5 results on '"Miyaji, Eliane Namie"'
Search Results
2. Novel method for production and purification of untagged pneumococcal surface protein A from clade 1.
- Author
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da Costa Rodrigues, Tasson, Zorzete, Patricia, Miyaji, Eliane Namie, and Gonçalves, Viviane Maimoni
- Subjects
STREPTOCOCCUS pneumoniae ,PRODUCTION methods ,AMINO acid sequence ,PNEUMOCOCCAL vaccines ,PROTEINS ,BIOMASS production - Abstract
Streptococcus pneumoniae can cause diseases with high mortality and morbidity. The licensed vaccines are based on capsular polysaccharides and induce antibodies with low cross reactivity, leading to restricted coverage of serotypes. For surpassing this limitation, new pneumococcal vaccines are needed for induction of broader protection. One important candidate is the pneumococcal surface protein A (PspA), which can be classified in 6 clades and 3 families. We have reported an efficient process for production and purification of untagged recombinant PspA from clade 4 (PspA4Pro). We now aim to obtain a highly pure recombinant PspA from clade 1 (PspA1) to be included, together with PspA4Pro, in a vaccine formulation to broaden response against pneumococci. The vector pET28a-pspA1 was constructed and used to transform Escherichia coli BL21(DE3) strain. One clone with high production of PspA1 was selected and adapted to high-density fermentation (HDF) medium. After biomass production in 6 L HDF using a bioreactor, the purification was defined after testing 3 protocols. During the batch bioreactor cultivation, plasmid stability remained above 90% and acetate formation was not detected. The final protein purification process included treatment with a cationic detergent after lysis, anion exchange chromatography, cryoprecipitation, cation exchange chromatography, and multimodal chromatography. The final purification process showed PspA1 purity of 93% with low endotoxin content and an overall recovery above 20%. The novel established process can be easily scaled-up and proved to be efficient to obtain a highly pure untagged PspA1 for inclusion in vaccine formulations. Key points: • Purification strategy for recombinant PspA1 from Streptococcus pneumoniae • Downstream processing for untagged protein antigens, the case of PspA1 • Purification strategy for PspA variants relies on buried amino acids in their sequences [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
3. Pneumococcal Surface Protein A-Hybrid Nanoparticles Protect Mice from Lethal Challenge after Mucosal Immunization Targeting the Lungs.
- Author
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Figueiredo, Douglas Borges de, Kaneko, Kan, Rodrigues, Tasson da Costa, MacLoughlin, Ronan, Miyaji, Eliane Namie, Saleem, Imran, and Gonçalves, Viviane Maimoni
- Subjects
LUNGS ,LACTOFERRIN ,PNEUMOCOCCAL vaccines ,ANTIGEN presentation ,DENDRITIC cells ,MICE ,IMMUNIZATION - Abstract
Pneumococcal disease remains a global burden, with current conjugated vaccines offering protection against the common serotype strains. However, there are over 100 serotype strains, and serotype replacement is now being observed, which reduces the effectiveness of the current vaccines. Pneumococcal surface protein A (PspA) has been investigated as a candidate for new serotype-independent pneumococcal vaccines, but requires adjuvants and/or delivery systems to improve protection. Polymeric nanoparticles (NPs) are biocompatible and, besides the antigen, can incorporate mucoadhesive and adjuvant substances such as chitosans, which improve antigen presentation at mucosal surfaces. This work aimed to define the optimal NP formulation to deliver PspA into the lungs and protect mice against lethal challenge. We prepared poly(glycerol-adipate-co-ω-pentadecalactone) (PGA-co-PDL) and poly(lactic-co-glycolic acid) (PLGA) NPs using an emulsion/solvent evaporation method, incorporating chitosan hydrochloride (HCl-CS) or carboxymethyl chitosan (CM-CS) as hybrid NPs with encapsulated or adsorbed PspA. We investigated the physicochemical properties of NPs, together with the PspA integrity and biological activity. Furthermore, their ability to activate dendritic cells in vitro was evaluated, followed by mucosal immunization targeting mouse lungs. PGA-co-PDL/HCl-CS (291 nm) or CM-CS (281 nm) NPs produced smaller sizes compared to PLGA/HCl-CS (310 nm) or CM-CS (299 nm) NPs. Moreover, NPs formulated with HCl-CS possessed a positive charge (PGA-co-PDL +17 mV, PLGA + 13 mV) compared to those formulated with CM-CS (PGA-co-PDL −20 mV, PLGA −40 mV). PspA released from NPs formulated with HCl-CS preserved the integrity and biological activity, but CM-CS affected PspA binding to lactoferrin and antibody recognition. PspA adsorbed in PGA-co-PDL/HCl-CS NPs stimulated CD80+ and CD86+ cells, but this was lower compared to when PspA was encapsulated in PLGA/HCl-CS NPs, which also stimulated CD40+ and MHC II (I-A/I-E)+ cells. Despite no differences in IgG being observed between immunized animals, PGA-co-PDL/HCl-CS/adsorbed-PspA protected 83% of mice after lethal pneumococcal challenge, while 100% of mice immunized with PLGA/HCl-CS/encapsulated-PspA were protected. Therefore, this formulation is a promising vaccine strategy, which has beneficial properties for mucosal immunization and could potentially provide serotype-independent protection. [ABSTRACT FROM AUTHOR]
- Published
- 2022
- Full Text
- View/download PDF
4. Progress in mucosal immunization for protection against pneumococcal pneumonia.
- Author
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Gonçalves, Viviane Maimoni, Kaneko, Kan, Solórzano, Carla, MacLoughlin, Ronan, Saleem, Imran, and Miyaji, Eliane Namie
- Subjects
PNEUMOCOCCAL pneumonia ,RESPIRATORY infections ,IMMUNIZATION ,PNEUMOCOCCAL vaccines ,LUNG infections - Abstract
Introduction: Lower respiratory tract infections are the fourth cause of death worldwide and pneumococcus is the leading cause of pneumonia. Nonetheless, existing pneumococcal vaccines are less effective against pneumonia than invasive diseases and serotype replacement is a major concern. Protein antigens could induce serotype-independent protection, and mucosal immunization could offer local and systemic immune responses and induce protection against pneumococcal colonization and lung infection. Areas covered: Immunity induced in the experimental human pneumococcal carriage model, approaches to address the physiological barriers to mucosal immunization and improve delivery of the vaccine antigens, different strategies already tested for pneumococcal mucosal vaccination, including live recombinant bacteria, nanoparticles, bacterium-like particles, and nanogels as well as, nasal, pulmonary, sublingual and oral routes of vaccination. Expert opinion: The most promising delivery systems are based on nanoparticles, bacterial-like particles or nanogels, which possess greater immunogenicity than the antigen alone and are considered safer than approaches based on living cells or toxoids. These particles can protect the antigen from degradation, eliminating the refrigeration need during storage and allowing the manufacture of dry powder formulations. They can also increase antigen uptake, control release of antigen and trigger innate immune responses. [ABSTRACT FROM AUTHOR]
- Published
- 2019
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5. Pertussis toxin improves immune responses to a combined pneumococcal antigen and leads to enhanced protection against Streptococcus pneumoniae.
- Author
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Salcedo-Rivillas C, Debrie AS, Miyaji EN, Ferreira JM Jr, Raw I, Locht C, Ho PL, Mielcarek N, and Oliveira ML
- Subjects
- Adjuvants, Immunologic, Administration, Intranasal, Animals, Antibodies, Bacterial immunology, Antigens, Bacterial immunology, B-Lymphocytes immunology, Bordetella pertussis immunology, Complement System Proteins immunology, Immunization, Passive, Immunoglobulin G administration & dosage, Immunoglobulin G immunology, Interleukin-17 metabolism, Leukocytes immunology, Mice, Mice, Inbred BALB C, Mice, Inbred C57BL, Mice, Knockout, Pertussis Vaccine immunology, Pneumococcal Infections prevention & control, Receptors, Immunologic immunology, Recombinant Proteins immunology, Streptococcal Vaccines immunology, Vaccination, Bacterial Proteins immunology, Pertussis Toxin immunology, Pneumococcal Infections immunology, Pneumococcal Vaccines immunology, Streptococcus pneumoniae immunology
- Abstract
Pneumococcal surface protein A (PspA) is a candidate antigen for the composition of protein-based vaccines against Streptococcus pneumoniae. While searching for efficient adjuvants for PspA-based vaccines, our group has described the potential of combining PspA with the whole-cell pertussis vaccine (wP). When given to mice through the nasal route, a formulation composed of PspA from clade 5 (PspA5) and wP (PspA5-wP) induced high levels of antibodies and protection against challenges with different pneumococcal strains. PspA5-wP also induced the secretion of interleukin 17 (IL-17) by splenocytes and the infiltration of leukocytes in the lungs after challenge. Here, we show that protection against a pneumococcal invasive challenge was completely abrogated in μMT(-/-) mice, which are deficient in the maturation of B cells, illustrating the importance of antibodies in the survival elicited by the PspA5-wP vaccine. Moreover, passive immunization showed that IgG purified from the sera of mice immunized with PspA5-wP conferred significant protection to naive mice, whereas the respective F(ab')2 did not. Additionally, in vivo depletion of complement abolished protection against the pneumococcal challenge. The combination of PspA5 with wild-type or mutant Bordetella pertussis strains or with purified components showed that the pertussis toxin (PT)-containing formulations induced the highest levels of antibodies and protection. This suggests that the adjuvant activity of wP in the PspA5 model is mediated at least in part by PT. The sera from mice immunized with such formulations displayed high IgG binding and induction of complement deposition on the pneumococcal surface in vitro, which is consistent with the in vivo results., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)
- Published
- 2014
- Full Text
- View/download PDF
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