An optimized nanoparticle delivery system based on chitosan and chondroitin sulfate molecules reduces the toxicity of amphotericin B and is effective in treating tegumentary leishmaniasis

Amphotericin B (AmpB) is active against leishmaniasis, but its use is hampered due to its high toxicity observed in patients. In this study, a nanoparticles-delivery system for AmpB (NQC-AmpB), containing chitosan and chondroitin sulfate molecules, was evaluated in BALB/c mice against Leishmania amazonensis. An in vivo biodistribution study, including biochemical and toxicological evaluations, was performed to evaluate the toxicity of AmpB. Nanoparticles were radiolabeled with technetium-99m and injected in mice. The products presented a similar biodistribution in the liver, spleen, and kidneys of the animals. Free AmpB induced alterations in the body weight of the mice, which, in the biochemical analysis, indicated hepatic and renal injury, as well as morphological damage to the kidneys of the animals. In general, no significant organic alteration was observed in the animals treated with NQC-AmpB. Mice were infected with L. amazonensis and treated with the nanoparticles or free AmpB; then, parasitological and immunological analyses were performed. The NQC-AmpB group, as compared to the control groups, presented significant reductions in the lesion size and in the parasite burden in all evaluated organs. These animals presented significantly higher levels of IFN-γ and IL-12, and low levels of IL-4 and IL-10, when compared to the control groups. The NQC-AmpB system was effective in reducing the infection in the animals, and proved to be effective in diminishing the toxicity evoked by AmpB, which was observed when it was administered alone. In conclusion, NQC-AmpB could be considered a viable possibility for future studies in the treatment of leishmaniasisThis work was supported by grants from Pró-Reitoria de Pesquisa from UFMG (Edital 01/2014), Instituto Nacional de Ciência e Tecnologia em Nano-biofarmacêutica (INCT-Nanobiofar), FAPEMIG (CBB-APQ-00496-11 and CBB-APQ-00819-12), and CNPq (APQ-472090/2011-9 and APQ-482976/2012-8). MACF is a grant recipient of FAPEMIG/CAPES. EAFC, VNC, and AAGF are grant recipients of CNPq. Eduardo AF Coelho and André AG Faraco are co-senior authors of this stud

Recent updates and perspectives on approaches for the development of vaccines against visceral leishmaniasis

All rights reserved. Visceral leishmaniasis (VL) is one of the most important tropical diseases worldwide. Although chemotherapy has been widely used to treat this disease, problems related to the development of parasite resistance and side effects associated with the compounds used have been noted. Hence, alternative approaches for VL control are desirable. Some methods, such as vector control and culling of infected dogs, are insufficiently effective, with the latter not ethically recommended. The development of vaccines to prevent VL is a feasible and desirable measure for disease control, for example, some vaccines designed to protect dogs against VL have recently been brought to market. These vaccines are based on the combination of parasite fractions or recombinant proteins with adjuvants that are able to induce cellular immune responses, however, their partial efficacy and the absence of a vaccine to protect against human leishmaniasis underline the need for characterization of new vaccine candidates. This review presents recent advances in control measures for VL based on vaccine development, describing extensively studied antigens, as well as new antigenic proteins recently identified using immuno-proteomic techniquesThis work was supported by grants from Instituto Nacional de Ciência e Tecnologia em Nano-Biofarmacêutica, Rede Nanobiotec/Brasil-Universidade Federal de Uberlândia/CAPES, PRONEX-FAPEMIG (APQ-01019-09), FAPEMIG (CBB-APQ-00819-12 and CBB-APQ-01778-2014), and CNPq (APQ-482976/2012-8, APQ-488237/2013-0, and APQ-467640/2014-9). EAFC and LRG are recipients of the grant from CNPq. MACF is the recipient of grants from FAPEMIG/CAPE

Antigenicity and Protective Efficacy of a Leishmania Amastigote-specific Protein, Member of the Super-oxygenase Family, against Visceral Leishmaniasis

Background:The present study aimed to evaluate a hypothetical Leishmania amastigote-specific protein (LiHyp1), previously identified by an immunoproteomic approach performed in Leishmania infantum, which showed homology to the super-oxygenase gene family, attempting to select a new candidate antigen for specific serodiagnosis, as well as to compose a vaccine against VL.Methodology/Principal Findings:The LiHyp1 DNA sequence was cloned; the recombinant protein (rLiHyp1) was purified and evaluated for its antigenicity and immunogenicity. The rLiHyp1 protein was recognized by antibodies from sera of asymptomatic and symptomatic animals with canine visceral leishmaniasis (CVL), but presented no cross-reactivity with sera of dogs vaccinated with Leish-Tec, a Brazilian commercial vaccine; with Chagas' disease or healthy animals. In addition, the immunogenicity and protective efficacy of rLiHyp1 plus saponin was evaluated in BALB/c mice challenged subcutaneously with virulent L. infantum promastigotes. rLiHyp1 plus saponin vaccinated mice showed a high and specific production of IFN-γ, IL-12, and GM-CSF after in vitro stimulation with the recombinant protein. Immunized and infected mice, as compared to the control groups (saline and saponin), showed significant reductions in the number of parasites found in the liver, spleen, bone marrow, and in the paws' draining lymph nodes. Protection was associated with an IL-12-dependent production of IFN-γ, produced mainly by CD4 T cells. In these mice, a decrease in the parasite-mediated IL-4 and IL-10 response could also be observed.Conclusions/Significance:The present study showed that this Leishmania oxygenase amastigote-specific protein can be used for a more sensitive and specific serodiagnosis of asymptomatic and symptomatic CVL and, when combined with a Th1-type adjuvant, can also be employ as a candidate antigen to develop vaccines against VL. © 2013 Martins et al.UFMG (Edital 07/2012), Instituto Nacional de Ciencia e Tecnologia em Nanobiofarmace utica; Fundacao de Amparo a Pesquisa do Estado de Minas Gerais (FAPEMIG; CBB-APQ-02364-08, PPSUS/MS/CNPq/FAPEMIG/SES-MG/CBB-APQ-00356-10, and CBB-APQ-00496-11); Conselho Nacional de Desenvolvimento Cientıfico e Tecnologico (CNPq; APQ-472090/2011-9); Instituto Nacional de Ciencia e Tecnologia; FAPEMIG/CAPES; Ministerio de Ciencia e Innovacion (FIS/PI1100095)Peer Reviewe

Antileishmanial activity and mechanism of action from a purified fraction of Zingiber officinalis Roscoe against Leishmania amazonensis

In recent years, considerable attention has been given to identify new antileishmanial products derived from medicinal plants, although, to date, no new effective compound has been recently applied to treat leishmaniasis. In the present study, the antileishmanial activity of a water extract from Zingiber officinalis Roscoe (ginger) was investigated and a purified fraction, named F10, was identified as responsible by this biological activity. The chemical characterization performed for this fraction showed that it is mainly composed by flavonoids and saponins. The water extract and the F10 fraction presented IC50 values of 125.5 and 49.8 μg/mL, respectively. Their selectivity indexes (SI) were calculated and values were seven and 40 times higher, respectively, in relation to the value found for amphotericin B, which was used as a control. Additional studies were performed to evaluate the toxicity of these compounds in human red blood cells, besides of the production of nitrite, as an indicator of nitric oxide (NO), in treated and infected macrophages. The results showed that both F10 fraction and water extract were not toxic to human cells, and they were able to stimulate the nitrite production, with values of 13.6 and 5.4 μM, respectively, suggesting that their biological activity could be due to macrophages activation via NO production. In conclusion, the present study shows that a purified fraction from ginger could be evaluated in future works as a therapeutic alternative, on its own or in association with other drugs, to treat disease caused by L. amazonensis

Novel targeting using nanoparticles: an approach to the development of an effective anti-leishmanial drug-delivery system

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2014-05-09T12:07:23Z No. of bitstreams: 1 Ribeiro, T.G. Novel....pdf: 2225100 bytes, checksum: 9b5bdc3df6a233f4b33f9f0d6c514586 (MD5)Made available in DSpace on 2014-05-09T12:07:23Z (GMT). No. of bitstreams: 1 Ribeiro, T.G. Novel....pdf: 2225100 bytes, checksum: 9b5bdc3df6a233f4b33f9f0d6c514586 (MD5) Previous issue date: 2014Universidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências Farmacêuticas. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências da Saúde: Infectologia e Medicina Tropical. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências Farmacêuticas. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências Farmacêuticas. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências da Saúde: Infectologia e Medicina Tropical. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências da Saúde: Infectologia e Medicina Tropical. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Departamento de Patologia Clínica, COLTEC. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Departamento de Patologia Clínica, COLTEC. Belo Horizonte, MG, BrasilUniversidade Federal da Bahia. Programa de Pós-Graduação em Patologia Humana. Salvador, BA, Brasil / Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Faculdade de Medicina. Salvador, BA, Brasil / Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilUniversidade Federal da Bahia. Programa de Pós-Graduação em Patologia Humana. Salvador, BA, Brasil / Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Faculdade de Medicina. Salvador, BA, Brasil / Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências da Saúde: Infectologia e Medicina Tropical. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências Farmacêuticas. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências da Saúde: Infectologia e Medicina Tropical. Belo Horizonte, MG, Brasil / Universidade Federal de Minas Gerais. Departamento de Patologia Clínica, COLTEC. Belo Horizonte, MG, BrasilUniversidade Federal de Minas Gerais. Faculdade de Farmácia. Programa de Pós-Graduação em Ciências Farmacêuticas. Belo Horizonte, MG, Brasil / Universidade Federal de Minas Gerais. Faculdade de Farmácia. Departamento de Produtos Farmacêuticos. Belo Horizonte, MG, BrasilThe study reported here aimed to develop an optimized nanoparticle delivery system for amphotericin B (AmpB) using a polyelectrolyte complexation technique. For this, two oppositely charged polymers presenting anti-leishmanial activity – chitosan (Cs) and chondroitin sulfate (ChS) – were used: Cs as a positively charged polymer and ChS as a negatively charged polymer. The chitosan (NQ) nanoparticles, chitosan-chondroitin sulfate (NQC) nanoparticles, and chitosan-chondroitin sulfate-amphotericin B (NQC-AmpB) nanoparticles presented a mean particle size of 79, 104, and 136 nm, respectively; and a polydispersity index of 0.2. The measured zeta potential of the nanoparticles indicated a positive charge in their surface, while scanning and transmission electron microscopy revealed spherical nanoparticles with a smooth surface. Attenuated total reflectance-Fourier transform infrared spectroscopy analysis showed an electrostatic interaction between the polymers, whereas the release profile of AmpB from the NQC-AmpB nanoparticles showed a controlled release. In addition, the Cs; ChS; and NQ, NQC, and NQC-AmpB nanoparticles proved to be effective against promastigotes of Leishmania amazonensis and Leishmania chagasi, with a synergistic effect observed between Cs and ChS. Moreover, the applied NQ, NQC, and NQC-AmpB compounds demonstrated low toxicity in murine macrophages, as well as null hemolytic activity in type O+ human red blood cells. Pure AmpB demonstrated high toxicity in the macrophages. The results show that cells infected with L. amazonensis and later treated with Cs, ChS, NQ, NQC, NQC-AmpB nanoparticles, or pure AmpB presented with a significant reduction in parasite number in the order of 24%, 31%, 55%, 66%, 90%, and 89%, respectively. The data presented indicate that the engineered NQC-AmpB nanoparticles could potentially be used as an alternative therapy to treat leishmaniasis, mainly due its low toxicity to mammals’ cells

Correction: Antigenicity and Protective Efficacy of a Leishmania Amastigote-specific Protein, Member of the Super-oxygenase Family, against Visceral Leishmaniasis.

[This corrects the article on p. e2148 in vol. 7.]

Antigenicity and protective efficacy of a Leishmania amastigote-specific protein, member of the super-oxygenase family, against visceral leishmaniasis.

BackgroundThe present study aimed to evaluate a hypothetical Leishmania amastigote-specific protein (LiHyp1), previously identified by an immunoproteomic approach performed in Leishmania infantum, which showed homology to the super-oxygenase gene family, attempting to select a new candidate antigen for specific serodiagnosis, as well as to compose a vaccine against VL.Methodology/principal findingsThe LiHyp1 DNA sequence was cloned; the recombinant protein (rLiHyp1) was purified and evaluated for its antigenicity and immunogenicity. The rLiHyp1 protein was recognized by antibodies from sera of asymptomatic and symptomatic animals with canine visceral leishmaniasis (CVL), but presented no cross-reactivity with sera of dogs vaccinated with Leish-Tec, a Brazilian commercial vaccine; with Chagas' disease or healthy animals. In addition, the immunogenicity and protective efficacy of rLiHyp1 plus saponin was evaluated in BALB/c mice challenged subcutaneously with virulent L. infantum promastigotes. rLiHyp1 plus saponin vaccinated mice showed a high and specific production of IFN-γ, IL-12, and GM-CSF after in vitro stimulation with the recombinant protein. Immunized and infected mice, as compared to the control groups (saline and saponin), showed significant reductions in the number of parasites found in the liver, spleen, bone marrow, and in the paws' draining lymph nodes. Protection was associated with an IL-12-dependent production of IFN-γ, produced mainly by CD4 T cells. In these mice, a decrease in the parasite-mediated IL-4 and IL-10 response could also be observed.Conclusions/significanceThe present study showed that this Leishmania oxygenase amastigote-specific protein can be used for a more sensitive and specific serodiagnosis of asymptomatic and symptomatic CVL and, when combined with a Th1-type adjuvant, can also be employ as a candidate antigen to develop vaccines against VL

Proteins of <i>Leishmania infantum</i> amastigotes-like identified by an immunoproteomic approach.

<p>a) Sera samples of dogs with VL. b) Name of the identified protein and species: Lmj, <i>L. major</i>; Lbr, <i>L. braziliensis</i>; Li, <i>L. infantum</i>; Ld, <i>L. donovani</i>. c) Accession numbers according to NCBI. d) Experimental expected and predicted molecular weights (<i>M</i>r, in KDa). e) Experimental expected and predicted isoeletric points (p<i>I</i>).</p

Comparison of spots identified in protein extracts from promastigote and amastigote-like stages of <i>Leishmania infantum</i>.

<p>Diagrams show the percentage of protein spots identified in either individual or combined parasite stages. In A, the percentage of total proteins identified by asymptomatic (19/18%), symptomatic (64/62%), and a combination of both sera classes (21/20%). In B, the percentage of proteins from the promastigote stage identified by asymptomatic (10/20%), symptomatic (25/49%), and a combination of both sera classes (16/31%). In C, the percentage of proteins from amastigote-like stage identified by asymptomatic (9/17%), symptomatic (39/74%), and a combination of both sera classes (5/9%).</p