AUTHOR=Magalhães Tatiana Barbosa dos Santos , Silva Dahara Keyse Carvalho , Teixeira Jessica da Silva , De Lima Juliana Dizaira Teles , Barbosa-Filho José Maria , Moreira Diogo Rodrigo Magalhães , Guimarães Elisalva Teixeira , Soares Milena Botelho Pereira 

TITLE=A Betulinic Acid Derivative, BA5, Induces G0/G1 Cell Arrest, Apoptosis Like-Death, and Morphological Alterations in Leishmania sp

JOURNAL=Frontiers in Pharmacology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/pharmacology/articles/10.3389/fphar.2022.846123

DOI=10.3389/fphar.2022.846123

ISSN=1663-9812

ABSTRACT=<p>Leishmaniasis are endemic diseases caused by different species of intracellular parasites of the genus <italic>Leishmania</italic>. Due to the high toxicity and drug resistance of current antileishmanial drugs, it is necessary to identify new and more effective drugs. Previously, we investigated the immunomodulatory and anti-<italic>Trypanosoma cruzi</italic> action of BA5, a derivative of betulinic acid. In the present study, we investigated the <italic>in vitro</italic> activity of BA5 against different species of <italic>Leishmania</italic> and their action mechanism. BA5 exhibited low cytotoxicity against macrophages and inhibited the proliferation of promastigote forms of <italic>Leishmania amazonensis</italic> (IC<sub>50</sub> = 4.5 ± 1.1 μM), <italic>Leishmania major</italic> (IC<sub>50</sub> = 3.0 ± 0.8 μM), <italic>Leishmania braziliensis</italic> (IC<sub>50</sub> = 0.9 ± 1.1 μM) and <italic>Leishmania infantum</italic> (IC<sub>50</sub> = 0.15 ± 0.05 μM). Incubation with BA5 reduced the percentage of <italic>Leishmania amazonensis</italic>-infected macrophages and the number of intracellular parasites (IC<sub>50</sub> = 4.1 ± 0.7 μM). To understand the mechanism of action underlying BA5 antileishmanial activity (incubation at IC<sub>50</sub>/2<sub>,</sub> IC<sub>50</sub> or 2xIC<sub>50</sub> values of the drug), we investigated ultrastructural changes by scanning electron microscopy and evaluated cell cycle, membrane mitochondrial potential, and cell death against promastigote forms of <italic>Leishmania amazonensis</italic> by flow cytometry. Promastigotes incubated with BA5 presented membrane blebbing, flagella damage, increased size, and body deformation. Flow cytometry analysis showed that parasite death is mainly caused by apoptosis-like death, arrested cell cycle in G0/G1 phase and did not alter the membrane mitochondrial potential of <italic>Leishmania amazonensis</italic>. Surprisingly, the combination of BA5 and amphotericin B, an assay used to determine the degree of drug interaction, revealed synergistic effects (CI = 0.15 ± 0.09) on promastigotes forms of <italic>Leishmania amazonensis</italic>. In conclusion, BA5 compound is an effective and selective antileishmanial agent.</p>