Research Article: Identification of Three Classes of Heteroaromatic Compounds with Activity against Intracellular Trypanosoma cruzi by Chemical Library Screening

Date Published: February 24, 2009

Publisher: Public Library of Science

Author(s): Esther Bettiol, Marie Samanovic, Andrew S. Murkin, Jayne Raper, Frederick Buckner, Ana Rodriguez, James H. McKerrow

Abstract: The development of new drugs against Chagas disease is a priority since the currently available medicines have toxic effects, partial efficacy and are targeted against the acute phase of disease. At present, there is no drug to treat the chronic stage. In this study, we have optimized a whole cell-based assay for high throughput screening of compounds that inhibit infection of mammalian cells by Trypanosoma cruzi trypomastigotes. A 2000-compound chemical library was screened using a recombinant T. cruzi (Tulahuen strain) expressing β-galactosidase. Three hits were selected for their high activity against T. cruzi and low toxicity to host cells in vitro: PCH1, NT1 and CX1 (IC50: 54, 190 and 23 nM, respectively). Each of these three compounds presents a different mechanism of action on intracellular proliferation of T. cruzi amastigotes. CX1 shows strong trypanocidal activity, an essential characteristic for the development of drugs against the chronic stage of Chagas disease where parasites are found intracellular in a quiescent stage. NT1 has a trypanostatic effect, while PCH1 affects parasite division. The three compounds also show high activity against intracellular T. cruzi from the Y strain and against the related kinetoplastid species Leishmania major and L. amazonensis. Characterization of the anti–T. cruzi activity of molecules chemically related to the three library hits allowed the selection of two compounds with IC50 values of 2 nM (PCH6 and CX2). These values are approximately 100 times lower than those of the medicines used in patients against T. cruzi. These results provide new candidate molecules for the development of treatments against Chagas disease and leishmaniasis.

Partial Text: Chagas disease or American trypanosomiasis is a devastating disease caused by the trypanosomatid protozoan Trypanosoma cruzi. It is endemic in 18 countries of Central and South America, putting 120 million of people at risk, with an estimated 16–18 million people currently infected [1]. The disease first manifests itself with an acute phase involving symptoms of swelling near the infection site, fever, fatigue, and enlarged lymphatic organs. It can then remain asymptomatic or manifest itself in a chronic form leading to cardiac insufficiency and megacolon. The two available drugs used to fight T. cruzi parasites during the acute stage are benznidazole (BZN) (Rochagan, Hoffmann-LaRoche) and nifurtimox (Lampit, Bayer). These drugs have toxic side effects and are not always effective. There is no drug available to treat the chronic stage of Chagas disease. Though some studies suggest that treatment with either BZN or nifurtimox decreases parasite load and slows disease progression, treatment of the chronic stage with these compounds is not officially recommended [2].

New compounds are desperately needed to fight efficiently T. cruzi, the parasite responsible for Chagas disease. To this aim, we optimized a simple and straight-forward assay that allows the HTS of compounds against T. cruzi replicating within mammalian cells. We tested 2000 compounds from the DIVERSet chemical library. This compound library has been useful to discover inhibitors of matrix metalloproteinase-9 in a whole cell assay [20] and to find inhibitors of the ribonucleic activity of angiogenin using a cell-free system [21]. However, to our knowledge this is the first time this library has been used to discover new anti-microbials.



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