RESUMEN
The multifactorial basis of therapeutic response can obscure the relation between antimicrobial drug susceptibility and clinical outcome. To discern the relationship between parasite susceptibility to meglumine antimoniate (SbV) and therapeutic outcome of cutaneous leishmaniasis, risk factors for treatment failure were considered in evaluating this relationship in ninety-one cutaneous leishmaniasis patients and corresponding clinical strains of Leishmania (Viannia) panamensis. Parasite susceptibility to 32 µg SbV/mL (plasma Cmax) was evaluated in primary human macrophages, PBMCs, and U937 macrophages. Early parasitological response to treatment was determined in lesions of a subgroup of patients, and pathogenicity of Sb-resistant and sensitive clinical strains was compared in BALB/c mice. Parasite survival in cell models and patient lesions was determined by qRT-PCR of Leishmania 7SLRNA transcript. Parasite loads in BALB/c mice were quantified by limiting dilution analysis. The disparate Sb-susceptibility of parasite subpopulations distinguished by isoenzyme profiles (zymodemes) was manifest in all cell models. Notably, Sb-resistance defined by parasite survival, was most effectively discerned in U937 macrophages compared with primary human host cells, significantly higher among strains from patients who failed treatment than cured and, significantly associated with treatment failure. Each unit increase in transformed survival rate corresponded to a 10.6-fold rise in the odds of treatment failure. Furthermore, treatment failure was significantly associated with naturally Sb-resistant zymodeme 2.3 strains, which also produced larger lesions and parasite burdens in BALB/c mice than Sb-sensitive zymodeme 2.2 strains. The confounding effect of host risk factors for treatment failure in discerning this association was evidenced in comparing strains from patients with and without the defined risk factors for treatment failure. These results establish the association of natural resistance to meglumine antimoniate with treatment failure, the importance of host risk factors in evaluating drug susceptibility and treatment outcome, and the clinical and epidemiological relevance of natural Sb-resistance in L. (V.) panamensis subpopulations.
Asunto(s)
Antiprotozoarios , Resistencia a Medicamentos , Leishmaniasis Cutánea , Macrófagos , Antimoniato de Meglumina , Meglumina , Ratones Endogámicos BALB C , Compuestos Organometálicos , Insuficiencia del Tratamiento , Animales , Leishmaniasis Cutánea/tratamiento farmacológico , Leishmaniasis Cutánea/parasitología , Antimoniato de Meglumina/uso terapéutico , Antimoniato de Meglumina/farmacología , Humanos , Antiprotozoarios/uso terapéutico , Antiprotozoarios/farmacología , Femenino , Meglumina/uso terapéutico , Meglumina/farmacología , Compuestos Organometálicos/uso terapéutico , Compuestos Organometálicos/farmacología , Ratones , Macrófagos/parasitología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Masculino , Leishmania guyanensis/efectos de los fármacos , Adulto , Persona de Mediana Edad , Adulto Joven , Carga de Parásitos , AdolescenteRESUMEN
Besides being scarce, the drugs available for treating cutaneous leishmaniasis have many adverse effects. Ozone is an option to enhance the standard treatment due to the wound-healing activity reported in the literature. In this study, we evaluated the efficiency of ozonated sunflower oil as an adjuvant in treating cutaneous lesions caused by Leishmania amazonensis. BALB/c mice were infected with L. amazonensis, and after the lesions appeared, they were treated in four different schedules using the drug treatment with meglumine antimoniate (Glucantime®), with or without ozonated oil. After thirty days of treatment, the lesions' thickness and their parasitic burden, blood leukocytes, production of NO and cytokines from peritoneal macrophages and lymph node cells were analyzed. The group treated with ozonated oil plus meglumine antimoniate showed the best performance, improving the lesion significantly. The parasitic burden showed that ozonated oil enhanced the leishmanicidal activity of the treatment, eliminating the parasites in the lesion. Besides, a decrease in the TNF levels from peritoneal macrophages and blood leukocytes demonstrated an immunomodulatory action of ozone in the ozonated oil-treated animals compared to the untreated group. Thus, ozonated sunflower oil therapy has been shown as an adjuvant in treating Leishmania lesions since this treatment enhanced the leishmanicidal and wound healing effects of meglumine antimoniate.
Asunto(s)
Antiprotozoarios , Leishmaniasis Cutánea , Ozono , Animales , Ratones , Antimoniato de Meglumina/farmacología , Antimoniato de Meglumina/uso terapéutico , Aceite de Girasol/uso terapéutico , Antiprotozoarios/farmacología , Meglumina/farmacología , Meglumina/uso terapéutico , Leishmaniasis Cutánea/tratamiento farmacológico , Leishmaniasis Cutánea/parasitología , Cicatrización de Heridas , Ozono/uso terapéutico , Ratones Endogámicos BALB CRESUMEN
Currently, zinc oxide (ZnO) particles are used in nanotechnology to destroy a wide range of microorganisms. Although pentavalent antimony compounds are used as antileishmanial drugs, they are associated with several limitations and side effects. Therefore, it is always desirable to try to find new and effective treatments. The aim of this research is to determine the antileishmanial effect of ZnO particles in comparison to the Antimoan Meglumine compound on promastigotes and amastigotes of Leishmania major (MRHO/IR/75/ER). After the extraction and purification of macrophages from the peritoneal cavity of C57BL/6 mice, L. major parasites were cultured in Roswell Park Memorial Institute-1640 culture medium containing fetal bovine serum (FBS) 10% and antibiotic. In this experimental study, the effect of different concentrations of nanoparticles was investigated using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) colorimetric method, in comparison to the glucantime on promastigotes, amastigotes and healthy macrophages in the culture medium. The amount of light absorption of the obtained color from the regeneration of tetrazolium salt to the product color of formazan by the parasite was measured by an enzyme-linked immunosorbent assay (ELISA) reader, and the IC50 value was calculated. IC50 after 24 h of incubation was calculated as IC50 = 358.6 µg/mL. The results showed, that the efficacy of ZnO nanoparticles was favorable and dose-dependent. The concentration of 500 µg/mL of ZnO nanoparticles induced 84.67% apoptosis after 72. Also, the toxicity of nanoparticles was less than the drug. Nanoparticles exert their cytotoxic effects by inducing apoptosis. They can be suitable candidates in the pharmaceutical industry in the future.
Asunto(s)
Antiprotozoarios , Leishmania major , Antimoniato de Meglumina , Óxido de Zinc , Óxido de Zinc/farmacología , Óxido de Zinc/química , Animales , Leishmania major/efectos de los fármacos , Ratones , Antiprotozoarios/farmacología , Antimoniato de Meglumina/farmacología , Ratones Endogámicos C57BL , Nanopartículas/química , Macrófagos/parasitología , Macrófagos/efectos de los fármacos , Concentración 50 Inhibidora , Macrófagos Peritoneales/parasitología , Macrófagos Peritoneales/efectos de los fármacos , Nanopartículas del Metal/químicaRESUMEN
This study aimed to investigate the in vitro and in silico antileishmanial activity of azacitidine (AZA) on Leishmania major promastigotes and amastigotes. The in silico method was used to evaluate the possibility of the interaction of AZA into the binding pocket of inducible nitric oxide synthase (iNOS), a leading defensive oxidative metabolite. Following that, in vitro anti-promastigote, and anti-amastigote activity of AZA was determined using an MTT assay and a macrophage model, respectively. Cytotoxic effects of AZA and meglumine antimoniate (MA) were also assessed by MTT assay on murine macrophages. All experiments were performed in triplicate. The results showed that AZA interacted with Ser133, Gln134, and Lys13 amino acids of iNOS, and the molecular docking score was obtained at -241.053 kcal/mol. AZA in combination with MA significantly (P<0.001) inhibited the growth rate of nonclinical promastigote (IC50 247.6±7.3 µM) and 8.5-fold higher of clinical intramacrophage amastigote stage (29.8±5.3 µM), compared to the untreated group. A significant upsurge of Th1 subsets and transcription genes and a meaningful decline in Th2 cytokines subclasses at the equivalent concentrations of AZA and MA was observed (P<0.001). The apoptosis effect of AZA along with MA was significantly induced on L. major in a dose-dependent manner (P<0.001). The present study demonstrated that AZA possesses antileishmanial activity in in vitro and in silico models. However, AZA combined with MA was more effective than AZA alone in inhibiting the growth rate of promastigotes and amastigotes of L. major. This study indicates that AZA in combination with MA demonstrated a potent antileishmanial mechanism, promoting immune response and enhancing an immunomodulatory role toward the Th1 pathway. This experimental study is a basic study for applying more knowledge about the mechanisms of AZA along with MA in animal models in the future.
Asunto(s)
Antiprotozoarios , Leishmania major , Animales , Ratones , Antimoniato de Meglumina/farmacología , Azacitidina , Simulación del Acoplamiento Molecular , Antiprotozoarios/farmacologíaRESUMEN
BACKGROUND: We decided to investigate the antileishmanial, cellular mechanisms, and cytotoxic effects of green synthesized Zinc nanoparticles (ZnNPs) alone and combined with glucantime against Leishmania major infection. METHODS: The effect of green synthesized ZnNP on L. major amastigote was studied through macrophage cells. The mRNA expression level of iNOS and IFN-γ followed by the exposure of J774-A1 macrophage cells to ZnNPs was assessed by Real-time PCR. The Caspase-3-like activity of promastigotes exposed to ZnNPs was studied. Effects of ZnNPs alone and combined with glucantime (MA) were studied on cutaneous leishmaniasis in BALB/c mice. RESULTS: ZnNPs displayed the spherical shape with sizes ranging from 30 to 80 nm. The obtained IC50 values for ZnNPs, MA, and ZnNPs + MA were 43.2, 26.3, and 12.6 µg/mL, respectively; indicating the synergistic effects of ZnNPs in combination with MA. CL lesions had completely improved in the mice received with ZnNPs in combination with MA. The mRNA expression level of iNOS, TNF-α, and IFN-γ was dose-dependently (p < 0.01) upregulated; whereas it was downregulated in IL-10. ZnNPs markedly stimulated the caspase-3 activation with no significant toxicity on normal cells. CONCLUSION: Based on these in vitro and in vivo results, green synthesized ZnNPs, mainly along with MA, showed that has the potential to be introduced as a new drug for CL therapy. Triggering of NO production, and inhibition of infectivity rate are revealed as mechanisms of action ZnNPs on L. major. But, supplementary investigations are necessary to clear the efficacy and safety of these agents.
Asunto(s)
Antineoplásicos , Antiprotozoarios , Leishmania major , Leishmaniasis Cutánea , Nanopartículas del Metal , Animales , Ratones , Antimoniato de Meglumina/farmacología , Caspasa 3/genética , Zinc/farmacología , Antiprotozoarios/farmacología , Leishmaniasis Cutánea/tratamiento farmacológico , Antineoplásicos/farmacología , Ratones Endogámicos BALB CRESUMEN
Leishmaniasis is a tropical zoonotic disease. It is found in 98 countries, with an estimated 1.3 million people being affected annually. During the life cycle, the Leishmania parasite alternates between promastigote and amastigote forms. The first line treatment for leishmaniasis are the pentavalent antimonials, such as N-methylglucamine antimoniate (Glucantime®) and sodium stibogluconate (Pentostam®). These drugs are commonly related to be associated with dangerous side effects such as cardiotoxicity, nephrotoxicity, hepatotoxicity, and pancreatitis. Considering these aspects, this work aimed to obtain a new series of limonene-acylthiosemicarbazides hybrids as an alternative for the treatment of leishmaniasis. For this, promastigotes, axenic amastigotes, and intracellular amastigotes of Leishmania amazonensis were used in the antiproliferative assay; J774-A1 macrophages for the cytotoxicity assay; and electron microscopy techniques were performed to analyze the morphology and ultrastructure of parasites. ATZ-S-04 compound showed the best result in both tests. Its IC50 , in promastigotes, axenic amastigotes and intracellular amastigotes was 0.35±0.08â µM, 0.49±0.06â µM, and 15.90±2.88â µM, respectively. Cytotoxicity assay determined a CC50 of 16.10±1.76â µM for the same compound. By electron microscopy, it was observed that ATZ-S-04 affected mainly the Golgi complex, in addition to morphological changes in promastigote forms of L. amazonensis.
Asunto(s)
Antiprotozoarios , Leishmania , Leishmaniasis , Humanos , Animales , Ratones , Limoneno/farmacología , Antiprotozoarios/farmacología , Antiprotozoarios/química , Leishmaniasis/parasitología , Macrófagos , Antimoniato de Meglumina/farmacología , Ratones Endogámicos BALB CRESUMEN
INTRODUCTION: Leishmaniasis is an important disease caused by parasites of the Leishmania. Due to the urgent need for financial incentives and research and development of new anti-Leishmania, a point that stands out is the creation of patents that comprise drugs and nanoformulations in treating the disease. AREAS COVERED: Information on individual patents and patent families containing potential drugs and nanoformulations were obtained by searching the Orbit software (QUESTEL SAS, France) using the following terms: Leishmania; treatment; nanoparticle*; drug×. The data obtained ranged from 2015 to 2022. EXPERT OPINION: Meglumine antimoniate is a pentavalent antimonial widely used in the classic treatment of leishmaniasis. It is part of the classic treatment recommended by WHO, being the first-choice drug globally about 75 years ago. Thus, the need to introduce new anti-Leishmania therapies into clinical medicine is evident since cases of resistance to monotherapy and multitherapy have increased greatly. Associated with this, the search for patents that are good candidates in treating this disease assues interest in investments of financial resources and raises a ray of hope for safe, effective, and low-cost therapies to become licensed for the treatment of leishmaniasis.
Asunto(s)
Antiprotozoarios , Leishmania , Leishmaniasis , Humanos , Antiprotozoarios/farmacología , Patentes como Asunto , Leishmaniasis/tratamiento farmacológico , Antimoniato de Meglumina/farmacología , Antimoniato de Meglumina/uso terapéuticoRESUMEN
Current therapeutic ways adopted for the treatment of leishmaniasis are toxic and expensive including parasite resistance is a growing problem. Given this scenario, it is urgent to explore treatment alternatives for leishmaniasis. The aim of this study was to evaluate the effect of 3-phenyl-lawsone (3-PL) naphthoquinone on Leishmania (Viannia) braziliensis infection, both in vitro and in vivo, using two local routes of administration: subcutaneous (higher dose) and tattoo (lower dose). In vitro 3-PL showed low toxicity for macrophages (CC50 >3200 µM/48h) and activity against intracellular amastigotes (IC50 = 193 ± 19 µM/48h) and promastigotes (IC50 = 116 ± 26 µM/72h), in which induced increased ROS generation. Additionally, 3-PL up-regulated the production of cytokines such as tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein 1 (MCP-1), interleukin-6 (IL-6) and IL-10 in infected macrophages. However, the anti-amastigote action was independent of nitric oxide production. Treatment of hamsters infected with L. (V.) braziliensis from one week after infection with 3-PL by subcutaneous (25 µg/Kg) or tattooing (2.5 µg/Kg) route, during 3 weeks (3 times/week) or 2 weeks (2 times/week) significantly decreased the parasite load (p<0.001) in the lesion. The reduction of parasite load by 3-PL treatment was comparable to reference drug meglumine antimoniate administered by the same routes (subcutaneous 1mg/Kg and tattoo 0.1mg/Kg). In addition, treatment started from five weeks after infection with 3-PL per tattoo also decreased the parasite load. These results show the anti-leishmanial effect of 3-PL against L. (V.) braziliensis and its efficacy by subcutaneous (higher dose) and tattoo (lower dose) routes. In addition, this study shows that drug delivery by tattooing the lesion allows the use of lower doses than the conventional subcutaneous route, which may support the development of a new therapeutic strategy that can be adopted for leishmaniasis.
Asunto(s)
Antiprotozoarios , Leishmania braziliensis , Leishmaniasis Cutánea , Naftoquinonas , Tatuaje , Cricetinae , Animales , Antimoniato de Meglumina/farmacología , Antimoniato de Meglumina/uso terapéutico , Antiprotozoarios/farmacología , Antiprotozoarios/uso terapéutico , Leishmaniasis Cutánea/tratamiento farmacológico , Leishmaniasis Cutánea/parasitología , Naftoquinonas/farmacología , Naftoquinonas/uso terapéutico , Carga de ParásitosRESUMEN
Lipophosphoglycan (LPG), the major Leishmania glycoconjugate, induces pro-inflammatory/immunosuppressive innate immune responses. Here, we evaluated functional/biochemical LPG properties from six Leishmania amazonensis strains from different hosts/clinical forms. LPGs from three strains (GV02, BA276, and LV79) had higher pro-inflammatory profiles for most of the mediators, including tumor necrosis factor alpha and interleukin 6. For this reason, glycoconjugates from all strains were biochemically characterized and had polymorphisms in their repeat units. They consisted of three types: type I, repeat units devoid of side chains; type II, containing galactosylated side chains; and type III, containing glucosylated side chains. No relationship was observed between LPG type and the pro-inflammatory properties. Finally, to evaluate the susceptibility against antileishmanial agents, two strains with high (GV02, BA276) and one with low (BA336) pro-inflammatory activity were selected for chemotherapeutic tests in THP-1 cells. All analyzed strains were susceptible to amphotericin B (AmB) but displayed various responses against miltefosine (MIL) and glucantime (GLU). The GV02 strain (canine visceral leishmaniasis) had the highest IC50 for MIL (3.34 µM), whereas diffuse leishmaniasis strains (BA276 and BA336) had a higher IC50 for GLU (6.87-12.19 mM). The highest IC50 against MIL shown by the GV02 strain has an impact on clinical management. Miltefosine is the only drug approved for dog treatment in Brazil. Further studies into drug susceptibility of L. amazonensis strains are warranted, especially in areas where dog infection by this species overlaps with those caused by Leishmania infantum.
Asunto(s)
Anfotericina B , Leishmania , Anfotericina B/farmacología , Animales , Perros , Glicoesfingolípidos , Interleucina-6 , Leishmania/genética , Antimoniato de Meglumina/farmacología , Ratones , Ratones Endogámicos BALB C , Fosforilcolina/análogos & derivados , Factor de Necrosis Tumoral alfaRESUMEN
Objective: Leishmaniasis is the second deadliest parasitic disease in the World Health Organisation's list of neglected diseases, following malaria. Cutaneous leishmaniasis (CL) is the most common form of the disease and it is one of the few communicable diseases with increasing incidence rates owing to factors like armed conflicts and climate change. CL can be divided into two major groups: Acute CL (ACL) and chronic CL (CCL). The aim of this study was to compare the in vitro efficacy of miltefosine and pentavalent antimony compounds in the CCL patient samples. Methods: Five isolates previously isolated from 5 CCL patients were included in this study. Genotyping is performed using internal transcribed spacer 1 (ITS 1) gene region real-time PCR. In vitro drug efficacy tests were applied to determine their activity against meglumine antimoniate (MA) and miltefosine. Serial dilutions (512, 256, 128, 64, 32, 16, 8 and 4 µg/mL) prepared from MA and miltefosine were prepared in 96-well flat-bottom cell culture plates and incubated at 24 °C for 48 hours. The efficacy of the drug on Leishmania spp. promastigotes after 24 and 48 hours was evaluated by hemocytometer slide and XTT cell viability test. Results: All of the samples were genotyped as L. tropica. Evaluation of 24 and 48 hours showed, 128 µg/mL and 256 µg/mL and 32 µg/mL and 64 µg/mL concentrations of miltefosine and MA were enough to kill all the promastigotes respectively. The results of the hemocytometer slide and XTT were consistent. Conclusion: There are no studies investigating the in vitro efficacy of miltefosine with the CCL patient group. To overcome the treatment challenges experienced in this special patient group, more studies are needed. According to our results, it is concluded that miltefosine is efficient for the treatment of CCL and further clinical studies with miltefosine will reveal valuable data.
Asunto(s)
Antiprotozoarios , Leishmaniasis Cutánea , Antiprotozoarios/farmacología , Antiprotozoarios/uso terapéutico , Humanos , Leishmaniasis Cutánea/tratamiento farmacológico , Antimoniato de Meglumina/farmacología , Antimoniato de Meglumina/uso terapéutico , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacología , Fosforilcolina/uso terapéuticoRESUMEN
INTRODUCTION: Leishmaniasis denotes a significant health challenge worldwide with no ultimate treatment. The current study investigated the biological effects of gamma-terpinene (GT) on Leishmania major in putative antileishmanial action, cytotoxicity, apoptosis induction, gene expression alteration, antioxidant activity, hemolysis, and ROS generation. METHODS: GT and meglumine antimoniate (MA) were probed alone and in combination (GT/MA) for their anti-leishmanial potentials using the MTT biochemical colorimetric assay and a model macrophage cell. In addition, their immunomodulatory properties were assessed by analyzing their effect on the transcription of cytokines related to Th1 and Th2 responses. GT and MA, alone and in combination, were also assessed for their potential to alter metacaspase gene expression in L. major promastigotes by real-time RT-PCR. The hemolytic potential of GT and MA-treated promastigotes were also measured by routine UV absorbance reading. Electrophoresis on agarose gel was employed to analyze genomic DNA fragmentation. RESULTS: GT demonstrated notable dose-dependent antileishmanial effects towards promastigotes and amastigotes of L. major. The IC50 values for GT against L. major promastigotes and amastigotes were 46.76 mM and 25.89 mM, respectively. GT was considerably safer towards murine macrophages than L. major amastigotes with an SI value of 3.17. Transcriptional expression of iNOS, JAK-1, Interleukin (IL-10), and TGF-ß was meaningfully decreased, while the levels of metacaspase mRNA were increased. Results also confirmed GT antioxidant activities. Also, increased levels of intracellular ROS were observed upon treatment of promastigotes with GT. The gel electrophoresis result indicated slight DNA fragmentation in the treated promastigotes by both drugs. A weak hemolytic effect was also observed for GT. CONCLUSION: The results demonstrated that GT showed potent activity against L. major stages. It seems that its mechanism of action involves representing an immunomodulatory role towards upregulation of iNOS and JAK-1, while downregulation of IL-10 and TGF- ß. Moreover, GT has an antioxidative potential and exerts its action through activating macrophages to kill the organism. Further in vivo and clinical studies are essential to explore its effect in future programs.
Asunto(s)
Antiprotozoarios , Leishmania major , Animales , Antioxidantes/farmacología , Antiprotozoarios/farmacología , Antiprotozoarios/uso terapéutico , Monoterpenos Ciclohexánicos , Interleucina-10 , Antimoniato de Meglumina/farmacología , Ratones , Ratones Endogámicos BALB C , Especies Reactivas de OxígenoRESUMEN
Treatment of leishmaniasis by conventional synthetic compounds has faced a serious challenge worldwide. This study was performed to evaluate the effect and modes of action of aromatic Turmerone on the Leishmania major intra-macrophage amastigotes, the causative agent of zoonotic cutaneous leishmaniasis in the Old World. In the findings, the mean numbers of L. major amastigotes in macrophages were significantly decreased in exposure to Turmerone plus meglumine antimoniate (Glucantime®; MA) than MA alone, especially at 50 µg/mL. In addition, Turmerone demonstrated no cytotoxicity as the selectivity index (SI) was 21.1; while it induced significant apoptosis in a dose-dependent manner on L. major promastigotes. In silico molecular docking analyses indicated an affinity of Turmerone to IL-12, with the MolDock score of - 96.8 kcal/mol; which may explain the increased levels of Th1 cytokines and decreased level of IL-10. The main mechanism of action is more likely associated with stimulating a powerful antioxidant and promoting the immunomodulatory roles in the killing of the target organism.
Asunto(s)
Antiprotozoarios , Leishmania major , Leishmaniasis Cutánea , Compuestos Organometálicos , Animales , Antioxidantes/farmacología , Antiprotozoarios/farmacología , Antiprotozoarios/uso terapéutico , Leishmaniasis Cutánea/tratamiento farmacológico , Leishmaniasis Cutánea/veterinaria , Meglumina/farmacología , Meglumina/uso terapéutico , Antimoniato de Meglumina/farmacología , Simulación del Acoplamiento Molecular , Compuestos Organometálicos/farmacología , Compuestos Organometálicos/uso terapéuticoRESUMEN
This study compared the therapeutic potential of the chemotherapy using meglumine antimoniate encapsulated in a mixture of conventional and PEGylated liposomes (Nano Sbv) and immunotherapy with anti-canine IL-10 receptor-blocking monoclonal antibody (Anti IL-10R) on canine visceral leishmaniasis (CVL). Twenty mongrel dogs naturally infected by L. infantum, displaying clinical signs of visceral leishmaniasis were randomly divided in two groups. In the first one, nine dogs received six intravenous doses of a mixture of conventional and PEGylated liposomes containing meglumine antimoniate at 6.5 mg Sb/kg/dose. In the second one, eleven dogs received two intramuscular doses of 4 mg of anti-canine IL-10 receptor-blocking monoclonal antibody. The animals were evaluated before (T0) and 30, 90, and 180 days after treatments. Our major results demonstrated that both treatments were able to maintain hematological and biochemical parameters, increase circulating T lymphocytes subpopulations, increase the IFN-γ producing T-CD4 lymphocytes, restore the lymphoproliferative capacity and improve the clinical status. However, although these improvements were observed in the initial post-treatment times, they did not maintain until the end of the experimental follow-up. We believe that the use of booster doses or the association of chemotherapy and immunotherapy (immunochemotherapy) is promising to improve the effectiveness of treating CVL for improving the clinical signs and possibly reducing the parasite burden in dogs infected with Leishmania infantum.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Enfermedades de los Perros/tratamiento farmacológico , Leishmaniasis Visceral/tratamiento farmacológico , Liposomas/química , Antimoniato de Meglumina/farmacología , Polietilenglicoles/química , Receptores de Interleucina-10/antagonistas & inhibidores , Alopurinol/farmacología , Animales , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/metabolismo , Enfermedades de los Perros/metabolismo , Perros , Factores Inmunológicos/metabolismo , Inmunoterapia/métodos , Leishmania infantum/efectos de los fármacos , Leishmaniasis Visceral/metabolismo , Compuestos Organometálicos/farmacologíaRESUMEN
OBJECTIVES: Leishmaniasis is a zoonotic disease and several drugs have been used in the treatment, including meglumine antimoniate (AME). The chemotherapy reaches clinical cure but does not eliminate parasites, contributing to drug resistance. To improve AME efficacy we incorporated it in anionic liposomes. The antiparasitic activity and intracellular localization were investigated in canine macrophages infected with Leishmania infantum. METHODS: Liposomes (L-AME) is composed of egg phosphatidylcholine, cholesterol, palmitoyl oleoyl phosphatidyl serine and α-tocopherol (4 : 3 : 0.4 : 0.07 mol%) plus AME. L-AME size, polydispersity, zeta potential and morphology were analysed as well as antileishmanial activity and intracellular localization in DH82 macrophages. KEY FINDINGS: Liposomes (360 nm) zeta potential range from -40 to -65 mV, had 23% encapsulation efficiency and were stable for 180 days at 4°C. Free AME was cytotoxic towards L. infantum infected macrophages (ID50 = 0.012 M) while L-AME did not reduce cell viability. L-AME colocalized with parasites inside macrophages in a time-dependent manner, and reduced the percentage of infected cells and the number of intracellular parasites, decreasing the infection index (75-80%) twice as compared with AME treatment. CONCLUSIONS: Liposomal AME is a promising delivery system for treating visceral leishmaniasis, improving meglumine efficacy against L. infantum and minimizing its cytotoxicity towards canine macrophages.
Asunto(s)
Leishmania infantum , Compuestos Organometálicos , Animales , Perros , Liposomas , Macrófagos , Antimoniato de Meglumina/farmacología , Ratones , Ratones Endogámicos BALB C , Compuestos Organometálicos/farmacología , Compuestos Organometálicos/uso terapéuticoRESUMEN
BACKGROUND: Leishmaniasis is an infectious disease caused by parasites of the genus Leishmania and presents different clinical manifestations. The adverse effects, immunosuppression and resistant strains associated with this disease necessitate the development of new drugs. Nanoparticles have shown potential as alternative antileishmanial drugs. We showed in a previous study the biosynthesis, characterization and ideal concentration of a nanocomposite that promoted leishmanicidal activity. In the present study, we conducted a specific analysis to show the mechanism of action of AgNP-PVP-MA (silver nanoparticle-polyvinylpyrrolidone-[meglumine antimoniate (Glucantime®)]) nanocomposite during Leishmania amazonensis infection in vitro. RESULTS: Through ultrastructural analysis, we observed significant alterations, such as the presence of small vesicles in the flagellar pocket and in the extracellular membrane, myelin-like structure formation in the Golgi complex and mitochondria, flagellum and plasma membrane rupture, and electrodense material deposition at the edges of the parasite nucleus in both evolutive forms. Furthermore, the Leishmania parasite infection index in macrophages decreased significantly after treatment, and nitric oxide and reactive oxygen species production levels were determined. Additionally, inflammatory, and pro-inflammatory cytokine and chemokine production levels were evaluated. The IL-4, TNF-α and MIP-1α levels increased significantly, while the IL-17 A level decreased significantly after treatment. CONCLUSIONS: Thus, we demonstrate in this study that the AgNP-PVP-MA nanocomposite has leishmanial potential, and the mechanism of action was demonstrated for the first time, showing that this bioproduct seems to be a potential alternative treatment for leishmaniasis.
Asunto(s)
Antiprotozoarios/farmacología , Antiprotozoarios/uso terapéutico , Leishmania/efectos de los fármacos , Nanocompuestos/uso terapéutico , Animales , Células Cultivadas , Técnicas In Vitro , Leishmania/fisiología , Leishmania/ultraestructura , Macrófagos/parasitología , Antimoniato de Meglumina/química , Antimoniato de Meglumina/farmacología , Antimoniato de Meglumina/uso terapéutico , Nanopartículas del Metal/química , Nanopartículas del Metal/uso terapéutico , Ratones , Ratones Endogámicos BALB C , Povidona/química , Povidona/farmacología , Povidona/uso terapéutico , Plata/química , Plata/farmacología , Plata/uso terapéuticoRESUMEN
Leishmaniasis is a human and animal disease endemic in tropical and subtropical areas treated by means of pentavalent antimony as first-line approach. Unfortunately, the formulations available on the market are characterized by significant side effects and a total remission of the disease is difficult to be obtained. The aim of this investigation is to describe the development and characterization of aqueous-core poly-l-lactide (PLA) nanocapsules containing glucantime (meglumine antimoniate, MA) with the aim of increasing the pharmacological efficacy of the active compound. The polymeric systems characterized by a mean diameter of ≈300 nm exert a great interaction with murine macrophages. MA-loaded PLA nanocapsules show a great antileishmanial activity on mice infected with Leishmania infantum with respect to the free drug, favoring a decrease of the administration times. The biodistribution profiles demonstrate a lower renal accumulation of MA after its nanoencapsulation and a significant increase of its plasmatic half-life. The parasite load evaluated by immunohistochemistry shows a significant decrease in liver, spleen, and kidneys when mice are treated with MA-loaded PLA nanocapsules especially after 45 days. The obtained results demonstrate the potential application of MA-loaded PLA nanocapsules as novel nanomedicine for the treatment of leishmaniasis.
Asunto(s)
Leishmania infantum , Nanocápsulas , Compuestos Organometálicos , Animales , Meglumina/química , Meglumina/farmacología , Meglumina/uso terapéutico , Antimoniato de Meglumina/farmacología , Ratones , Ratones Endogámicos BALB C , Compuestos Organometálicos/química , Poliésteres/farmacología , Distribución TisularRESUMEN
Cutaneous leishmaniasis (CL) is a neglected tropical skin disease caused by the protozoan genus Leishmania. The treatment is restricted to a handful number of drugs that exhibit toxic effects, limited efficacy, and drug resistance. Additionally, developing an effective topical treatment is still an enormous unmet medical challenge. Natural oils, e.g. the oleoresin from P. emarginatus fruits (SO), contain various bioactive molecules, especially terpenoid compounds such as diterpenes and sesquiterpenes. However, its use in topical formulations can be impaired due to the natural barrier of the skin for low water solubility compounds. Nanoemulsions (NE) are drug delivery systems able to increase penetration of lipophilic compounds throughout the skin, improving their topical effect. In this context, we propose the use of SO-containing NE (SO-NE) for CL treatment. The SO-NE was produced by a low energy method and presented suitable physicochemical characteristic: average diameter and polydispersity index lower than 180 nm and 0.2, respectively. Leishmania (Leishmania) amazonensis-infected BALB/c mice were given topical doses of SO or SO-NE. The topical use of a combination of SO-NE and intraperitoneal meglumine antimoniate reduced lesion size by 41 % and tissue regeneration was proven by histopathological analyses. In addition, a reduction in the parasitic load and decreased in the level of IFN-γ in the lesion may be associated, as well as a lower level of the cytokine IL-10 may be associated with a less intense inflammatory process. The present study suggests that SO-NE in combination meglumine antimoniate represents a promising alternative for the topical treatment of CL caused by L. (L.) amazonensis.
Asunto(s)
Fabaceae , Leishmania mexicana/efectos de los fármacos , Leishmaniasis Cutánea/tratamiento farmacológico , Extractos Vegetales/farmacología , Piel/efectos de los fármacos , Tripanocidas/farmacología , Administración Tópica , Animales , Citocinas/metabolismo , Modelos Animales de Enfermedad , Composición de Medicamentos , Quimioterapia Combinada , Emulsiones , Fabaceae/química , Femenino , Interacciones Huésped-Parásitos , Leishmania mexicana/crecimiento & desarrollo , Leishmaniasis Cutánea/parasitología , Leishmaniasis Cutánea/patología , Antimoniato de Meglumina/farmacología , Mesocricetus , Ratones Endogámicos BALB C , Nanopartículas , Carga de Parásitos , Extractos Vegetales/aislamiento & purificación , Piel/parasitología , Piel/patología , Tripanocidas/aislamiento & purificaciónRESUMEN
BACKGROUND: Drug resistance is a common phenomenon frequently observed in countries where leishmaniasis is endemic. Due to the production of the pteridine reductase enzyme (PTR1), drugs lose their efficacy, and consequently, the patient becomes unresponsive to treatment. This study aimed to compare the in vitro effect of meglumine antimoniate (MA) on non- healing Leishmania tropica isolates and on MA transfected non-healing one to PTR1. METHODS: Two non-healing and one healing isolates of L. tropica were collected from patients who received two courses or one cycle of intralesional MA along with biweekly liquid nitrogen cryotherapy or systemic treatment alone, respectively. After confirmation of L. tropica isolates by polymerase chain reaction (PCR), the recombinant plasmid pcDNA-rPTR (antisense) was transfected via electroporation and cultured on M199. Isolates in form of promastigotes were treated with different concentrations of MA and read using an enzyme-linked immunosorbent assay (ELISA) reader and the half inhibitory concentration (IC50 ) value was calculated. The amastigotes were grown in mouse macrophages and were similarly treated with various concentrations of MA. The culture glass slides were stained, and the mean number of intramacrophage amastigotes and infected macrophages were assessed in triplicate for both stages. RESULTS: All three transfected isolates displayed a reduction in optical density compared with the promastigotes in respective isolates, although there was no significant difference between non-healing and healing isolates. In contrast, in the clinical form (amastigotes), there was a significant difference between non-healing and healing isolates (p < 0.05). CONCLUSION: The results indicated that the PTR1 gene reduced the efficacy of the drug, and its inhibition by antisense and could improve the treatment of non-healing cases. These findings have future implications in the prophylactic and therapeutic modality of non- healing Leishmania isolates to drug.
Asunto(s)
Leishmania tropica/genética , Leishmaniasis Cutánea/tratamiento farmacológico , Leishmaniasis Cutánea/genética , Oxidorreductasas/genética , Proteínas Protozoarias/genética , Adulto , Animales , Antiprotozoarios/farmacología , Antiprotozoarios/uso terapéutico , ADN sin Sentido , Farmacorresistencia Microbiana/efectos de los fármacos , Farmacorresistencia Microbiana/genética , Femenino , Humanos , Leishmania tropica/efectos de los fármacos , Leishmania tropica/aislamiento & purificación , Leishmaniasis Cutánea/parasitología , Masculino , Antimoniato de Meglumina/farmacología , Antimoniato de Meglumina/uso terapéutico , Ratones Endogámicos BALB C , TransfecciónRESUMEN
World Health Organization reported that approximately one billion people are at risk in endemic areas, one million cases of cutaneous leishmaniasis (CL) and approximately 300,000 cases of visceral leishmaniasis (VL) were reported per year in the last five years. The number of deaths due to VL is reported to be approximately 20,000 per year. Approximately 2500 cases/year have been reported as CL, caused by Leishmania tropica and Leishmania infantum, in Turkey. The significant increase observed in many cities mainly in the provinces of Mediterranean and Aegean regions in cases and foci in recent years, suggests that there may be an increase in this infections in the following years as well. In Turkey, the causative agent of CL is L.tropica and meglumine antimoniate is used in the treatment of CL. We aimed to determine antimony resistance genes specific for L.tropica by comparing the gene and protein expressions of antimony-resistant and non-resistant L.tropica strains. L.tropica isolates obtained from 3 CL patients without antimonate resistance from Aegean, Mediterranean and Southeastern regions of Turkey were provided to transform into 3 resistant isolates against meglumine antimony in the laboratory conditions. Gene expression alterations by microarray method; protein profiles by two-dimensional gel electrophoresis (2D-PAGE) and relevant proteins by MALDI-TOF/TOF MS of these isolates were accomplished and compared. L.tropica isolates from 10 CL patients who did not respond to antimony therapy were analyzed for resistance to antimonial compounds and quantitative real-time polymerase chain reaction was performed to detect the expression of genes responsible for resistance development. Moreover, differences in protein expression levels in isolates with and without antimony resistance were determined by comparing protein profiles and identification of proteins with different expression levels was carried out. Enolase, elongation factor-2, heat shock protein 70, tripanthione reductase, protein kinase C and metallo-peptidase proteins have been shown to play roles in L.tropica isolates developing resistance to antimonial compounds and similar expression changes have also been demonstrated in naturally resistant isolates from patients. In conclusion, it was revealed that L.tropica strains in our country may gain resistance to meglumine antimoniate in a short time. It is foreseen that if the patients living in our country or entering the country are treated inadequately and incompletely, there may be new, resistant leishmaniasis foci that may increase the number of resistant strains and cases rapidly.
Asunto(s)
Resistencia a Medicamentos , Leishmania tropica , Leishmaniasis Cutánea , Antimoniato de Meglumina , Resistencia a Medicamentos/genética , Humanos , Leishmania tropica/efectos de los fármacos , Leishmaniasis Cutánea/tratamiento farmacológico , Antimoniato de Meglumina/farmacología , Antimoniato de Meglumina/uso terapéutico , TurquíaRESUMEN
The inadequacy of available treatments for leishmaniasis has presented up to 40% therapeutic failure. This fact suggests an urgency in the discovery of new drugs or alternative approaches for treating this disease. The objective of this study was to evaluate the antileishmanial activity of combined therapy between crotamine (CTA) from Crotalus durissus terrificus and the pentavalent antimonial Glucantime® (GLU). The assays were in vitro performed measuring the inhibition of Leishmania amazonensis amastigotes, followed by the evaluation of cellular production of cytokines and nitrites. After that, analytical methods were performed in order to characterize the molecules involved in the study by Mass Spectrometry, molecular affinity through an in silico assay and Surface Plasmon Resonance. In vivo experiments with BALB/c mice were performed by analyzing parasitemia, lesion size and immunological mediators. In the in vitro experiments, the pharmacological association improved the inhibition of the amastigotes, modulated the production of cytokines and nitric oxide. The therapy improved the effectiveness of the GLU, demonstrating a decreased parasitemia in the infected tissues. Altogether, the results suggest that the combined approach with CTA and GLU may be a promising alternative for the treatment of cutaneous leishmaniasis.