Single-dose treatment for cutaneous leishmaniasis with an easily synthesized chalcone entrapped in polymeric microparticles.

Cutaneous leishmaniasis (CL) is a major health problem in many countries and its current treatment involves multiple parenteral injections with toxic drugs and requires intensive health services. Previously, the efficacy of a single subcutaneous injection with a slow-release formulation consisting of poly(lactide-co-glycolide) (PLGA) microparticles loaded with an antileishmanial 3-nitro-2-hydroxy-4,6-dimethoxychalcone (CH8) was demonstrated in mice model. In the search for more easily synthesized active chalcone derivatives, and improved microparticle loading, CH8 analogues were synthesized and tested for antileishmanial activity in vitro and in vivo. The 3-nitro-2',4',6'-trimethoxychalcone (NAT22) analogue was chosen for its higher selectivity against intracellular amastigotes (selectivity index = 1489, as compared with 317 for CH8) and more efficient synthesis (89% yield, as compared with 18% for CH8). NAT22 was loaded into PLGA / polyvinylpyrrolidone (PVP) polymeric blend microspheres (NAT22-PLGAk) with average diameter of 1.9 µm. Although NAT22-PLGAk showed similar activity to free NAT22 in killing intracellular parasites in vitro (IC50 ~ 0.2 µm), in vivo studies in Leishmania amazonensis - infected mice demonstrated the significant superior efficacy of NAT22-PLGAk to reduce the parasite load. A single intralesional injection with NAT22-PLGAk was more effective than eight injections with free NAT22. Together, these results show that NAT22-PLGAk is a promising alternative for single-dose localized treatment of CL.

Encapsulation in lipid-core nanocapsules improves topical treatment with the potent antileishmanial compound CH8.

Cutaneous leishmaniasis (CL) is a neglected parasitic disease conventionally treated by multiple injections with systemically toxic drugs. Aiming at a more acceptable therapy, we developed lipid-core nanocapsules (LNCs) entrapping the potent antileishmanial chalcone (CH8) for topical application. Rhodamine-labeled LNC (Rho-LNC-CH8) was produced for imaging studies. LNC-CH8 and Rho-LNC-CH8 had narrow size distributions (polydispersity index <0.10), with similar mean sizes (~180 nm) by dynamic light scattering. In vitro, Rho-LNC-CH8 was rapidly internalized by extracellular Leishmania amazonensis parasites macrophages in less than 15 min. LNC-CH8 activated macrophage oxidative mechanisms more efficiently than CH8, and was more selectively toxic against the intracellular parasites. In vivo, topically applied Rho-LNC-CH8 efficiently permeated mouse skin. In L. amazonensis-infected mice, LNC-CH8 reduced the parasite load by 86% after three weeks of daily topical treatment, while free CH8 was ineffective. In conclusion, LNC-CH8 has strong potential as a novel topical formulation for CL treatment.

Novel and safe single-dose treatment of cutaneous leishmaniasis with implantable amphotericin B-loaded microparticles.

The development of an effective amphotericin B (AmB) topical formulation to replace the systemically toxic injections currently used in cutaneous leishmaniasis (CL) treatment is challenging due to poor absorption through the skin. Aiming at an effective local chemotherapy, we designed PLGA (poly(lactide-co-glycolide acid) microparticles loaded with deoxycholate amphotericin B (d-AmB) for both macrophage intracellular targeting and sustained extracellular release. For that, d-AmB/PLGA microparticles with sizes ranging from 0.5 µm to 20 µm were synthesized and tested both in vitro and in vivo. In vitro, d-AmB/PLGA was more selectively active against intracellular amastigotes of Leishmania amazonensis than free d-AmB (selectivity index = 50 and 25, respectively). In vivo, the efficacy of a single intralesional (i.l) injection with d-AmB/PLGA was determined in early and established BALB/c mouse ear lesions. In early lesions, a single injection given on day 10 of infection was more effective in controlling parasite growth than eight i.l. injections with free d-AmB, as measured on day 120. Such d-AmB/PLGA injection was also effective in established lesions (day 30), leading to 97% parasite burden reduction, as compared with d-AmB or liposomal AmB (Ambisome®) i.l. injection containing the same AmB dose. Pharmacokinetic studies showed that following d-AmB/PLGA injection, AmB leaked slower from non-infected than infected ears, yet remaining in the ear tissue for as long as 30 days. Of interest, AmB was not detectable in the circulating plasma for at least two weeks of d-AmB/PLGA injection, contrasting with the rapid and durable (2 days) detection after free d-AmB injection. Despite the transient ear swelling and local cell infiltration, no alterations in AST, ALT and creatinine serum levels was induced by d-AmB/PLGA. For its approved components, local efficacy, and single-dose applicability, this novel and safe AmB microparticle depot formulation has strong potential as a new therapy for human CL.

Broad Spectrum and Safety of Oral Treatment with a Promising Nitrosylated Chalcone in Murine Leishmaniasis.

The oral efficacy and safety of a leishmanicidal nitrochalcone (CH8) were studied in BALB/c mouse infections with Leishmania amazonensis and Leishmania infantum Although 10-fold-higher doses of CH8 were needed to produce the same antiparasitic effect as that seen with the reference drug miltefosine, the latter was nephrotoxic, whereas CH8 restored disease toxicity markers to normal. This study shows the therapeutic potential of an orally active and hepato-/nephroprotective chalcone against cutaneous and visceral leishmaniases.

Improved drug loading via spray drying of a chalcone implant for local treatment of cutaneous leishmaniasis.

Current chemotherapy of cutaneous leishmaniasis (CL), even the mildest forms, encompasses multiple and painful injections with toxic drugs that cause systemic adverse effects. Recently, we showed the promising use of poly(lactic-co-glycolic acid) (PLGA) microparticles loaded with an antileishmanial nitrosylated chalcone (CH8) for effective, safe, local, and single-dose treatment of CL. Here, we proposed to optimize the delivery system by increasing the CH8 loading in PLGA-microparticles using spray drying instead of emulsification-solvent evaporation. The effect of solvent composition and polymeric matrix changes on thermal properties, loading efficiency, particle size, morphology, and spatial drug distribution of the CH8-loaded microparticles was evaluated. The results showed that spray drying allowed a higher CH8 content (18% w/w), as contrasting with the previous solvent evaporation technique that maximally incorporated 7.8% of CH8. In vitro studies on 96-hour incubation with L. amazonensis-infected macrophages showed that entrapment in spray-dried PLGA microparticles rendered CH8 safer, preserved its antileishmanial activity, and did not affect its antioxidant properties.