Associação de produtos naturais e anfotericina b para o tratamento da leishmaniose visceral

Abstract

Visceral leishmaniasis (VL) is a neglected disease, caused by protozoa of the genus Leishmania, closely linked to poverty. It is also the second parasitic disease that kills the most in the world. Treatment is the main form of VL control, however, it has several limitations: it is prolonged, presents toxicity and high cost, among others. The selection of parasites resistant to available medications is also another important limitation, especially in some countries, such as Brazil. Thus, there is a need to implement new therapeutic strategies for the treatment of VL, such as repositioning and drug association, which have proven to be very important tools in the management of several diseases, such as HIV, malaria and cancer. Natural products have always been present in the search for new treatment alternatives and new drugs. The present study aimed to evaluate the action of natural products bisabolol, eugenol and lapachol, with antileishmania activity described in the literature, in combination with amphotericin b, reference drug, in vitro and in vivo models of VL. The inhibitory concentration (IC50) and cytotoxic concentration (CC50) of each drug were calculated and the therapeutic indices (IT) were determined. Next, the interaction of the combinations between natural products and amphotericin b in Leishmania infantum promastigotes was performed. The results were analyzed by the fixed proportion isobologram method, and fractional inhibitory concentrations (CIF) were analyzed; The sum of CIF (ΣCIF) and the mean ΣCIF (XΣCIF) were calculated for each combination. The nature of the interactions was classified according to the mean of XΣCIF. The combination of eugenol and bisabolol with amphotericin b showed XΣCIF values that indicated additive or indifferent interaction. Due to promising results in the in vitro assays, eugenol was selected for evaluation of association with amphotericin b in vivo. In this trial, BALB/c mice with VL were randomly assigned to five groups (n = 6) and treated (i) intraperitoneally (ip) with amphotericin b 5 mg/kg/48h/10 days (standard dose); (ii) amphotericin b at the dose of 1mg/kg/24h/10 days (subdose) ip; (iii) eugenol 75mg/kg/24h/10 days orally; with the combination of amphotericin b 1mg/kg/24h/10 days / ip and eugenol 75mg/kg/24h/10 days orally (iv), and a control group without treatment (v). After treatment, the animals were euthanized and the parasite burden of the liver and spleen determined by qPCR. There was a significant reduction in parasite load in the spleen (p <0.05) and liver (p <0.01) of the animals treated with the combination of eugenol and amphotericin b sub-dose compared to the untreated control group. Surprisingly, the impact on reducing the parasite burden on the organs of mice provided by the combination of eugenol + amphotericin b was similar to that induced by standard treatment with amphotericin b, which used a 5-fold higher dose of the drug than used in the combination. The results suggest that the combination eugenol + amphotericin b had an additive or synergic effect in a murine VL model. In conclusion, this paper proposes the association between the natural product eugenol and the reference drug amphotericin b in sub-dose as a therapeutic alternative for VL. However, further studies are needed to evaluate other combinations of natural products and conventional drugs, as well as to establish the optimal therapeutic dose and toxicity of the combination eugenol + amphotericin b, in order to propose a new therapeutic regimen for VL.