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Abstract:

Proliferation of Leishmania mexicana promastigotes in synthetic medium can be blocked by the depletion of intracellular polyamine pools induced by the presence of D,L-α-difluoromethylornithine (DFMO), a specific and irreversible inhibitor of ornithine decarboxylase (ODC). Here we report that DEMO-resistant cell lines growing normally at DFMO levels of 10 mM have been obtained from non-proliferating cultures after a single-step selection in the presence of high concentrations of the drug. The DFMO-resistant promastigotes underwent a morphological transformation into an 'amastigote-like' form after incubation for several hours at gradually increasing temperatures up to 35°C. The uptake of DEMO was not significantly altered in the drug-resistant cell lines but in both cases (promastigote and 'amastigote-like' forms) the ODC specific activity was increased approx. 15-fold over the normal enzymic levels found in the wild-type Leishmania. The enzyme affinities for its substrate and for DFMO gave very similar values in the drug-resistant promastigotes and the wild-type parasites. In contrast, ODC from the 'amastigote-like' Leishmania showed a higher affinity for ornithine and a decreased capacity for the binding of DEMO. An 80-fold amplification of the ODC gene and a corresponding increase in its transcripts have been detected in both DEMO-resistant Leishmania cell lines. The drug-resistant phenotypes with their characteristic morphologies, the increased levels of ODC activity and the amplification of the ODC gene have been stable for at least 6 months in the absence of selective pressure.

Registro:

Documento: Artículo
Título:α-Difluoromethylornithine-resistant cell lines obtained after one-step selection of Leishmania mexicana promastigote cultures
Autor:Sánchez, C.P.; Mucci, J.; González, N.S.; Ochoa, A.; Zakin, M.M.; Algranati, I.D.
Filiación:Institute de Investigaciones Bioquímicas Fundación Campomar, Facultad de Ciencias Exactas y Naturales, University of Buenos Aires, Patricias Argentinas 435, 1405 Buenos Aires, Argentina
Unité d'Expression des Gènes Eucaryotes, Institut Pasteur, Paris, France
Zentrum für Infektionsforschung, Universität Würzburg, Röntgenring 11, D-97070 Würzburg, Germany
Palabras clave:antitrypanosomal agent; eflornithine; messenger RNA; ornithine; ornithine decarboxylase; ornithine decarboxylase inhibitor; article; binding affinity; cell culture; cell selection; controlled study; drug resistance; drug uptake; enzyme activity; enzyme inhibitor interaction; enzyme specificity; gene amplification; Leishmania mexicana; nonhuman; priority journal; promastigote; Leishmania mexicana
Año:1997
Volumen:324
Número:3
Página de inicio:847
Página de fin:853
DOI: http://dx.doi.org/10.1042/bj3240847
Título revista:Biochemical Journal
Título revista abreviado:BIOCHEM. J.
ISSN:02646021
CODEN:BIJOA
CAS:eflornithine, 67037-37-0, 70052-12-9; ornithine, 70-26-8, 7006-33-9; ornithine decarboxylase, 9024-60-6
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_02646021_v324_n3_p847_Sanchez

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Citas:

---------- APA ----------
Sánchez, C.P., Mucci, J., González, N.S., Ochoa, A., Zakin, M.M. & Algranati, I.D. (1997) . α-Difluoromethylornithine-resistant cell lines obtained after one-step selection of Leishmania mexicana promastigote cultures. Biochemical Journal, 324(3), 847-853.
http://dx.doi.org/10.1042/bj3240847
---------- CHICAGO ----------
Sánchez, C.P., Mucci, J., González, N.S., Ochoa, A., Zakin, M.M., Algranati, I.D. "α-Difluoromethylornithine-resistant cell lines obtained after one-step selection of Leishmania mexicana promastigote cultures" . Biochemical Journal 324, no. 3 (1997) : 847-853.
http://dx.doi.org/10.1042/bj3240847
---------- MLA ----------
Sánchez, C.P., Mucci, J., González, N.S., Ochoa, A., Zakin, M.M., Algranati, I.D. "α-Difluoromethylornithine-resistant cell lines obtained after one-step selection of Leishmania mexicana promastigote cultures" . Biochemical Journal, vol. 324, no. 3, 1997, pp. 847-853.
http://dx.doi.org/10.1042/bj3240847
---------- VANCOUVER ----------
Sánchez, C.P., Mucci, J., González, N.S., Ochoa, A., Zakin, M.M., Algranati, I.D. α-Difluoromethylornithine-resistant cell lines obtained after one-step selection of Leishmania mexicana promastigote cultures. BIOCHEM. J. 1997;324(3):847-853.
http://dx.doi.org/10.1042/bj3240847