Study of Ethinyl Estradiol Activity Against Promastigotes, Axenic and Macrophage-Dwelling Amastigotes of Leishmania infantum by Using Atomic Force Microscopy and Methyl Thiazolyl Tetrazolium Methods

In the era of drug resistant organisms, there exists a huge need to overcome this issue. Drug repositioning is the action of repurposing current drugs against alternative targets. Herein weas sessed the antileishmanial activity of ethinyl estradiolandtolmetinonthe promastigotes and amastigotes of Leishmania infantum, using methyl thiazolyl tetrazolium (MTT) and atomic force microscopy (AFM) methods. Following previous in silico evaluation of 3358 FDA-approved compounds, ethinyl estradiol and tolmetin drugs were selected. Promastigote assay was done in RPMI 1640 culture in 96-well plates (105/100L). Also, axenic amastigotes cultured in acidic RPMI 1640 were seeded in 96-well plates (5  104 parasites). Raw 264.7 macrophages were seeded at a density of 3  106 cells/well. Then, the macrophages were infected with metacyclic promastigotes of L. infantum. Also, drug solvent and amphotericin B were considered as control wells. Drug challenge was performed in triplicate with different concentrations of ethinyl estradiol (250, 125, 62.5, 31.25, 15.62 and 7.81 g/mL) and tolmetin (200, 100, 50, 25, 12.5 and 6.25 g/mL) and cell viability was assessed by MTT assay. AFM imaging was done to observe morphological changes. The main finding of our investigation was the substantial effect of ethinyl estradiol on promastigotes, and amastigotes of L. infantum during 24, 48 and 72 h, according toMTT results and cellular morphology observations. Thus, the IC50 of this drug on promastigote, axenic amastigotes and macrophage-dwelling amastigotes during 72 h was 23, 45 and 32 g/mL, respectively. However, tolmetin was not effective against L. infantum parasites. Cellular alteration, was observed by AFM technique. This study showed that ethinyl estradiol had good anti-leishmanial activity and it is recommended to test this drug under in vivo condition. With the aid of drug repurposing we could substitute the old drugs with novel compounds for a specific disease.