Evaluation of anti-leishmanial activity of artemisinin combined with amphotericin B or miltefosine in Leishmania donovani promastigotes

Abstract

Background: The increasing incidence of drug resistance in Leishmaniasis necessitates evaluation of combination chemotherapy. Miltefosine and amphotericin B are established anti-leishmanial drugs, while artemisinin has shown significant leishmanicidal activity in experimental models. In this study, we have evaluated the additive/synergistic effect of artemisinin with amphotericin B or miltefosine. Methods: Leishmania parasites were isolated from the bone marrow aspirate of a patient with visceral leishmaniasis. Parasites were typed as Leishmania donovani by restriction fragment length polymorphism of internal transcribed spacer 1 region of Leishmania genome. Promastigotes were incubated in a fixed ratio combination of artemisinin (0-500 µM) and amphotericin B (0-100 nM) or miltefosine (0-100 µM) and cell viability was assessed. An isobologram was constructed to evaluate the additive/synergistic effect, wherein it was considered additive if the mean sum fractional inhibitory concentration (mean ΣFIC) at the IC₅₀ level was <2, but ≥1 and synergism, if the mean ΣFIC was <1. Results: The isobologram showed an additive effect for three combinations of artemisinin-amphotericin B and artemisinin-miltefosine, the mean ΣFICs ranging from 1.02 to 1.44 and 1.08 to 1.33 along with a synergistic effect with one combination, the mean ΣFICs being 0.58 and 0.81 respectively. Conclusions: This study supports the combination use of artemisinin-amphotericin B and artemisinin-miltefosine, worthy of future pharmacological consideration.