RESUMO
Emetic tartar (ET), was used in the treatment of leishmaniasis but its use was discontinued due to its low therapeutic index. Liposomes have been shown to be a promising strategy for delivery of bioactive substances in the region of interest, in order to reduce and/or eliminate undesirable effects. In the present study, liposomes containing ET were prepared and characterized to evaluate acute toxicity as well as their leishmanicidal action using BALB/c mice with an inoculum of Leishmania (Leishmania) infantum. Liposomes were composed of egg phosphatidylcholine and 3ß-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol, with an average diameter of 200 nm, zeta potential of +18 mV, and ET encapsulated into liposomes at a concentration near 2 g/L. Healthy mice were treated with ET or liposome containing ET (Lip-ET) in a single dose of 16 mg/kg of Sb3+ intravenously and observed for 14 days. The death of two animals in the ET-treated group and no deaths in the Lip-ET-treated group was observed. Higher hepatic and cardiac toxicity were observed in animals treated with ET when compared to animals treated with Lip-ET, blank liposomes (Blank-Lip) and PBS. The study of antileishmanial efficacy was conducted by intraperitoneal administration of Lip-ET, for ten consecutive days. It was observed by limiting dilution that treatments with liposomal formulations containing ET, as well as Glucantime®, led to a significant reduction in parasitic load in spleen and liver (p < 0.05) when compared to the untreated control group.
RESUMO
The treatment against visceral leishmaniasis (VL) presents problems, mainly related to the toxicity and/or high cost of the drugs. In this context, a prophylactic vaccination is urgently required. In the present study, a Leishmania protein called LiHyE, which was suggested recently as an antigenic marker for canine and human VL, was evaluated regarding its immunogenicity and protective efficacy in BALB/c mice against Leishmania infantum infection. In addition, the protein was used to stimulate peripheral blood mononuclear cells (PBMCs) from VL patients before and after treatment, as well as from healthy subjects. Vaccination results showed that the recombinant (rLiHyE) protein associated with liposome or saponin induced effective protection in the mice, since significant reductions in the parasite load in spleen, liver, draining lymph nodes, and bone marrow were found. The parasitological protection was associated with Th1-type cell response, since high IFN-γ, IL-12, and GM-CSF levels, in addition to low IL-4 and IL-10 production, were found. Liposome induced a better parasitological and immunological protection than did saponin. Experiments using PBMCs showed rLiHyE-stimulated lymphoproliferation in treated patients' and healthy subjects' cells, as well as high IFN-γ levels in the cell supernatant. In conclusion, rLiHyE could be considered for future studies as a vaccine candidate against VL.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Antígenos de Protozoários/administração & dosagem , Leishmania infantum/imunologia , Leishmaniose Visceral/prevenção & controle , Animais , Antígenos de Protozoários/imunologia , Feminino , Humanos , Imunogenicidade da Vacina , Leishmaniose Visceral/imunologia , Leishmaniose Visceral/parasitologia , Leucócitos Mononucleares/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Carga Parasitária , Células Th1/imunologia , VacinaçãoRESUMO
The control measures against visceral leishmaniasis (VL) include a precise diagnosis of disease, the treatment of human cases, and reservoir and vector controls. However, these are insufficient to avoid the spread of the disease in specific countries worldwide. As a consequence, prophylactic vaccination could be interesting, although no effective candidate against human disease is available. In the present study, the Leishmania infantum amastin protein was evaluated regarding its immunogenicity and protective efficacy against experimental VL. BALB/c mice immunized with subcutaneous injections of the recombinant protein with or without liposome/saponin (Lip/Sap) as an adjuvant. After immunization, half of the animals per group were euthanized and immunological evaluations were performed, while the others were challenged with L. infantum promastigotes. Forty-five days after infection, the animals were euthanized and parasitological and immunological evaluations were performed. Results showed the development of a Th1-type immune response in rAmastin-Lip and rAmastin-Sap/vaccinated mice, before and after infection, which was based on the production of protein and parasite-specific IFN-γ, IL-12, GM-CSF, and nitrite, as well as the IgG2a isotype antibody. CD4+ T cells were mainly responsible for IFN-γ production in vaccinated mice, which also presented significant reductions in parasitism in their liver, spleen, draining lymph nodes, and bone marrow. In addition, PBMC cultures of treated VL patients and healthy subjects stimulated with rAmastin showed lymphoproliferation and higher IFN-γ production. In conclusion, the present study shows the first case of an L. infantum amastin protein associated with distinct delivery systems inducing protection against L. infantum infection and demonstrates an immunogenic effect of this protein in human cells.
Assuntos
Leishmania infantum/imunologia , Leishmaniose Visceral/imunologia , Proteínas de Protozoários/imunologia , Adjuvantes Imunológicos/farmacologia , Sequência de Aminoácidos , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/parasitologia , Células Cultivadas , Feminino , Humanos , Imunidade/imunologia , Interferon gama/imunologia , Leishmaniose Visceral/parasitologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/parasitologia , Linfonodos/imunologia , Linfonodos/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/imunologia , Células Th1/imunologia , Células Th1/parasitologiaRESUMO
Breast cancer is the most incident and mortal cancer type in women, with an estimated 2 million new cases expected by 2020 worldwide, with 600,000 deaths. As not all breast cancer types respond to the anti-hormonal therapy, the development of new antineoplastic drugs is necessary. Lawsone (2-hydroxy-1,4-naphtoquinone) is a natural bioactive naphtoquinone displaying a range of activities, with dozens of derivatives described in the literature, including some glycosides possessing antitumor activity. Here, a series of glycosides of lawsone are reported for the first time and all compounds displayed good activity against the SKBR-3 cell line, with IC50 below 10 µM. The most promising derivative was the glycosyl triazole derived from peracetylated d-glucose (11), which showed better cytotoxicity against SKBR-3 (IC50 = 0.78 µM), being the most selective toward this tumoral cell (SI > 20). All compounds described in this work were more active than lawsone, indicating the importance of the carbohydrate and glycosyl triazole moiety for activity.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Glicosídeos/síntese química , Glicosídeos/uso terapêutico , Naftoquinonas/síntese química , Naftoquinonas/uso terapêutico , Feminino , Humanos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
The era of chemotherapy began in the 1940s, but it was in the 1960s that it was seen as really promising when the first patients with childhood acute lymphoblastic leukemia were cured with combination chemotherapy. Today, it is known that due to resistance to single agents, combination therapy is essential for tumor eradication and cure. In the last decade, studies have shown that anticancer drug combinations can act synergistically or antagonistically against tumor cells in vitro, depending on the ratios of the individual drugs forming the combination. From this observation and facing the possibility of maintaining the in vivo synergistic ratio of combinations came the idea of co-encapsulating anticancer agents in nanosystems. In vivo studies validated this idea by showing that the co-encapsulation of anticancer agents in liposomes allows the maintenance of drug ratios in the plasma and the delivery of fixed drug ratios directly to tumor tissue, leading to a better efficacy compared to the administration of the free drugs combination. Liposomes co-encapsulating irinotecan/floxuridine are now in Phase II trial, and liposomes co-encapsulating cytarabine/daunorubicin were recently approved by the FDA for treatment of patients with acute myeloid leukemia.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias/tratamento farmacológico , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Combinação de Medicamentos , Sinergismo Farmacológico , Humanos , Lipossomos/administração & dosagem , Lipossomos/efeitos adversos , Lipossomos/química , Neoplasias/patologiaRESUMO
Leishmania proteins have been evaluated as vaccine candidates against leishmaniasis; however, most antigens present low immunogenicity and need to be added with immune adjuvants. A low number of licensed adjuvants exist on the market today; therefore, research conducted to produce new products is desirable. The present study sought to evaluate the immunogenicity and protective efficacy of a recombinant Leishmania hypothetical protein, namely LiHyR, administered with saponin or liposomes in BALB/c mice. Immunological and parasitological parameters were evaluated, and results showed significant protection against Leishmania infantum infection produced by both compositions in the immunized animals; however, this was not identified when the antigen was used alone. In addition, the liposomal formulation was more effective in inducing a polarized Th1 response in the vaccinated animals, which was maintained after challenge and reflected by lower parasitism found in all evaluated organs when the limiting dilution technique and RT-PCR assay were employed. The protected animals showed higher levels of protein and parasite-specific IFN-γ IL-2, IL-12, GM-CSF, and TNF-α, which were evaluated by capture ELISA and flow cytometry, in addition to a higher production of anti-protein and anti-parasite IgG2a antibodies, both before and after challenge. The Lip/rLiHyR combination induced higher IFN-γ production through both CD4+ and CD8+ T cell subtypes. Results indicate the possibility of using the LiHyR, containing a liposomal formulation, as a vaccine candidate against visceral leishmaniasis.
Assuntos
Citocinas/imunologia , Imunogenicidade da Vacina , Leishmania infantum/imunologia , Vacinas contra Leishmaniose/farmacologia , Leishmaniose Visceral/prevenção & controle , Proteínas de Protozoários/farmacologia , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Vacinas contra Leishmaniose/imunologia , Leishmaniose Visceral/imunologia , Leishmaniose Visceral/patologia , Lipossomos , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Protozoários/imunologiaRESUMO
A range of antitumor agents for cancer treatment is available; however, they show low specificity, which often limit their use. Recently, we have reported the preparation of folate-coated long-circulating and pH-sensitive liposomes (SpHL-folate-PTX) loaded with paclitaxel (PTX), an effective drug for the treatment of solid tumors, including breast cancer. The purpose of this study was to prepare and characterize SpHL-PTX and SpHL-folate-PTX radiolabeled with technetium-99m (99mTc). Biodistribution studies and scintigraphic images were performed after intravenous administration of 99mTc-PTX, 99mTc-SpHL-PTX and 99mTc-SpHL-folate-PTX into healthy and tumor-bearing mice. High radiochemical purity (>98%) and in vitro stability (>90%) were achieved for both liposome formulations. The pharmacokinetic properties of 99mTc-SpHL-DTPA-PTX and 99mTc-SpHL-folate-DTPA-PTX decreased in a monophasic manner showing half-life of 400.1 and 541.8min, respectively. Scintigraphic images and biodistribution studies showed a significant uptake in liver, spleen and kidneys, demonstrating these routes as way for excretion. At 8h post-injection, the liposomal tumor uptake was higher than 99mTc-PTX. Interesting, 4h after administration, the liposome folate coated showed higher tumor-to-muscle ratio than 99mTc-SpHL-DTPA-PTX and 99mTc-PTX. In conclusion, the liposomal systems, showed high tumor uptake by scintigraphic images, especially the 99mTc-SpHL-folate-DTPA-PTX that showed a sustained and higher tumor-to-muscle ratio than non-functionalized liposome, which indicate its feasibility as a PTX delivery system to folate positive tumors.
Assuntos
Sistemas de Liberação de Medicamentos/métodos , Ácido Fólico/administração & dosagem , Paclitaxel/administração & dosagem , Tecnécio/administração & dosagem , Animais , Neoplasias da Mama/sangue , Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Portadores de Fármacos/administração & dosagem , Portadores de Fármacos/metabolismo , Feminino , Ácido Fólico/sangue , Humanos , Lipossomos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Paclitaxel/sangue , Tecnécio/sangue , Distribuição TecidualRESUMO
Qa-2 is believed to mediate a protective immune response against cancer; however, little is known about the role of Qa-2 in tumorigenesis. Here, we used 4T1 breast cancer cells to study the involvement of Qa-2 in tumor progression in a syngeneic host. Qa-2 expression was reduced during in vivo tumor growth and in cell lines derived from 4T1-induced tumors. Tumor-derived cells elicited an epithelial-mesenchymal transition associated with upregulation of Zeb1 and Twist1/2 and enhanced tumor initiating and invasive capacities. Furthermore, these cells showed increased stem characteristics, as demonstrated by upregulation of Hes1, Sox2 and Oct3/4, and enrichment of CD44high/CD24median/low cells. Remarkably, Qa-2 cell-surface expression was excluded from the CD44high/CD24median/low subpopulation. Tumor-derived cells showed increased Src activity, and treatment of these cells with the Src kinase inhibitor PP2 enhanced Qa-2 but reduced Sox2 and CD44high/CD24median/low expression levels, suggesting that Src signaling, while positively associated with stemness, negatively regulates Qa-2 expression in breast cancer. Finally, overexpression of the Qa-2 family member Q7 on the cell surface slowed down in vivo tumor growth and reduced the metastatic potential of 4T1 cells. These results suggest an anti-malignant role for Qa-2 in breast cancer development, which appears to be absent from cancer stem cells.
Assuntos
Neoplasias da Mama/patologia , Transição Epitelial-Mesenquimal , Antígenos de Histocompatibilidade Classe I/metabolismo , Células-Tronco Neoplásicas/patologia , Animais , Apoptose , Neoplasias da Mama/metabolismo , Carcinogênese , Movimento Celular , Proliferação de Células , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Células-Tronco Neoplásicas/metabolismo , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The high incidence and mortality of breast cancer supports efforts to develop innovative imaging probes to effectively diagnose, evaluate the extent of the tumor, and predict the efficacy of tumor treatments while concurrently and selectively delivering anticancer agents to the cancer tissue. In the present study we described the preparation of technetium-99m (99mTc)-labeled paclitaxel (PTX) and evaluated its feasibility as a radiotracer for breast tumors (4T1) in BALB/c mice. Thin Layer Chromatography (TLC) was used to determine the radiochemical purity and in vitro stability of 99mTc-PTX. PTX micelles showed a unimodal distribution with mean diameter of 13.46±0.06nm. High radiochemical purity (95.8±0.3%) and in vitro stability (over than 95%), up to 24h, were observed. Blood circulation time of 99mTc-PTX was determined in healthy BALB/c mice. 99mTc-PTX decays in a one-phase manner with a half-life of 464.3 minutes. Scintigraphic images and biodistribution were evaluated at 4, 8 and 24h after administration of 99mTc-PTX in 4T1 tumor-bearing mice. The data showed a significant uptake in the liver, spleen and kidneys, due to the importance of these routes for excretion. Moreover, high tumor uptake was achieved, indicated by high tumor-to-muscle ratios. These findings indicate the usefulness of 99mTc-PTX as a radiotracer to identify 4T1 tumor in animal models. In addition, 99mTc-PTX might be used to follow-up treatment protocols in research, being able to provide information about tumor progression after therapy.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias da Mama/diagnóstico por imagem , Paclitaxel/farmacologia , Compostos Radiofarmacêuticos/farmacologia , Animais , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacocinética , Feminino , Meia-Vida , Humanos , Marcação por Isótopo , Camundongos , Camundongos Endogâmicos BALB C , Especificidade de Órgãos , Paclitaxel/química , Paclitaxel/farmacocinética , Cintilografia , Compostos Radiofarmacêuticos/química , Compostos Radiofarmacêuticos/farmacocinética , Tecnécio , Distribuição TecidualRESUMO
Nowadays cancer is one of the most common causes of deaths worldwide. Conventional antitumor agents still present various problems related to specificity for tumor cells often leading to therapeutic failure. Nanoscale particles are considered potential alternative to direct access of drugs into tumor cells, therefore increasing the drug accumulation and performance. The aim of this study was to evaluate the antitumor activity of doxorubicin (DOX)-loaded nanostructured lipid carriers (NLC) versus liposomes against a breast cancer animal experimental model. NLC-DOX and liposomes-DOX were successfully prepared and characterized. Tumor-bearing mice were divided into five groups (blank-NLC, blank-liposome, DOX, NLC-DOX, liposome-DOX). Each animal received by the tail vein four doses of antitumoral drugs (total dose, 16mg/kg), every 3 days. Antitumor efficacy was assessed by measuring 1) tumor volume, calculating the inhibitory ratio (TV-IR, see after) and 2) acquiring scintigraphic images of the tumor using doxorubicin radiolabeled with technetium-99m as an imaging tumor probe. Liposome-DOX and free DOX did not showed differences in the tumor mean volume, whereas NLC-DOX proved to be the best treatments in controlling the tumor growth. NLC-DOX showed an inhibition ration (TV-IR) of 73.5% while free DOX and liposome-DOX decreased TV-RI of 48.8% and 68.0%, respectively. Tumor was clearly visualized in controls, DOX, and liposome-DOX groups. Yet, regarding the NLC-DOX group, tumor was barely identified by the image, indicating antitumor efficacy. Moreover, both NLC and liposomes proved to be able to delay the occurrence of lung metastasis. In conclusion, results of this study indicated that NLC-DOX might be an alternative strategy to achieve an efficient antitumor activity.