Introduction Leishmaniasis is a major parasitic disease. WHO estimates that 1.3 million new cases and 20000 to 30000 deaths occur annually, and a population of 350 million is at risk. Available chemotherapy is far from satisfaction because antileishmanial drugs are costly with unpleasant side effects. The situation has further worsened with emergence of drug resistance in various regions of endemicity. The anti-metabolite 6-aminonicotinamide (6AN) is converted to 6-aminoNAD and 6-aminoNADP, the latter being a strong inhibitor of the pentose phosphate pathway. We tested 6AN ability to inhibit parasite growth. Methods L. infantum promastigotes were grown at 25°C in RPMI with 15% FBS. The number of viable cells was measured either by Alamar Blue fluorescence or using a hemocytometer. 6-phosphogluconate dehydrogenase (6PGD) activity was measured spectrophotometrically. Results Effect of 6AN on oxidative stress. Parasites incubated 48 h with 6AN, were treated with 100 uM H2O2 for 45 min. The number of viable parasites in control cultures decreased by 7.2% (±5), while in cultures preincubated with 100 or 250 uM 6AN decreased by 38.7% (±1.2) and 50% (±2.6). Effect of 6AN on the growth rate in absence of oxidative stress. When the parasites were grown in the presence of 200 uM 6AN, the growth rate, measured after 48 and 96 h, decreased by 33% (p<0.05). Effect of 6AN on 6PGD activity. 6PGD is the main target of 6-aminoNADP. The parasites, grown with or without 6AN, were lysed and the 6PGD activity was assayed. The enzymatic activity of 6AN treated cells was reduced to 30% of the activity of untreated cells. The inhibition extent does not change by changing the dilution of the cell extract, meaning that the inhibition is not due to the residual presence of the inhibitor. Conclusions 6AN shows good antileishmanial activity. In fact its effect on the parasites is not limited to an increased sensitivity to the oxidative stress. 6PGD is confirmed as main target of the inhibitor

Effect of the antimetabolite 6-aminonicotinamide on Leishmania infantum promastigotes

BELLINI, Tiziana;CONTINI, Carlo;MARITATI, Martina;TRENTINI, Alessandro;DALLOCCHIO, Franco Pasquale Filippo;HANAU, Stefania
2015

Abstract

Introduction Leishmaniasis is a major parasitic disease. WHO estimates that 1.3 million new cases and 20000 to 30000 deaths occur annually, and a population of 350 million is at risk. Available chemotherapy is far from satisfaction because antileishmanial drugs are costly with unpleasant side effects. The situation has further worsened with emergence of drug resistance in various regions of endemicity. The anti-metabolite 6-aminonicotinamide (6AN) is converted to 6-aminoNAD and 6-aminoNADP, the latter being a strong inhibitor of the pentose phosphate pathway. We tested 6AN ability to inhibit parasite growth. Methods L. infantum promastigotes were grown at 25°C in RPMI with 15% FBS. The number of viable cells was measured either by Alamar Blue fluorescence or using a hemocytometer. 6-phosphogluconate dehydrogenase (6PGD) activity was measured spectrophotometrically. Results Effect of 6AN on oxidative stress. Parasites incubated 48 h with 6AN, were treated with 100 uM H2O2 for 45 min. The number of viable parasites in control cultures decreased by 7.2% (±5), while in cultures preincubated with 100 or 250 uM 6AN decreased by 38.7% (±1.2) and 50% (±2.6). Effect of 6AN on the growth rate in absence of oxidative stress. When the parasites were grown in the presence of 200 uM 6AN, the growth rate, measured after 48 and 96 h, decreased by 33% (p<0.05). Effect of 6AN on 6PGD activity. 6PGD is the main target of 6-aminoNADP. The parasites, grown with or without 6AN, were lysed and the 6PGD activity was assayed. The enzymatic activity of 6AN treated cells was reduced to 30% of the activity of untreated cells. The inhibition extent does not change by changing the dilution of the cell extract, meaning that the inhibition is not due to the residual presence of the inhibitor. Conclusions 6AN shows good antileishmanial activity. In fact its effect on the parasites is not limited to an increased sensitivity to the oxidative stress. 6PGD is confirmed as main target of the inhibitor
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11392/2330080
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