Approved drugs successfully repurposed against based on machine learning predictions.

Drug repurposing is a promising approach towards the discovery of novel treatments against Neglected Tropical Diseases, such as Leishmaniases, presenting the advantage of reducing both costs and duration of the drug discovery process. In previous work, our group developed a Machine Learning pipeline for the repurposing of FDA-approved drugs against parasites. The present study is focused on an validation of this approach by assessing the antileishmanial effects of 10 predicted drug candidates. First, we evaluated the drugs' activity against promastigotes from two strains of and one of , which caused distinct clinical manifestations, using an MTT assay. The standard anti- drug Amphotericin B was used as a positive control. Five molecules demonstrated anti- effects, out of which Acebutolol, Prilocaine and Phenylephrine are described herein for the first time. When tested on promastigote growth, Acebutolol displayed IC values ranging from 69.28 to 145.53 µg/mL. Prilocaine exhibited IC values between 33.10 and 45.81 µg/mL. Phenylephrine, on the other hand, presented IC values >200 µg/mL. The two remaining drugs, Dibucaine and Domperidone, exhibited significantly low IC values varying between 0.58 and 1.05 µg/mL, and 6.30 and 8.17 µg/mL, respectively. Both compounds were previously described as anti- agents . All five compounds demonstrated no notable cytotoxic effects on THP-1-derived macrophages at the IC concentrations, allowing for their testing on the intracellular form of and parasites. Interestingly, all compounds exhibited antileishmanial activity on amastigotes with enhanced IC values compared to the corresponding promastigotes. Noticeably, Dibucaine and Domperidone displayed IC values of at most 1.99 µg/mL. Acebutolol, Prilocaine and Phenylephrine showed IC values ranging from 13.84 to 66.81 µg/mL. Our previously published Computer-Aided repositioning pipelines of FDA-approved drugs as antileishmanial agents identified Dibucaine and Domperidone as candidates in support of previous studies. This study consolidates such findings through the validation against 2 species, highly prevalent in Africa and Middle East, and reveals Acebutolol, Prilocaine, and Phenylephrine as novel anti- effectors, confirming the relevance of our approach and calling for further investigations.