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Simple dialkyl pyrazole-3,5-dicarboxylates show in vitro and in vivo activity against disease-causing trypanosomatids


The synthesis and antiprotozoal activity of some simple dialkyl pyrazole-3,5-dicarboxylates (compounds 2–6) and their sodium salts (pyrazolates) (compounds 7–9) against Trypanosoma cruzi, Leishmania infantum and Leishmania braziliensis are reported. In most cases the studied compounds showed, especially against the clinically significant amastigote forms, in vitro activities higher than those of the reference drugs (benznidazole for T. cruzi and glucantime for Leishmania spp.); furthermore, the low non-specific cytotoxicities against Vero cells and macrophages shown by these compounds led to good selectivity indexes, which are 8–72 times higher for T. cruzi amastigotes and 15–113 times higher for Leishmania spp. amastigotes than those of the respective reference drugs. The high efficiency of diethyl ester 3 and its sodium salt 8 against the mentioned protozoa was confirmed by further in vitro assays on infection rates and by an additional in vivo study in a murine model of acute and chronic Chagas disease. The inhibitory capacity of compounds 3 and 8 on the essential iron superoxide dismutase of the aforementioned parasites may be related to the observed anti-trypanosomatid activity. The low acute toxicity of compounds 3 and 8 in mice is also reported in this article.

Corresponding author
*Corresponding authors: Instituto de Química Médica, CSIC, c/Juan de la Cierva 3, 28006-Madrid, Spain. E-mail: and Departamento de Parasitología, Instituto de Investigación Biosanitaria (ibs.GRANADA), Hospitales Universitarios de Granada/Universidad de Granada, Granada, Spain. E-mail:
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These authors contributed equally to this work.

Present address: Department of Pathogen Molecular Biology, London School of Hygiene & Tropical Medicine, London, UK.


Present address: Department of Cell Biology, University of Alberta, Edmonton, Alberta, Canada.

Present address: GIBI 230, Instituto de Investigación Sanitaria y Hospital Universitario y Politécnico La Fe, Valencia, Spain.

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Askew, B. C., Bednar, R. A., Bednar, B., Claremon, D. A., Cook, J. J., McIntyre, C. J., Hunt, C. A., Gould, R. J., Lynch, R. J., Lynch, J. J. Jr., Gaul, S. L., Stranieri, M. T., Sitko, G. R., Holahan, M. A., Glass, J. D., Hamill, T., Gorham, L. M., Prueksaritanont, T., Baldwin, J. J. and Hartman, G. D. (1997). Non-peptide glycoprotein IIb/IIIa inhibitors. 17. Design and synthesis of orally active, long-acting non-peptide fibrinogen receptor antagonists. Journal of Medicinal Chemistry 40, 17791788.
Bermudez, J., Davies, C., Simonazzi, A., Real, J. P. and Palma, S. (2016). Current drug therapy and pharmaceutical challenges for Chagas disease. Acta Tropica 156, 116.
Berneman, A., Montout, L., Goyard, S., Chamond, N., Cosson, A., d'Archivio, S., Gouault, N., Uriac, P., Blondel, A. and Minoprio, P. (2013). Combined approaches for drug design points the way to novel proline racemase inhibitor candidates to fight Chagas’ disease. PLoS ONE 8, e60955.
Beyer, W. F. Jr. and Fridovich, I. (1987). Assaying for superoxide dismutase activity: some large consequences of minor changes in conditions. Analytical Biochemistry 161, 559566.
Cavalli, A. and Bolognesi, M. L. (2009). Neglected Tropical Diseases: multi-target-directed ligands in the search for novel lead candidates against Trypanosoma and Leishmania . Journal of Medicinal Chemistry 52, 73397359.
Cencig, S., Coltel, N., Truyens, C. and Carlier, Y. (2011). Parasitic loads in tissues of mice infected with Trypanosoma cruzi and treated with AmBisome. PLoS Neglected Tropical Diseases 5, e1216.
Cerecetto, H. and González, M. (2010). Synthetic medicinal chemistry in Chagas’ disease: compounds at the final stage of “hit-to-lead” phase. Pharmaceuticals 3, 810838.
Dujardin, J.-C., González-Pacanowska, D., Croft, S. L., Olesen, O. F. and Späth, G. F. (2010). Collaborative actions in anti-trypanosomatid chemotherapy with partners from disease endemic areas. Trends in Parasitology 26, 395403.
Escartí, F., Miranda, C., Lamarque, L., Latorre, J., García-España, E., Kumar, M., Arán, V. J. and Navarro, P. (2002). Cu2+-induced formation of cage-like compounds containing pyrazole macrocycles. Journal of the Chemical Society, Chemical Communications 936937.
Espuelas, S., Plano, D., Nguewa, P., Font, M., Palop, J. A., Irache, J. M. and Sanmartín, C. (2012). Innovative lead compounds and formulation strategies as newer kinetoplastid therapies. Current Medicinal Chemistry 19, 42594288.
Fonseca-Berzal, C., Ibáñez-Escribano, A., Reviriego, F., Cumella, J., Morales, P., Jagerovic, N., Nogal-Ruiz, J. J., Escario, J. A., da Silva, P. B., Soeiro, M. N. C., Gómez-Barrio, A., Arán, V. J. (2016). Antichagasic and trichomonacidal activity of 1-substituted 2-benzyl-5-nitroindazolin-3-ones and 3-alkoxy-2-benzyl-5-nitro-2H-indazoles. European Journal of Medicinal Chemistry 115, 295310.
González, P., Marín, C., Rodríguez-González, I., Hitos, A. B., Rosales, M. J., Reina, M., Díaz, J. G., González-Coloma, A. and Sánchez-Moreno, M. (2005). In vitro activity of C20-diterpenoid alkaloid derivatives in promastigotes and intracellular amastigotes of Leishmania infantum . International Journal of Antimicrobial Agents 25, 136141.
Guedes, P. M. M., Silva, G. K., Gutierrez, F. R. S. and Silva, J. S. (2011). Current status of Chagas disease chemotherapy. Expert Review of Anti-infective Therapy 9, 609620.
Lamarque, L., Navarro, P., Miranda, C., Arán, V. J., Ochoa, C., Escartí, F., García-España, E., Latorre, J., Luis, S. V. and Miravet, F. (2001). Dopamine interaction in the absence and in the presence of Cu2+ ions with macrocyclic and macrobicyclic polyamines containing pyrazole units. Crystal structures of [Cu2(L1)(H2O)2](ClO4)4 and [Cu2(H−1L3)](ClO4)3·2H2O. Journal of the American Chemical Society 123, 1056010570.
Longoni, S. S., Marín, C., Sauri-Arceo, C. H., López-Cespedes, A., Rodríguez-Vivas, R. I., Villegas, N., Escobedo-Ortegón, J., Barrera-Pérez, M. A., Bolio-Gonzalez, M. E. and Sánchez-Moreno, M. (2011). An iron-superoxide dismutase antigen-based serological screening of dogs indicates their potential role in the transmission of cutaneous leishmaniasis and trypanosomiasis in Yucatan, Mexico. Vector-Borne and Zoonotic Diseases 11, 815821.
Marín, C., Ramírez-Macías, I., López-Céspedes, A., Olmo, F., Villegas, N., Díaz, J. G., Rosales, M. J., Gutiérrez-Sánchez, R. and Sánchez-Moreno, M. (2011). In vitro and in vivo trypanocidal activity of flavonoids from Delphinium staphisagria against Chagas disease. Journal of Natural Products 74, 744750.
Marín, C., Clares, M. P., Ramírez-Macías, I., Blasco, S., Olmo, F., Soriano, C., Verdejo, B., Rosales, M. J., Gomez-Herrera, D., García-España, E. and Sánchez-Moreno, M. (2013). In vitro activity of scorpiand-like azamacrocycle derivative in promastigotes and intracellular amastigotes of Leishmania infantum and Leishmania braziliensis . European Journal of Medicinal Chemistry 62, 466477.
Miranda, C., Escartí, F., Lamarque, L., García-España, E., Navarro, P., Latorre, J., Lloret, F., Jiménez, H. R. and Yunta, M. J. R. (2005). CuII and ZnII coordination chemistry of pyrazole-containing polyamine receptors – influence of the hydrocarbon side chain length on the metal coordination. European Journal of Inorganic Chemistry 2005, 189208.
Mishra, J., Saxena, A. and Singh, S. (2007). Chemotherapy of leishmaniasis: past, present and future. Current Medicinal Chemistry 14, 11531169.
Navarro, P., Sánchez-Moreno, M., Marín, C., García-España, E., Ramírez-Macías, I., Olmo, F., Rosales, M. J., Gómez-Contreras, F., Yunta, M. J. R., Gutiérrez-Sánchez, R. (2014). In vitro leishmanicidal activity of pyrazole-containing polyamine macrocycles which inhibit the Fe-SOD enzyme of Leishmania infantum and Leishmania braziliensis species. Parasitology 141, 10311043.
Olmo, F., Clares, M. P., Marín, C., González, J., Inclán, M., Soriano, C., Urbanová, K., Tejero, R., Rosales, M. J., Krauth-Siegel, R. L., Sánchez-Moreno, M. and García-España, E. (2014 a). Synthetic single and double aza-scorpiand macrocycles acting as inhibitors of the antioxidant enzymes iron superoxide dismutase and trypanothione reductase in Trypanosoma cruzi with promising results in a murine model. RSC Advances 4, 6510865120 (Electronic supplementary information:
Olmo, F., Escobedo-Ortegón, J., Palma, P., Sánchez-Moreno, M., Mejía-Jaramillo, A., Triana, O. and Marín, C. (2014 b). Specific primers design based on the superoxide dismutase b gene for Trypanosoma cruzi as a screening tool: validation method using strains from Colombia classified according to their discrete typing unit. Asian Pacific Journal of Tropical Medicine 7, 854859.
Olmo, F., Rotger, C., Ramírez-Macías, I., Martínez, L., Marín, C., Carreras, L., Urbanová, K., Vega, M., Chaves-Lemaur, G., Sampedro, A., Rosales, M. J., Sánchez-Moreno, M. and Costa, A. (2014 c). Synthesis and biological evaluation of N,N′ -squaramides with high in vivo efficacy and low toxicity: toward a low-cost drug against Chagas disease. Journal of Medicinal Chemistry 57, 987999.
Olmo, F., Guardia, J. J., Marín, C., Messouri, I., Rosales, M. J., Urbanová, K., Chayboun, I., Chahboun, R., Alvarez-Manzaneda, E. J. and Sánchez-Moreno, M. (2015). Prospects of an alternative treatment against Trypanosoma cruzi based on abietic acid derivatives show promising results in Balb/c mouse model. European Journal of Medicinal Chemistry 89, 683690.
Rajasekaran, R. and Chen, Y.-P. P. (2015). Potential therapeutic targets and the role of technology in developing novel antileishmanial drugs. Drug Discovery Today 20, 958968.
Ramírez-Macías, I., Marín, C., Es-Samti, H., Fernández, A., Guardia, J. J., Zentar, H., Agil, A., Chahboun, R., Alvarez-Manzaneda, E. and Sánchez-Moreno, M. (2012). Taiwaniaquinoid and abietane quinone derivatives with trypanocidal activity against T. cruzi and Leishmania spp. Parasitology International 61, 405413.
Reviriego, F., Rodríguez-Franco, M. I., Navarro, P., García-España, E., Liu-González, M., Verdejo, B. and Domènech, A. (2006). The sodium salt of diethyl 1H-pyrazole-3,5-dicarboxylate as an efficient amphiphilic receptor for dopamine and amphetamines. Crystal structure and solution studies. Journal of the American Chemical Society 128, 1645816459.
Sánchez-Moreno, M., Sanz, A. M., Gómez-Contreras, F., Navarro, P., Marín, C., Ramírez-Macias, I., Rosales, M. J., Olmo, F., Garcia-Aranda, I., Campayo, L., Cano, C., Arrebola, F. and Yunta, M. J. R. (2011). In vivo trypanosomicidal activity of imidazole- or pyrazole-based benzo[g]phthalazine derivatives against acute and chronic phases of Chagas disease. Journal of Medicinal Chemistry 54, 970979.
Sánchez-Moreno, M., Gómez-Contreras, F., Navarro, P., Marín, C., Olmo, F., Yunta, M. J. R., Sanz, A. M., Rosales, M. J., Cano, C. and Campayo, L. (2012 a). Phthalazine derivatives containing imidazole rings behave as Fe-SOD inhibitors and show remarkable anti-T. cruzi activity in immunodeficient-mouse model of infection. Journal of Medicinal Chemistry 55, 99009913.
Sánchez-Moreno, M., Gómez-Contreras, F., Navarro, P., Marín, C., Ramírez-Macías, I., Olmo, F., Sanz, A. M., Campayo, L., Cano, C. and Yunta, M. J. R. (2012 b). In vitro leishmanicidal activity of imidazole- or pyrazole-based benzo[g]phthalazine derivatives against Leishmania infantum and Leishmania braziliensis species. Journal of Antimicrobial Chemotherapy 67, 387397.
Sánchez-Moreno, M., Marín, C., Navarro, P., Lamarque, L., García-España, E., Miranda, C., Huertas, O., Olmo, F., Gómez-Contreras, F., Pitarch, J. and Arrebola, F. (2012 c). In vitro and in vivo trypanosomicidal activity of pyrazole-containing macrocyclic and macrobicyclic polyamines: their action on acute and chronic phases of Chagas disease. Journal of Medicinal Chemistry 55, 42314243.
Sánchez-Sancho, F., Campillo, N. E. and Páez, J. A. (2010). Chagas disease: progress and new perspectives. Current Medicinal Chemistry 17, 423452.
Santos, D. O., Coutinho, C. E. R., Madeira, M. F., Bottino, C. G., Vieira, R. T., Nascimento, S. B., Bernardino, A., Bourguignon, S. C., Corte-Real, S., Pinho, R. T., Rodrigues, C. R. and Castro, H. C. (2008). Leishmaniasis treatment – a challenge that remains: a review. Parasitology Research 103, 110.
Schenck, T. G., Downes, J. M., Milne, C. R. C., Mackenzie, P. B., Boucher, H., Whelan, J. and Bosnich, B. (1985). Bimetallic reactivity. Synthesis of bimetallic complexes containing a bis(phosphino)pyrazole ligand. Inorganic Chemistry 24, 23342337.
Sharma, M. K., Arán, V. J., Navarro, P., Ramos-Gallardo, A. and Vegas, A. (1994). Dinuclear Cu(II) complexes with two pyrazolate bridging groups formed from 26 membered oxaimine and polyamine macrocycles of 3,5-disubstituted 1H-pyrazole. Tetrahedron Letters 35, 57235726.
Singh, N., Kumar, M. and Singh, R. K. (2012). Leishmaniasis: current status of available drugs and new potential drug targets. Asian Pacific Journal of Tropical Medicine 5, 485497.
Soeiro, M. N. C. and de Castro, S. L. (2009). Trypanosoma cruzi targets for new chemotherapeutic approaches. Expert Opinion on Therapeutic Targets 13, 105121.
Téllez-Meneses, J., Mejía-Jaramillo, A. M. and Triana-Chávez, O. (2008). Biological characterization of Trypanosoma cruzi stocks from domestic and sylvatic vectors in Sierra Nevada of Santa Marta, Colombia. Acta Tropica 108, 2634.
Urbina, J. A. (2010). Specific chemotherapy of Chagas disease: relevance, current limitations and new approaches. Acta Tropica 115, 5568.
WHO (World Health Organization) (2015). Investing to Overcome the Global Impact of Neglected Tropical Diseases. Third WHO report on neglected tropical diseases, Geneva, Switzerland.
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