Association between tamoxifen-induced apoptosis and acidic glycosphingolipid levels in Trypanosoma cruzi

DIANA M. ACOSTA; LUCIANA L. SOPRANO,; MAXIMILIANO R. FERRERO; PATRICIA GARAVAGLIA; MONICA I ESTEVA; ALICIA S. COUTO; VILMA G. DUSCHAK

Santa Fe- Argentina

Simposio; XXIII Reunión Científica Anual Sociedad Argentina de Protozoología; 2009

Sociedad Argentina de Protozoología

Association between tamoxifen-induced apoptosis and acidic glycosphingolipid levels in Trypanosoma cruzi Diana M. Acosta1, Luciana L. Soprano1, Maximiliano R. Ferrero1, Patricia Garavaglia1, Monica I Esteva1, Alicia S. Couto2 and Vilma G. Duschak1 1 Departamento de Investigación, Inst. Nac. de Parasitología ‘Dr Mario Fatala Chaben’, ANLIS-Malbrán, Ministerio de Salud, Bs As, Argentina. 2CIHIDECAR, Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Bs As , Argentina Chagas’ disease, caused by the kinetoplastid protozoan Trypanosoma cruzi is one of the major health problems in Latin America. Acidic glycosphingolipids (AGSLs) are active participants in adhesion processes in many cellular systems and appear to be involved in the regulation of cell proliferation, differentiation, host interaction and other developmental cellular events. On the other hand, the drug tamoxifen, (Z)-1-(4-(2-(diimethylamino)ethoxyl-phenyl)-1,2-diphenyl-1-butane, is a non steroidal anti-oestrogen widely used for the treatment and prevention of breast cancer. In addition to its activity as antioestrogen receptor modulator, the modulation of caspases, calmodulin and kinases by tamoxifen was also reported, involved in mechanisms responsible for inducing apoptosis. Besides, this drug is a potent inhibitor of lipid peroxidation induced by Fe(III)-ascorbate in ox-brain phospholipid liposomes and interferes in glycosphingolipid metabolism, retarding it by inhibition of ceramide glycosylation in human cancer cells.  Herein, the activity of tamoxifen was assessed in vivo against axenically cultured epimastigotes and in vitro on blood trypomastigotes finding a dose-dependant epimastigote growth inhibition reaching up to complete inhibition of proliferation at 50 µM and a 90% lysis percentage with 10 µM in blood trypomastigote forms. Electron microscopy analysis of tamoxifen treated parasites (10-50 µM) showed characteristic apoptotic signs at the two concentrations tested but a significant increase of apoptotic cells percentage was observed at 50 µM. On the other hand, after saponification, purification of the acidic components of tamoxifen-treated (10-50 µM) and control parasites was achieved and acidic components were characterized. TLC analysis of purified showed increased levels of AGSLs at growing concentrations of tamoxifen in comparison with those synthesized by equal number of control parasites without treatment with tamoxifen. In this report we show for the first time by UV-MALDI-TOF mass spectrometry analysis a sulfatide and a ganglioside structure in T. cruzi and suggest a possible association between the increase in the biosynthesis of AGSLs and tamoxifen-induced apoptosis. In addition, tamoxifen treatment showed a significant trypanocidal activity on blood trypomastigotes in different strains under the conditions tested. Furthermore, high levels of sulfatides associated to blocking of cytokinesis found in parasites treated with 50 µM tamoxifen may be responsible for the complete inhibition of proliferation. Supported by CONICET-ANPCYT-UBA,.INP-ANLIS-Malbrán