Leishmanioses are zoonotic diseases caused by intracellular protozoans of the genus Leishmania. Recent research has revealed the extensive distribution and expansion of canine leishmaniosis in large areas of the world, where the high prevalence of canine infection is associated with an increased risk of human disease. There are not specific pharmacologic treatments for canine leishmaniasis. The only way to manage the situation is the euthanasia of the infected dogs. The sacrifice of the dog was used to try to control the expansion of the infection since decades without success. Also there is a lot of other animal species that can act as host for the disease, also with human contact. Then, to achieve a solution, we must develop a vaccine or a specific drug for canine leishmaniasis. The Pathogen Box is a project led by Medicines for Malaria Venture (MMV, Switzerland; http://www.pathogenbox.org/) that aims to identify novel drugs with activity against diseases such as tuberculosis, malaria, toxoplasmosis, and dengue, among others. The box consists of 400 mostly novel synthetic chemicals that were initially selected from a set of ~4 million compounds due to their low toxicity for mammalian cells and activity against specific microbial pathogens. In fact, the compounds display cytotoxicity at levels that are thought to be reasonable for drug discovery programs. In this study, we screened the Pathogen Box compounds for antiparasitic activity against Leishmania infantum (reference strain and clinical isolates). This screen led to the discovery of 5 novel hits for drug development and drug design.