Oral Efficacy of Apigenin against Cutaneous Leishmaniasis: Involvement of Reactive Oxygen Species and Autophagy as a Mechanism of Action.

BACKGROUND: The treatment for leishmaniasis is currently based on pentavalent antimonials and amphotericin B; however, these drugs result in numerous adverse side effects. The lack of affordable therapy has necessitated the urgent development of new drugs that are efficacious, safe, and more accessible to patients. Natural products are a major source for the discovery of new and selective molecules for neglected diseases. In this paper, we evaluated the effect of apigenin on Leishmania amazonensis in vitro and in vivo and described the mechanism of action against intracellular amastigotes of L. amazonensis. METHODOLOGY/PRINCIPAL FINDING: Apigenin reduced the infection index in a dose-dependent manner, with IC50 values of 4.3 µM and a selectivity index of 18.2. Apigenin induced ROS production in the L. amazonensis-infected macrophage, and the effects were reversed by NAC and GSH. Additionally, apigenin induced an increase in the number of macrophages autophagosomes after the infection, surrounding the parasitophorous vacuole, suggestive of the involvement of host autophagy probably due to ROS generation induced by apigenin. Furthermore, apigenin treatment was also effective in vivo, demonstrating oral bioavailability and reduced parasitic loads without altering serological toxicity markers. CONCLUSIONS/SIGNIFICANCE: In conclusion, our study suggests that apigenin exhibits leishmanicidal effects against L. amazonensis-infected macrophages. ROS production, as part of the mechanism of action, could occur through the increase in host autophagy and thereby promoting parasite death. Furthermore, our data suggest that apigenin is effective in the treatment of L. amazonensis-infected BALB/c mice by oral administration, without altering serological toxicity markers. The selective in vitro activity of apigenin, together with excellent theoretical predictions of oral availability, clear decreases in parasite load and lesion size, and no observed compromises to the overall health of the infected mice encourage us to supports further studies of apigenin as a candidate for the chemotherapeutic treatment of leishmaniasis.

In vitro and in vivo effects of (-)-epigallocatechin 3-O-gallate on Leishmania amazonensis.

(-)-Epigallocatechin 3-O-gallate (1), the most abundant flavanol in green tea, has been reported to have antiproliferative effects on Trypanosoma cruzi. The present study reports the effects in vitro and in vivo of 1 on Leishmania amazonensis. L. amazonensis-infected macrophages treated with 1 exhibited a significant reduction of the infection index in a dose-dependent manner, with an IC50 value of 1.6 µM. Oral administration of 1 on L. amazonensis-infected BALB/c mice (30 mg/kg/day) resulted in a decrease in the lesion size and parasite burden, without altering serological markers of toxicity. These data demonstrate the in vitro and in vivo leishmanicidal effects of compound 1.

Reactive oxygen species production by quercetin causes the death of Leishmania amazonensis intracellular amastigotes.

The present study reports the mechanism of the antileishmanial activity of quercetin against the intracellular amastigote form of Leishmania amazonensis. Treatment with 1 reduced the infection index in L. amazonensis-infected macrophages in a dose-dependent manner, with an IC50 value of 3.4 µM and a selectivity index of 16.8, and additionally increased ROS generation also in a dose-dependent manner. Quercetin has been described as a pro-oxidant that induces the production of reactive oxygen species, which can cause cell death. Taken together, these results suggest that ROS production plays a role in the mechanism of action of 1 in the control of intracellular amastigotes of L. amazonensis.

Reactive oxygen species production and mitochondrial dysfunction contribute to quercetin induced death in Leishmania amazonensis.

BACKGROUND: Leishmaniasis, a parasitic disease caused by protozoa of the genus Leishmania, affects more than 12 million people worldwide. Quercetin has generated considerable interest as a pharmaceutical compound with a wide range of therapeutic activities. One such activity is exhibited against the bloodstream parasite Trypanosoma brucei and amastigotes of Leishmania donovani. However, the mechanism of protozoan action of quercetin has not been studied. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, we report here the mechanism for the antileishmanial activity of quercetin against Leishmania amazonensis promastigotes. Quercetin inhibited L. amazonensis promastigote growth in a dose- and time- dependent manner beginning at 48 hours of treatment and with maximum growth inhibition observed at 96 hours. The IC(50) for quercetin at 48 hours was 31.4 µM. Quercetin increased ROS generation in a dose-dependent manner after 48 hours of treatment. The antioxidant GSH and NAC each significantly reduced quercetin-induced cell death. In addition, quercetin caused mitochondrial dysfunction due to collapse of mitochondrial membrane potential. CONCLUSIONS/SIGNIFICANCE: The effects of several drugs that interfere directly with mitochondrial physiology in parasites such as Leishmania have been described. The unique mitochondrial features of Leishmania make this organelle an ideal drug target while minimizing toxicity. Quercetin has been described as a pro-oxidant, generating ROS which are responsible for cell death in some cancer cells. Mitochondrial membrane potential loss can be brought about by ROS added directly in vitro or induced by chemical agents. Taken together, our results demonstrate that quercetin eventually exerts its antileishmanial effect on L. amazonensis promastigotes due to the generation of ROS and disrupted parasite mitochondrial function.