We investigated a chemical strategy to boost the trypanocidal activity of 2,4dihydroxybenzoic acid (2,4-DHBA)-and salicylhydroxamic acid (SHAM)-based trypanocides with triphenylphosphonium and quinolinium lipophilic cations (LC). Three series of LC conjugates were synthesized that were active in the submicromolar (5a-d and 10d-f) to low nanomolar (6a-f) range against wild-type and multi-drug resistant strains of African trypanosomes (Trypanosoma brucei brucei and T. congolense). This represented an improvement in trypanocidal potency of at least 200fold, and up to >10,000-fold, compared with the non-LC coupled parent compounds 2,4-DHBA and SHAM. Selectivity over human cells was >500 and reached >23,000 for 6e. Mechanistic studies showed that 6e did not inhibit the cell cycle but affected parasite respiration in a dose-dependent manner. Inhibition of the trypanosome alternative oxidase (TAO) and the mitochondrial membrane potential was also studied for selected compounds. We conclude that effective mitochondrial targeting greatly potentiated the activity of these compound series.

Abstract:In Trypanosoma brucei, a parasite that causes African trypanosomiasis in humans (sleeping sickness) and in livestock (nagana) throughout sub-Saharan Africa, the trypanosome alternative oxidase (TAO) is essential for the respiration of bloodstream form parasites (i.e. the human-infective form). Since TAO has no counterpart in mammalian cells and it is conserved among T. brucei subspecies, it has been validated as a promising target for the chemotherapy of African trypanosomiasis.We present here a successful approach to boost the activity of TAO inhibitors based on the conjugation of the inhibitor with lipophilic cations (LC) that can cross lipid bilayers by non-carrier mediated transport, and thus accumulate specifically into mitochondria, driven by the plasma and mitochondrial transmembrane potentials (negative inside). This design afforded several LC-TAO inhibitor conjugates active in the submicromolar to low nanomolar range against wild type and resistant strains of African trypanosomes (T. b. brucei, T. congolense), with selectivity over human cells >500.