Multi-target withaferin-A analogues as promising anti-kinetoplastid agents through the programmed cell death

Leishmaniasis and Chagas disease, two of the most prevalent neglected tropical diseases, are a world health problem. The harsh reality of these infective diseases is the absence of effective and safe therapies. In this framework, natural products play an important role in overcoming the current need to development new antiparasitic agents. The present study reports the synthesis, antikinetoplastid screening, mechanism study of fourteen withaferin A derivatives (2-15). Nine of them (2-6, 8-10 and 12) showed a potent dose-dependent inhibitory effect on the proliferation of Leishmania amazonensis and L. donovani promastigotes and Trypanosoma cruzi epimastigotes with IC₅₀ values ranging from 0.19 to 24.01 µM. Outstandingly, the fully acetylated derivative 10 (4,27-diacetylwithaferin A) was the most potent compound showing IC₅₀ values of 0.36, 2.82 and 0.19 µM against L. amazonensis, L. donovani and T. cruzi, respectively. Furthermore, analogue 10 exhibited approximately 18 and 36-fold greater antikinetoplastid activity, on L. amazonensis and T. cruzi, than the reference drugs. The activity was accompanied by significantly lower cytotoxicity on the murine macrophage cell line. Moreover, compounds 2, 3, 5-7, 9 and 10 showed more potent activity than the reference drug against the intracellular amastigotes forms of L. amazonensis and T.cruzi, with a good selectivity index on a mammalian cell line. In addition, withaferin A analogues 3, 5-7, 9 and 10 induce programmed cell death through a process of apoptosis-like and autophagy. These results strengthen the anti-parasitic potential of withaferin A-related steroids against neglected tropical diseases caused by Leishmania spp. and T. cruzi parasites.