Navegando por Autor "Freitas, Camila Simões de"

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Acarbose presents in vitro and in vivo antileishmanial activity against Leishmania infantum and is a promising therapeutic candidate against visceral leishmaniasis.
(2020) Costa, Rafaella Rodrigues; Silva, João Augusto Oliveira da; Reis, Thiago Alves Rosa dos; Tavares, Grasiele de Sousa Vieira; Mendonça, Débora Vasconcelos Costa; Freitas, Camila Simões de; Lage, Daniela Pagliara; Martins, Vívian Tamietti; Antinarelli, Luciana Maria Ribeiro; Machado, Amanda Sanchez; Bandeira, Raquel Soares; Ribeiro, Fernanda Ludolf; Santos, Thaís Teodoro de Oliveira; Brito, Rory Cristiane Fortes de; Humbert, Maria Victoria; Souza, Daniel Menezes; Duarte, Mariana Costa; Chávez Fumagalli, Miguel Angel; Roatt, Bruno Mendes; Coimbra, Elaine Soares; Coelho, Eduardo Antônio Ferraz
Treatment against visceral leishmaniasis (VL) is mainly hampered by drug toxicity, long treatment regimens and/or high costs. Thus, the identifcation of novel and low-cost antileishmanial agents is urgent. Acarbose (ACA) is a specifc inhibitor of glucosidase-like proteins, which has been used for treating diabetes. In the present study, we show that this molecule also presents in vitro and in vivo specifc antileishmanial activity against Leishmania infantum. Results showed an in vitro direct action against L. infantum promastigotes and amastigotes, and low toxicity to mammalian cells. In addition, in vivo experiments performed using free ACA or incorporated in a Pluronic® F127-based polymeric micelle system called ACA/ Mic proved efective for the treatment of L. infantum-infected BALB/c mice. Treated animals presented signifcant reductions in the parasite load in their spleens, livers, bone marrows and draining lymph nodes when compared to the controls, as well as the development of antileishmanial Th1-type humoral and cellular responses based on high levels of IFN-γ, IL-12, TNF-α, GM-CSF, nitrite and IgG2a isotype antibodies. In addition, ACA or ACA-treated animals sufered from low organ toxicity. Treatment with ACA/Mic outperformed treatments using either Miltefosine or free ACA based on parasitological and immunological evaluations performed one and 15 days post-therapy. In conclusion, data suggest that the ACA/Mic is a potential therapeutic agent against L. infantum and merits further consideration for VL treatment.
A clioquinol-containing Pluronic ® F127 polymeric micelle system is effective in the treatment of visceral leishmaniasis in a murine model.
(2020) Tavares, Grasiele de Sousa Vieira; Mendonça, Débora Vasconcelos Costa; Pereira, Isabela Amorim Gonçalves; Silva, João Augusto Oliveira da; Ramos, Fernanda Fonseca; Lage, Daniela Pagliara; Machado, Amanda Sanchez; Carvalho, Lívia Mendes; Reis, Thiago Alves Rosa dos; Melo, Luísa Helena Perin de; Carvalho, Ana Maria Ravena Severino; Ottoni, Flaviano Melo; Ribeiro, Fernanda Ludolf; Freitas, Camila Simões de; Bandeira, Raquel Soares; Silva, Alessandra M.; Chávez Fumagalli, Miguel Angel; Duarte, Mariana Costa; Souza, Daniel Menezes; Alves, Ricardo José; Roatt, Bruno Mendes; Coelho, Eduardo Antônio Ferraz
A clioquinol (ICHQ)-containing Pluronic F127 polymeric micelle system (ICHQ/Mic) was recently shown to be effective against Leishmania amazonensis infection in a murine model. In the present study, ICHQ/ Mic was tested against L. infantum infection. BALB/c mice (n = 12 per group) were infected with L. infantum stationary promastigotes through subcutaneous injection and, 45 days after challenge, received saline or were treated via the subcutaneous route with empty micelles, ICHQ or ICHQ/Mic. In addition, animals were treated with miltefosine by the oral route, as a drug control. Half of the animals were euthanized 1 and 15 days after treatment, aiming to evaluate two endpoints after therapy, when parasitological and immunological parameters were investigated. Results showed that the treatment using miltefosine, ICHQ or ICHQ/Mic induced significantly higher anti-parasite IFN-c, IL-12, GM-CSF, nitrite and IgG2a isotype antibody levels, which were associated with low IL-4 and IL-10 production. In addition, a higher frequency of IFN-c and TNF-a-producing CD4+ and CD8+ T-cells was found in these animals. The parasite load was evaluated in distinct organs, and results showed that the treatment using miltefosine, ICHQ or ICHQ/Mic induced significant reductions in organic parasitism in the treated and infected mice. A comparison between the treatments suggested that ICHQ/Mic was the most effective in inducing a highly polarized Th1-type response, as well as reducing the parasite load in significant levels in the treated and infected animals. Data obtained 15 days after treatment suggested maintenance of the immunological and parasitological responses. In conclusion, ICHQ/Mic could be considered in future studies for the treatment of visceral leishmaniasis.
Digitoxigenin presents an effective and selective antileishmanial action against Leishmania infantum and is a potential therapeutic agent for visceral leishmaniasis.
(2020) Freitas, Camila Simões de; Silva, João Augusto Oliveira da; Lage, Daniela Pagliara; Costa, Rafaella Rodrigues; Mendonça, Débora Vasconcelos Costa; Martins, Vívian Tamietti; Reis, Thiago Alves Rosa dos; Antinarelli, Luciana Maria Ribeiro; Machado, Amanda Sanchez; Tavares, Grasiele de Sousa Vieira; Ramos, Fernanda Fonseca; Coelho, Vinicio Tadeu da Silva; Brito, Rory Cristiane Fortes de; Ribeiro, Fernanda Ludolf; Chávez Fumagalli, Miguel Angel; Roatt, Bruno Mendes; Ramos, Gabriela S.; Munkert, Jennifer; Ottoni, Flaviano Melo; Campana, Priscilla Rodrigues Valadares; Humbert, Maria Victoria; Coimbra, Elaine Soares; Braga, Fernão Castro; Pádua, Rodrigo Maia de; Coelho, Eduardo Antônio Ferraz
Treatment for visceral leishmaniasis (VL) is hampered mainly by drug toxicity, their high cost, and parasite resistance. Drug development is a long and pricey process, and therefore, drug repositioning may be an alternative worth pursuing. Cardenolides are used to treat cardiac diseases, especially those obtained from Digitalis species. In the present study, cardenolide digitoxigenin (DIGI) obtained from a methanolic extract of Digitalis lanata leaves was tested for its antileishmanial activity against Leishmania infantum species. Results showed that 50% Leishmania and murine macrophage inhibitory concentrations (IC50 and CC50, respectively) were of 6.9 ± 1.5 and 295.3 ± 14.5 μg/mL, respectively. With amphotericin B (AmpB) deoxycholate, used as a control drug, values of 0.13 ± 0.02 and 0.79 ± 0.12 μg/mL, respectively, were observed. Selectivity index (SI) values were of 42.8 and 6.1 for DIGI and AmpB, respectively. Preliminary studies suggested that the mechanism of action for DIGI is to cause alterations in the mitochondrial membrane potential, to increase the levels of reactive oxygen species and induce accumulation of lipid bodies in the parasites. DIGI was incorporated into Pluronic® F127-based polymeric micelles, and the formula (DIGI/Mic) was used to treat L. infantum–infected mice. Miltefosine was used as a control drug. Results showed that animals treated with either miltefosine, DIGI, or DIGI/Mic presented significant reductions in the parasite load in their spleens, livers, bone marrows, and draining lymph nodes, as well as the development of a specific Th1-type response, when compared with the controls. Results obtained 1 day after treatment were corroborated with data corresponding to 15 days after therapy. Importantly, treatment with DIGI/Mic induced better parasitological and immunological responses when compared with miltefosine- and DIGI-treated mice. In conclusion, DIGI/Mic has the potential to be used as a therapeutic agent to protect against L. infantum infection, and it is therefore worth of consideration in future studies addressing VL treatment.
Exploring drug repositioning for leishmaniasis treatment : ivermectin plus polymeric micelles induce immunological response and protection against tegumentary leishmaniasis.
(2023) Freitas, Camila Simões de; Lage, Daniela Pagliara; Machado, Amanda Sanchez; Vale, Danniele Luciana; Martins, Vívian Tamietti; Cardoso, Jamille Mirelle de Oliveira; Silva, João Augusto Oliveira da; Reis, Thiago Alves Rosa dos; Tavares, Grasiele de Sousa Vieira; Ramos, Fernanda Fonseca; Ribeiro, Fernanda Ludolf; Pereira, Isabela Amorim Gonçalves; Bandeira, Raquel Soares; Fujiwara, Ricardo Toshio; Bueno, Lilian Lacerda; Roatt, Bruno Mendes; Chávez Fumagalli, Miguel Angel; Coelho, Eduardo Antônio Ferraz
Leishmania amazonensis can cause a wide spectrum of the clinical manifestations of leishmaniasis in humans. The development of new therapeutics is a long and expensive task; in this context, drug repositioning could be considered a strategy to identify new biological actions of known products. In the present study, ivermectin (IVE) was tested against distinct Leishmania species able to cause disease in humans. In vitro experiments showed that IVE was effective to reduce the infection degree and parasite load in Leishmania donovani- and L. amazonensisinfected macrophages that were treated with it. In addition, using the culture supernatant of treated macrophages, higher production of IFN-γ and IL-12 and lower levels of IL-4 and IL-10 were found. Then, IVE was used in a pure form or incorporated into Poloxamer 407-based polymeric micelles (IVE/M) for the treatment of L. amazonensis-infected BALB/c mice. Animals (n = 16 per group) were infected and later received saline, empty micelles, amphotericin B (AmpB), IVE, or IVE/M. They were euthanized at one (n = 8 per group) and 30 (n = 8 per group) days after treatment and, in both endpoints, immunological, parasitological, and biochemical evaluations were performed. Results showed that both IVE and IVE/M induced higher levels of IFN-γ, IL-12, GM-CSF, nitrite, and IgG2a antibodies, as well as higher IFN-γ expression evaluated by RT-qPCR in spleen cell cultures. Such animals showed low organic toxicity, as well as significant reductions in the lesion’s average diameter and parasite load in their infected tissue, spleen, liver, and draining lymph node. The efficacy was maintained 30 days post-therapy, while control mice developed a polarized Th2-type response and high parasite load. In this context, IVE could be considered as a new candidate to be applied in future studies for the treatment against distinct Leishmania species.
Flau-A, a naphthoquinone derivative, is a promising therapeutic candidate against visceral leishmaniasis : a preliminary study.
(2022) Mendonça, Débora Vasconcelos Costa; Tavares, Grasiele de Sousa Vieira; Pereira, Isabela Amorim Gonçalves; Silva, João Augusto Oliveira da; Ramos, Fernanda Fonseca; Lage, Daniela Pagliara; Machado, Amanda Sanchez; Carvalho, Lívia Mendes; Reis, Thiago Alves Rosa dos; Carvalho, Ana Maria Ravena Severino; Ottoni, Flaviano Melo; Ribeiro, Fernanda Ludolf; Freitas, Camila Simões de; Martins, Vivian Tamietti; Chávez Fumagalli, Miguel Angel; Duarte, Mariana Costa; Humbertf, Maria V.; Roatt, Bruno Mendes; Souza, Daniel Menezes; Alves, Ricardo José; Coelho, Eduardo Antônio Ferraz
Visceral leishmaniasis (VL) is a neglected tropical disease found in tropical and subtropical regions in the world. The therapeutics used for the treatment against disease presents problems, mainly related to drug toxicity, route of administration, high cost and/or by emergence of resistant strains. In this context, the search for alternative antileishmanial candidates is desirable. Recently, a naphthoquinone derivative namely 2-(2,3,4-tri-O-acetyl-6- deoxy-β-L-galactopyranosyloxy)-1,4-naphthoquinone or Flau-A showed an effective in vitro biological action against Leishmania infantum. In the present study, the efficacy of this naphthoquinone derivative was evaluated in an in vivo infection model. BALB/c mice (n = 12 per group) were infected and later received saline or were treated with empty micelles (B/Mic), free Flau-A or it incorporated in Poloxamer 407-based micelles (Flau-A/ Mic). The products were administered subcutaneously in the infected animals, which were then euthanized one (n = 6 per group) and 15 (n = 6 per group) days post-therapy, when immunological and parasitological eval- uations were performed. Results showed that animals treated with Flau-A or Flau-A/Mic produced significantly higher levels of antileishmanial IFN-γ, IL-12, TNF-α, GM-CSF, nitrite and IgG2a isotype antibody, when compared to data found in the control (saline and B/Mic) groups; which showed significantly higher levels of parasite- specific IL-4, IL-10 and IgG1 antibody. In addition, animals receiving free Flau-A or Flau-A/Mic presented also significant reductions in the parasite load in their spleens, livers, bone marrows and draining lymph nodes, when compared to the controls. A low hepatic and renal toxicity was also found. Overall, Flau-A/Mic showed better immunological and parasitological results, when compared to the use of free molecule. In conclusion, pre- liminary data suggest that this composition could be considered in future studies as promising therapeutic candidate against VL.
Immunotherapy using immunogenic mimotopes selected by phage display plus amphotericin B inducing a therapeutic response in mice infected with Leishmania amazonenses.
(2023) Soyer, Tauane Gonçalves; Ramos, Fernanda Fonseca; Pereira, Isabela Amorim Gonçalves; Lage, Daniela Pagliara; Bandeira, Raquel Soares; Jesus, Marcelo Moreira de; Costa, Guilherme de Paula; Machado, Amanda Sanchez; Freitas, Camila Simões de; Vale, Danniele Luciana; Martins, Vivian Tamietti; Galdino, Alexsandro Sobreira; Chávez Fumagalli, Miguel Angel; Souza, Daniel Menezes; Duarte, Mariana Costa; Roatt, Bruno Mendes; Coelho, Eduardo Antônio Ferraz; Tavares, Grasiele de Sousa Vieira
Leishmania amazonensis can cause cutaneous and visceral clinical manifestations of leish- maniasis in infected hosts. Once the treatment against disease is toxic, presents high cost, and/or there is the emergence of parasite-resistant strains, alternative means through which to control the disease must be developed. In this context, immunotherapeutics combining known drugs with immunogens could be applied to control infections and allow hosts to recover from the disease. In this study, immunotherapeutics protocols associating mimotopes selected by phage display and amphotericin B (AmpB) were evaluated in L. amazonensis-infected mice. Immunogens, A4 and A8 phages, were administered alone or associated with AmpB. Other animals received saline, AmpB, a wild-type phage (WTP), or WTP/AmpB as controls. Evaluations performed one and thirty days after the application of immunotherapeutics showed that the A4/AmpB and A8/AmpB combinations induced the most polarized Th1-type immune responses, which reflected in significant reductions in the lesion’s average diameter and in the parasite load in the infected tissue and distinct organs of the animals. In addition, the combination also reduced the drug toxicity, as compared to values found using it alone. In this context, preliminary data presented here suggest the potential to associate A4 and A8 phages with AmpB to be applied in future studies for treatment against leishmaniasis.
In vitro and in vivo antileishmanial activity of b-acetyl-digitoxin, a cardenolide of Digitalis lanata potentially useful to treat visceral leishmaniasis.
(2021) Freitas, Camila Simões de; Lage, Daniela Pagliara; Silva, João Augusto Oliveira da; Costa, Rafaella Rodrigues; Mendonça, Débora Vasconcelos Costa; Martins, Vívian Tamietti; Reis, Thiago Alves Rosa dos; Antinarelli, Luciana Maria Ribeiro; Machado, Amanda Sanchez; Tavares, Grasiele de Sousa Vieira; Ramos, Fernanda Fonseca; Brito, Rory Cristiane Fortes de; Ribeiro, Fernanda Ludolf; Chávez Fumagalli, Miguel Angel; Roatt, Bruno Mendes; Ramos, Gabriela S.; Munkert, Jennifer; Ottoni, Flaviano Melo; Campana, Priscilla Rodrigues Valadares; Duarte, Mariana Costa; Gonçalves, Denise Utsch; Coimbra, Elaine Soares; Braga, Fernão Castro; Pádua, Rodrigo Maia de; Coelho, Eduardo Antônio Ferraz
Current treatments of visceral leishmaniasis face limitations due to drug side effects and/or high cost, along with the emergence of parasite resistance. Novel and low-cost antileishmanial agents are therefore required. We report herein the antileishmanial activity of b-acetyl-digitoxin (b-AD), a cardenolide isolated from Digitalis lanata leaves, assayed in vitro and in vivo against Leishmania infantum. Results showed direct action of b-AD against parasites, as well as efficacy for the treatment of Leishmania-infected macrophages. In vivo experiments using b-AD-containing Pluronic F127 polymeric micelles (b-AD/Mic) to treat L. infantum-infected mice showed that this composition reduced the parasite load in distinct organs in more significant levels. It also induced the development of anti-parasite Th1-type immunity, attested by high levels of IFN-c, IL-12, TNF-a, GM-CSF, nitrite and specific IgG2a antibodies, in addition to low IL-4 and IL-10 contents, along with higher IFN-c-producing CD4+ and CD8+ T-cell frequency. Furthermore, low toxicity was found in the organs of the treated animals. Comparing the therapeutic effect between the treatments, b-AD/Mic was the most effective in protecting animals against infection, when compared to the other groups including miltefosine used as a drug control. Data found 15 days after treatment were similar to those obtained one day post-therapy. In conclusion, the results obtained suggest that b-AD/Mic is a promising antileishmanial agent and deserves further studies to investigate its potential to treat visceral leishmaniasis.
Inhalation of dimethyl fumarate-encapsulated solid lipid nanoparticles attenuate clinical signs of experimental autoimmune encephalomyelitis and pulmonary inflammatory dysfunction in mice.
(2022) Pinto, Bárbara Fernandes; Ribeiro, Lorena Natasha Brito; Silva, Gisela Bevilacqua Rolfsen Ferreira da; Freitas, Camila Simões de; Rocha, Lucas Kraemer; Oliveira, Fabrício Marcus Silva; Clímaco, Marianna Carvalho; Mourão, Flavio Afonso Gonçalves; Santos, Gabryella Soares Pinheiro dos; Béla, Samantha Ribeiro; Gurgel, Isabella Luísa da Silva; Leite, Fábio de Lima; Oliveira, Anselmo Gomes de; Vilela, Maura Regina Silva da Páscoa; Lima, Onésia Cristina Oliveira; Soriani, Frederico Marianetti; Fujiwara, Ricardo Toshio; Birbrair, Alexander; Russo, Remo de Castro; Tavares, Juliana Carvalho
Rationale: The FDA-approved Dimethyl Fumarate (DMF) as an oral drug for Multiple Sclerosis (MS) treatment based on its immunomodulatory activities. However, it also caused severe adverse effects mainly related to the gastrointestinal system. Objective: Investigated the potential effects of solid lipid nanoparticles (SLNs) containing DMF, administered by inhalation on the clinical signs, central nervous system (CNS) inflammatory response, and lung function changes in mice with experimental autoimmune encephalomyelitis (EAE). Materials and methods: EAE was induced using MOG35–55 peptide in female C57BL/6J mice and the mice were treated via inhalation with DMF-encapsulated SLN (CTRL/SLN/DMF and EAE/SLN/DMF), empty SLN (CTRL/SLN and EAE/SLN), or saline solution (CTRL/saline and EAE/saline), every 72 h during 21 days. Results: After 21 days post-induction, EAE mice treated with DMF-loaded SLN, when compared with EAE/saline and EAE/SLN, showed decreased clinical score and weight loss, reduction in brain and spinal cord injury and inflammation, also related to the increased influx of Foxp3+ cells into the spinal cord and lung tissues. Moreover, our data revealed that EAE mice showed signs of respiratory disease, marked by increased vascular permeability, leukocyte influx, production of TNF-α and IL-17, perivascular and peribronchial inflammation, with pulmonary mechanical dysfunction associated with loss of respiratory volumes and elasticity, which DMF-encapsulated reverted in SLN nebulization. Conclusion: Our study suggests that inhalation of DMF-encapsulated SLN is an effective therapeutic protocol that reduces not only the CNS inflammatory process and disability progression, characteristic of EAE disease, but also protects mice from lung inflammation and pulmonary dysfunction.
Ivermectin presents effective and selective antileishmanial activity in vitro and in vivo against Leishmania infantum and is therapeutic against visceral leishmaniasis.
(2021) Reis, Thiago Alves Rosa dos; Silva, João Augusto Oliveira da; Tavares, Grasiele de Sousa Vieira; Mendonça, Débora Vasconcelos Costa; Freitas, Camila Simões de; Costa, Rafaella Rodrigues; Lage, Daniela Pagliara; Martins, Vívian Tamietti; Machado, Amanda Sanchez; Ramos, Fernanda Fonseca; Silva, Alessandra M.; Ribeiro, Fernanda Ludolf; Antinarelli, Luciana Maria Ribeiro; Brito, Rory Cristiane Fortes de; Chávez Fumagalli, Miguel Angel; Humbert, Maria Victoria; Roatt, Bruno Mendes; Coimbra, Elaine Soares; Coelho, Eduardo Antônio Ferraz
Treatment for visceral leishmaniasis (VL) is hindered mainly by the toxicity and/or high cost of therapeutic drugs. In addition, parasite resistance has been registered. Thus, there is an urgent need for the identification of novel, effective and low-cost antileishmanial agents. Since drug discovery is a long and expensive process, drug repositioning for treatment of leishmaniasis should be considered. In the present study, Ivermectin (IVE), a broad-spectrum drug used for treatment of parasitic diseases, was evaluated in vitro and in vivo against Leishmania infantum species. Results in vitro showed that IVE presented 50% Leishmania and macrophage inhibitory concentrations (IC50 and CC50, respectively) of 3.64 ± 0.48 μM and 427.50 ± 17.60 μM, respectively, with a selectivity index (SI) of 117.45; whereas Amphotericin B (AmpB), which was used as control, showed IC50 and CC50 values of 0.12 ± 0.05 μM and 1.06 ± 0.23 μM, respectively, with a corresponding SI of 8.90. Treatment with IVE effectively reduced the infection percentage and parasite burden in infected and treated macrophages and displayed a prophylactic activity by inhibiting macrophage infection with pre-treated parasites. Furthermore, preliminary studies suggested that IVE targets the parasite’s mitochondria. Activity of IVE in its free format or incorporated into Pluronic® F127-based polymeric micelles (IVE/Mic) was also evaluated in vivo as a treating drug for L. infantum-infected BALB/c mice. Miltefosine was used as a control. Results showed that Miltefosine, IVE and IVE/Mic-treated animals presented significant reductions in the parasite load in their spleens, livers, bone marrows and draining lymph nodes, as well as development of an antileishmanial Th1-type immune response one and 15 days after treatment. Notably, IVE/Mic showed a better parasitological and immunological response in comparison to other alternative treatments. In conclusion, results suggest that IVE/Mic could be considered in future studies as a therapeutic alternative to treat VL.
A Leishmania amastigote-specific hypothetical protein evaluated as recombinant protein plus Th1 adjuvant or DNA plasmid-based vaccine to protect against visceral leishmaniasis.
(2020) Silva, João Augusto Oliveira da; Machado, Amanda Sanchez; Ramos, Fernanda Fonseca; Tavares, Grasiele de Sousa Vieira; Lage, Daniela Pagliara; Mendonça, Débora Vasconcelos Costa; Pereira, Isabela Amorim Gonçalves; Santos, Thaís Teodoro de Oliveira; Martins, Vivian Tamietti; Carvalho, Lívia Mendes; Freitas, Camila Simões de; Ribeiro, Fernanda Ludolf; Reis, Thiago Alves Rosa dos; Bandeira, Raquel Soares; Silva, Alessandra M.; Costa, Lourena Emanuele; Oliveira, Jamil Silvano de; Duarte, Mariana Costa; Roatt, Bruno Mendes; Teixeira, Antônio Lúcio; Coelho, Eduardo Antônio Ferraz
Most studies evaluating vaccine candidates against visceral leishmaniasis (VL) have used parasite promastigoteexpressed antigens; however, Leishmania proteins expressed in the amastigote forms should be considered, since few hours after infection this stage comes into contact with the host immune system and is responsible for the development of the disease. In this context, in the present study, a Leishmania amastigote-specific hypothetical protein, called LiHyJ, was evaluated as a recombinant protein plus saponin as an adjuvant or DNA vaccine to protect against VL. The vaccine effect was evaluated by means of the evaluation of immunological and parasitological analyses performed in BALB/c mice against Leishmania infantum infection. Results showed that rLiHyJ/saponin and DNA LiHyJ induced significantly higher levels of anti-protein and anti-parasite IFN-γ, IL-12, GM-CSF, and IgG2a isotype antibodies, which were associated with a low presence of IL-4 and IL-10. DNA vaccination induced higher IFN-γ production, mainly by CD8+ T cells, while rLiHyJ/saponin stimulated the production of this cytokine, mainly by CD4+ T cells. The parasite load evaluated in distinct organs showed that both immunization schedules significantly reduced organic parasitism, when compared to the controls. Similar results were found in the immunological and parasitological assays when using the recombinant protein or DNA, although the vaccination with rLiHyJ plus saponin induced a slightly higher Th1 response and lower parasite load, when compared to the use of DNA plasmid. The protein also proved to be immunogenic when peripheral blood mononuclear cells of treated VL patients and healthy subjects were in vitro stimulated, since higher IFN-γ and lower IL-4 and IL-10 levels were found in the culture supernatants. In conclusion, LiHyJ should be considered in future studies as a vaccine candidate to protect against VL.
Leishmania eukaryotic elongation Factor-1 beta protein is immunogenic and induces parasitological protection in mice against Leishmania infantum infection.
(2021) Santos, Thaís Teodoro de Oliveira; Machado, Amanda Sanchez; Ramos, Fernanda Fonseca; Silva, João Augusto Oliveira da; Lage, Daniela Pagliara; Tavares, Grasiele de Sousa Vieira; Mendonça, Débora Vasconcelos Costa; Cardoso, Mariana Santos; Siqueira, Williane Fernanda; Martins, Vívian Tamietti; Ribeiro, Fernanda Ludolf; Reis, Thiago Alves Rosa dos; Carvalho, Lívia Mendes; Freitas, Camila Simões de; Bandeira, Raquel Soares; Silva, Alessandra M.; Oliveira, Jamil Silvano de; Moreira, Ricardo Luiz Fontes; Fujiwara, Ricardo Toshio; Roatt, Bruno Mendes; Chávez Fumagalli, Miguel Angel; Humbert, Maria Victoria; Teixeira, Antônio Lúcio; Coelho, Eduardo Antônio Ferraz
Treatment for visceral leishmaniasis (VL) is hampered mainly by the toxicity and/or high cost of antileishmanial drugs. What is more, variability on sensitivity and/or specificity of diagnostic tests hinders effective disease management. In this context, prophylactic vaccination should be considered as a strategy to prevent disease. In the present study, immunogenicity of the Leishmania eukaryotic Elongation Factor-1 beta (EF1b) protein, classified as a Leishmania virulence factor, was evaluated in vitro and in vivo and tested, for the first time, as a vaccine candidate against Leishmania infantum infection. The antigen was administered as DNA vaccine or as recombinant protein (rEF1b) delivered in saponin. BALB/c mice immunization with a DNA plasmid and recombinant protein plus saponin induced development of specific Th1-type immunity, characterized by high levels of IFN-γ, IL-12, GM-CSF, both T cell subtypes and antileishmanial IgG2a isotype antibodies, before and after infection. This immunological response to the vaccines was corroborated further by parasitological analysis of the vaccinated and then challenged mice, which showed significant reductions in the parasite load in their liver, spleen, bone marrow and draining lymph nodes, when compared to the controls. Vaccination using rEF1b/saponin induced a more robust Th1 response and parasitological protection when compared to the DNA vaccine. Furthermore, in vitro analysis of lymphoproliferation, IFN-γ and IL-10 levels in human PBMC cultures showed as well development of a specific Th1-type response. In conclusion, data suggest that EF1b could be a promising vaccine candidate to protect against L. infantum infection.
Leishmania LiHyC protein is immunogenic and induces protection against visceral leishmaniasis.
(2022) Machado, Amanda Sanchez; Lage, Daniela Pagliara; Vale, Danniele Luciana; Freitas, Camila Simões de; Linhares, Flávia Prata; Cardoso, Jamille Mirelle de Oliveira; Silva, João Augusto Oliveira da; Pereira, Isabela Amorim Gonçalves; Ramos, Fernanda Fonseca; Tavares, Grasiele de Sousa Vieira; Ribeiro, Fernanda Ludolf; Bandeira, Raquel Soares; Maia, Luiz Gustavo Nobre; Souza, Daniel Menezes; Duarte, Mariana Costa; Chávez Fumagalli, Miguel Angel; Roatt, Bruno Mendes; Christodoulides, Myron; Martins, Vivian Tamietti; Coelho, Eduardo Antônio Ferraz
Treatment against visceral leishmaniasis (VL) presents problems by the toxicity of drugs, high cost and/or emergence of resistant strains. The diagnosis is hampered by variable sensitivity and/or specificity of tests. In this context, prophylactic vaccina- tion could represent a control measure against disease. In this study, the protective efficacy of Leishmania LiHyC protein was evaluated in a murine model against Leish- mania infantum infection. LiHyC was used as recombinant protein (rLiHyC) associated with saponin (rLiHyC/S) or Poloxamer 407-based polymeric micelles (rLiHyC/M) to immunize mice. Animals received also saline, saponin or empty micelles as controls. The immunogenicity was evaluated before and after the challenge, and results showed that vaccination with rLiHyC/S or rLiHyC/M induced the production of high levels of interferon-gamma (IFN-γ), interleukin (IL)-12 and granulocyte-macrophage colony-stimulating factor in cell culture supernatants, as well as higher IFN-γ expres- sion evaluated by RT-qPCR and involvement from CD4+ and CD8+ T-cell subtypes producing IFN-γ, tumor necrosis factor-α and IL-2. A positive lymphoproliferative response was also found in cell cultures from vaccinated animals, besides high levels of rLiHyC- and parasite-specific nitrite and IgG2a antibodies. Immunological assays correlated with significant reductions in the parasite load in the spleens, livers, bone marrows and draining lymph nodes from vaccinated mice, when compared to values found in the controls. The micellar composition showed slightly better immunological and parasitological data, as compared to rLiHyC/S. Results suggest that rLiHyC asso- ciated with adjuvants could be considered for future studies as a vaccine candidate against VL.
Liposomal formulation of ChimeraT, a multiple T-cell epitope-containing recombinant protein, is a candidate vaccine for human Visceral Leishmaniasis.
(2020) Lage, Daniela Pagliara; Ribeiro, Patrícia Aparecida Fernandes; Dias, Daniel Silva; Mendonça, Débora Vasconcelos Costa; Ramos, Fernanda Fonseca; Carvalho, Lívia Mendes; Steiner, Bethina Trevisol; Tavares, Grasiele de Sousa Vieira; Martins, Vivian Tamietti; Machado, Amanda Sanchez; Silva, João Augusto Oliveira da; Santos, Thaís Teodoro de Oliveira; Freitas, Camila Simões de; Oliveira, Jamil Silvano de; Roatt, Bruno Mendes; Ávila, Ricardo Andrez Machado de; Humbert, Maria Victoria; Christodoulides, Myron; Coelho, Eduardo Antônio Ferraz
Background: Leishmaniases are neglected diseases caused by infection with Leishmania parasites and there are no human vaccines in use routinely. The purpose of this study was to examine the immunogenicity of ChimeraT, a novel synthetic recombinant vaccine against visceral leishmaniasis (VL), incorporated into a human-compatible liposome formulation. Methods: BALB/c mice were immunized subcutaneously with ChimeraT/liposome vaccine, ChimeraT/saponin adjuvant, or ChimeraT/saline and immune responses examined in vitro and in vivo. Results: Immunization with the ChimeraT/liposome formulation induced a polarized Th1-type response and significant protection against L. infantum infection. ChimeraT/liposome vaccine stimulated significantly high levels of interferon (IFN)-γ, interleukin (IL)-12, and granulocyte macrophage-colony stimulating factor (GM-CSF) cytokines by both CD4 and CD8 T-cells, with correspondingly lower levels of IL-4 and IL-10 cytokines. Induced antibodies were predominantly IgG2a isotype, and homologous antigen-stimulated spleen cells produced significant nitrite as a proxy for nitric oxide (NO). Furthermore, we examined a small number of treated VL patients and found higher levels of circulating anti-ChimeraT protein IgG2 antibodies, compared to IgG1 levels. Conclusions: Overall, the liposomal formulation of ChimeraT induced a protective Th1-type immune response and thus could be considered in future studies as a vaccine candidate against human VL.
Recombinant endonuclease III protein from Leishmania infantum associated with Th1-type adjuvants is immunogenic and induces protection against visceral leishmaniasis.
(2023) Lage, Daniela Pagliara; Machado, Amanda Sanchez; Freitas, Camila Simões de; Vale, Danniele Luciana; Linhares, Flávia Prata; Cardoso, Jamille Mirelle de Oliveira; Silva, João Augusto Oliveira da; Ramos, Fernanda Fonseca; Pereira, Isabela Amorim Gonçalves; Ribeiro, Fernanda Ludolf; Tavares, Grasiele de Sousa Vieira; Bandeira, Raquel Soares; Oliveira, Jamil Silvano de; Souza, Daniel Menezes; Duarte, Mariana Costa; Galdino, Alexsandro Sobreira; Christodoulides, Myron; Chávez Fumagalli, Miguel Angel; Roatt, Bruno Mendes; Martins, Vívian Tamietti; Coelho, Eduardo Antônio Ferraz
Vaccination against visceral leishmaniasis (VL) should be considered as a safe and effective measure to disease control; however, few vaccines are available against canine VL and there is no an approved human vaccine. In this context, in the present study, we evaluated the endonuclease III (ENDO) protein, which was recently showed to be antigenic for human disease, as a vaccine candidate against Leishmania infantum infection. The recombinant protein (rENDO) was administered in BALB/c mice alone or associated with saponin (rENDO/Sap) or micelles (rENDO/Mic) as adjuvants. Controls received saline, saponin or empty micelles. Results showed that both rENDO/Sap and rENDO/Mic compositions induced higher levels of IFN-γ, IL-12, TNF-α, and GM-CSF cytokines, besides nitrite and IgG2a isotype antibodies, before and after challenge infection, which were related to both CD4+ and CD8+ T cell subtypes. The immunological results contributed to significant reductions in the parasite load found in the spleens, livers, bone marrows and draining lymph nodes of the vaccinated animals. In general, mice immunized with rENDO/Mic presented a slightly higher Th1-type cellular and humoral immune response, as compared to those receiving rENDO/Sap. In addition, saponin caused a slight to moderate inflammatory edema in their vaccinated footpads, which was not observed when micelles were used with rENDO. In addition, a preliminary analysis showed that the recombinant protein was immunogenic to human cells cultures, since PBMCs from treated VL patients and healthy subjects showed higher lymphoproliferation and IFN-γ production in the culture supernatants. In conclusion, data suggest that rENDO could be considered as a candidate to be evaluated in future studies as vaccine to protect against VL.
Recombinant guanosine-5--triphosphate (GTP)-binding protein associated with Poloxamer 407-based polymeric micelles protects against Leishmania infantum infection.
(2022) Lage, Daniela Pagliara; Machado, Amanda Sanchez; Vale, Danniele Luciana; Freitas, Camila Simões de; Linhares, Flávia Prata; Cardoso, Jamille Mirelle de Oliveira; Pereira, Isabela Amorim Gonçalves; Ramos, Fernanda Fonseca; Tavares, Grasiele de Sousa Vieira; Ribeiro, Fernanda Ludolf; Silva, João Augusto Oliveira da; Bandeira, Raquel Soares; Silva, Alessandra M.; Simões, Luciana C.; Reis, Thiago Alves Rosa dos; Oliveira, Jamil Silvano de; Christodoulides, Myron; Chávez Fumagalli, Miguel Angel; Roatt, Bruno Mendes; Martins, Vivian Tamietti; Coelho, Eduardo Antônio Ferraz
Leishmania virulence proteins should be considered as vaccine candidates against disease, since they are involved in developing infection in mammalian hosts. In a previous study, a Leishmania guanosine-5′ -triphosphate (GTP)- binding protein was identified as a potential parasite virulence factor. In the present work, the gene encoding GTP was cloned and the recombinant protein (rGTP) was evaluated as a vaccine candidate against Leishmania infantum infection. The protein was associated with saponin (rGTP/Sap) or Poloxamer 407-based micelles (rGTP/ Mic) as adjuvants, and protective efficacy was investigated in BALB/c mice after parasite challenge. Both rGTP/ Sap and rGTP/Mic compositions induced a Th1-type immune response in vaccinated animals, with significantly higher levels of IFN-γ, IL-12, IL-2, TNF-α, GM-CSF, nitrite, specific IgG2a isotype antibody and positive lym- phoproliferation, when compared to the control groups. This response was accompanied by significantly lower parasite load in the spleens, livers, bone marrows and draining lymph nodes of the animals. Immunological and parasitological evaluations indicated that rGTP/Mic induced a more polarized Th1-type response and higher reduction in the organ parasitism, and with lower hepatotoxicity, when compared to the use of rGTP/Sap. In conclusion, our preliminary data suggest that rGTP could be considered for further development as a vaccine candidate to protect against VL.
A recombinant Leishmania amastigote-specific protein, rLiHyG, with adjuvants, protects against infection with Leishmania infantum.
(2022) Machado, Amanda Sanchez; Lage, Daniela Pagliara; Vale, Danniele Luciana; Freitas, Camila Simões de; Linhares, Flávia Prata; Cardoso, Jamille Mirelle de Oliveira; Pereira, Isabela Amorim Gonçalves; Ramos, Fernanda Fonseca; Tavares, Grasiele de Sousa Vieira; Ribeiro, Fernanda Ludolf; Silva, João Augusto Oliveira da; Bandeira, Raquel Soares; Simoes, Aratti Cãndido; Duarte, Mariana Costa; Oliveira, Jamil Silvano de; Christodoulides, Myron; Chávez Fumagalli, Miguel Angel; Roatt, Bruno Mendes; Martins, Vivian Tamietti; Coelho, Eduardo Antônio Ferraz
Vaccination against visceral leishmaniasis (VL) should be considered as a control measure to protect against disease, and amastigote-specific proteins could help to develop such vaccines, since this parasite form is in contact with the host immune system during the active disease. In this study, a Leishmania amastigote-specific protein, LiHyG, was evaluated as recombinant protein (rLiHyG) as vaccine candidate against Leishmania infan- tum infection in BALB/c mice. The protein was associated with saponin (rLiHyG/Sap) or Poloxamer 407-based polymeric micelles (rLiHyG/Mic) as adjuvants, and animals receiving saline, saponin or micelle as controls. Immunological and parasitological analyses were performed before (n = 8 per group; as primary endpoint) and after (n = 8 per group; as secondary endpoint) infection. Results showed that, in both endpoints, rLiHyG/Sap and rLiHyG/Mic induced higher levels of IFN-γ, IL-12 and GM-CSF in spleen cell cultures from vaccinated animals, besides elevated presence of IgG2a isotype antibodies. Decreased hepatotoxicity and ‘positive lymphoprolifer- ative response were also found after challenge. Such findings reflected in significantly lower levels of parasite load found in their spleens, livers, bone marrows and draining lymph nodes. In conclusion, rLiHyG associated with Th1-type adjuvant could be considered for future studies as vaccine candidate to protect against VL.