interactions between compound 2a and miltefosine exhibited a better effect at the lowest concentration of compound 2a (combination 1:4) in both promastigote and amastigote forms of L. amazonensis. In silico analysis showed promising physicochemical properties for compound 2a. Conclusion: Compound 2a displays strong antileishmanial activity against L. amazonensis and L. infantum, demonstrating a broad ability to inhibit the growth of Leishmania species associated with cutaneous and visceral manifestations.