Indirubin derivatives are potent and selective anti-<i>Trypanosoma cruzi</i> agents

Current treatment for combatting Chagas disease, a life-threatening illness caused by the kinetoplastid protozoan parasite <i>Trypanosoma cruzi</i> is inadequate, and thus the discovery of new antiparasitic compounds is of prime importance. Previous studies identified the indirubins, a class of ATP kinase inhibitors, as potent growth inhibitors of the related kinetoplastid <i>Leishmania</i>. Herein, we evaluated the inhibitory activity of a series of 69 indirubin analogues screened against <i>T. cruzi</i> trypomastigotes and intracellular amastigotes. Seven indirubins were identified as potent <i>T. cruzi</i> inhibitors (low μΜ, nM range). Cell death analysis of specific compounds [3'oxime-6-bromoindirubin(6-BIO) analogues 10, 11 and 17, bearing a bulky extension on the oxime moiety and one 7 substituted analogue 32], as evaluated by electron microscopy and flow cytometry, showed a different mode of action between compound 32 compared to the three 6-BIO oxime- substituted indirubins, suggesting that indirubins may kill the parasite by different mechanisms dependent on their substitution. Moreover, the efficacy of four compounds that show the most potent anti-parasitic effect in both trypomastigotes and intracellular amastigotes (10, 11, 17, 32), was evaluated in a mouse model of <i>T. cruzi</i> infection. Compound 11 (3ʹpiperazine-6-BIO) displayed the best <i>in vivo</i> efficacy (1/6 mortality, 94.5% blood parasitaemia reduction, 12 dpi) at a dose five times reduced over the reference drug benznidazole (20 mg/kg vs100 mg/kg). We propose 3ʹpiperazine-6-BIO as a potential lead for the development of new treatments of Chagas disease.