Development of Therapeutics for the Treatment of Leishmaniasis and Chagas Disease

Abstract

Neglected tropical diseases (NTDs) are an increasing global health challenge exacerbated by a shortfall in funding and new developments in the field. The social and economic repercussions of these diseases can be as overwhelming as their health effects. Current frontline treatments for leishmaniasis and chagas disease suffer from range of issues such as, high cost, prolonged treatments, severe side effects and drug resistance. Hence, there is a need for new drugs. A library of molecules with highly diverse chemical space was screened against Leishmania mexicana parasites. Four of the derivatives from Class One and Two, 40, 46, 60 and 64 showed EC50 values of less than 10 µM against L. mexicana intracellular amastigotes (3.12, 7.37, 4.86 and 3.27 µM respectively) and 9,10-phenanthrenedione (Class One) and 1,10-phenanthroline (Class Two) were identified as novel chemical scaffolds which could be developed as anti-leishmanial therapeutic agents. The repurposing ability of three FDA approved drugs; pyrithione (104), ciclopirox (105) and piroctone olamine (106) as a treatment for cutaneous leishmaniasis was investigated. Having EC50 values lower than 2 µM against L. mexicana intracellular amastigotes and selectivity index (SI) values greater than 10 proved that all three drugs have the potential to be repurposed. The promising anti-leishmanial activities reported for the four novel derivatives (116-119) synthesized from pyrithione (104) showed that the activity is not due to the metal chelation and modification of the N-oxide functionality of pyrithione (104) did not significantly change its biological activity. Furthermore, it was evident from in-gel fluorescence assays that pyrithione (104) and the novel derivatives (116-119) inhibit DUB16 enzyme, an enzyme essential for the viability of procyclic promastigotes, in L. mexicana promastigotes. However, DUB16 overexpression did not provide any resistance towards antileishmanial activity, meaning that these inhibitors may have other targets beyond this enzyme. Two fluorinated compound libraries in were screened against L. mexicana and Trypanosoma cruzi parasites. From these libraries 4-(benzenesulfonyl)-2,3,5,6-tetrafluoropyridine (198) was the only molecule found to be active against both the parasite species [EC50 (L. mexicana promastigotes); 1.33 µM, EC50 (L. mexicana axenic amastigotes); 0.433 µM, IC50 (T. cruzi epimastigotes); 1.55 µM, EC50 (T. cruzi infected stage); 0.05 µM]. Subsequently, the effect of 198 on programmed cell death in T. cruzi epimastigotes was investigated and results indicated that 198 triggers plasma membrane permeabilization and alters the T. cruzi epimastigotes mitochondrial function by collapsing mitochondrial potential, decreasing the intracellular Ca2+ concentration and disrupting the intracellular ATP levels. Additionally, it was discovered that 198 also arrest the G0 phase of cell cycle at higher concentrations. This suggests that 198 could be an interesting lead compound in Chagas disease drug discovery and development. A second-generation library derived from 198 was synthesised and their anti-parasitic activities were evaluated against L. mexicana axenic amastigotes. Finally, a label free MS-based proteomics analysis was performed to identify the target proteins of 230 in L. mexicana promastigotes and 19 proteins were identified as potential target proteins of 230.