application): This is an application about further pursuing the use of inhibitors of cysteine proteases for the arrest of replication and attenuation of in vivo infectivity of the intracellular parasites Trypanosoma cruzi and Leishmania. In the previous TDRU funding period, these investigators showed that treatment with vinyl sulfone-derivatized peptidomimetics can rescue mice from a lethal T. cruzi infection, and in some cases eliminate any detectable parasites, achieving parasitological cure. A series of additional synthetic inhibitors, through collaborations with several members of the present multiproject application, were evaluated and found to be effective in the nM and low microM range against the T. cruzi cysteine protease cruzain. Several studies were performed to analyze the metabolites of the inhibitors generated in vivo, and a T. cruzi strain resistant to protease inhibitors was produced in culture. In addition, the mechanism of T. cruzi growth arrest by cysteine protease inhibitors was investigated, and concluded to be related to the accumulation of unprocessed cruzain in the parasite's Golgi apparatus and ER. The present application will be focused on screening additional compounds for enhanced efficacy and specificity against recombinant cruzain, and then examining activity of the most effective inhibitors on parasite replication and differentiation, using culture assays involving the three life cycle stages of T. cruzi. Ultrastructural studies to analyze the cellular effect of the inhibitors are also proposed. The best inhibitor leads identified in these previous steps will be subsequently evaluated in infectivity assays in mice. Essentially the same studies are proposed for Leishmania major, L. donovani and L. mexicana, but in this case the targeting will be on cathepsin B and L homologues identified in L. major. Since deletion mutants in some members of the cysteine protease family are available in L. mexicana, collaborative studies will be performed with this species to investigate the possible differential targeting of inhibitors to different gene family members.
| Lee, Gregory M; Balouch, Eaman; Goetz, David H et al. (2012) Mapping inhibitor binding modes on an active cysteine protease via nuclear magnetic resonance spectroscopy. Biochemistry 51:10087-98 |
| Doyle, Patricia S; Zhou, Yuan M; Hsieh, Ivy et al. (2011) The Trypanosoma cruzi protease cruzain mediates immune evasion. PLoS Pathog 7:e1002139 |
| Boyom, Fabrice Fekam; Fokou, Patrick Valere Tsouh; Yamthe, Lauve Rachel Tchokouaha et al. (2011) Potent antiplasmodial extracts from Cameroonian Annonaceae. J Ethnopharmacol 134:717-24 |
| Swenerton, Ryan K; Zhang, Shuyi; Sajid, Mohammed et al. (2011) The oligopeptidase B of Leishmania regulates parasite enolase and immune evasion. J Biol Chem 286:429-40 |
| Robertson, Stephanie A; Renslo, Adam R (2011) Drug discovery for neglected tropical diseases at the Sandler Center. Future Med Chem 3:1279-88 |
| Huang, Niu; Jacobson, Matthew P (2010) Binding-site assessment by virtual fragment screening. PLoS One 5:e10109 |
| Chen, Yen Ting; Brinen, Linda S; Kerr, Iain D et al. (2010) In vitro and in vivo studies of the trypanocidal properties of WRR-483 against Trypanosoma cruzi. PLoS Negl Trop Dis 4: |
| Chen, Chiung-Kuang; Leung, Siegfried S F; Guilbert, Christophe et al. (2010) Structural characterization of CYP51 from Trypanosoma cruzi and Trypanosoma brucei bound to the antifungal drugs posaconazole and fluconazole. PLoS Negl Trop Dis 4:e651 |
| Brak, Katrien; Kerr, Iain D; Barrett, Kimberly T et al. (2010) Nonpeptidic tetrafluorophenoxymethyl ketone cruzain inhibitors as promising new leads for Chagas disease chemotherapy. J Med Chem 53:1763-73 |
| Guiguemde, W Armand; Shelat, Anang A; Bouck, David et al. (2010) Chemical genetics of Plasmodium falciparum. Nature 465:311-5 |
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