Publications by authors named "Maria Grazia Aruta"

6 Publications

  • Page 1 of 1

Effect of O-Antigen Chain Length Regulation on the Immunogenicity of and Generalized Modules for Membrane Antigens (GMMA).

Int J Mol Sci 2021 Jan 28;22(3). Epub 2021 Jan 28.

GSK Vaccines Institute for Global Health (GVGH) s.r.l, Via Fiorentina 1, 53100 Siena, Italy.

Recently, generalized modules for membrane antigens (GMMA) technology has been proposed as an alternative approach to traditional glycoconjugate vaccines for O-antigen delivery. Saccharide length is a well-known parameter that can impact the immune response induced by glycoconjugates both in terms of magnitude and quality. However, the criticality of O-antigen length on the immune response induced by GMMA-based vaccines has not been fully elucidated. Here, and GMMA-producing strains were further mutated in order to display homogeneous polysaccharide populations of different sizes on a GMMA surface. Resulting GMMA were compared in mice immunization studies. Athymic nude mice were also used to investigate the involvement of T-cells in the immune response elicited. In contrast with what has been reported for traditional glycoconjugate vaccines and independent of the pathogen and the sugar structural characteristics, O-antigen length did not result in being a critical parameter for GMMA immunogenicity. This work supports the identification of critical quality attributes to optimize GMMA vaccine design and improve vaccine efficacy and gives insights on the nature of the immune response induced by GMMA.
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http://dx.doi.org/10.3390/ijms22031309DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7865430PMC
January 2021

Dissecting the contribution of O-Antigen and proteins to the immunogenicity of Shigella sonnei generalized modules for membrane antigens (GMMA).

Sci Rep 2021 Jan 13;11(1):906. Epub 2021 Jan 13.

GSK Vaccines Institute for Global Health (GVGH), via Fiorentina 1, 53100, Siena, Italy.

GMMA are exosomes released from engineered Gram-negative bacteria resembling the composition of outer membranes. We applied the GMMA technology for the development of an O-Antigen (OAg) based vaccine against Shigella sonnei, the most epidemiologically relevant cause of shigellosis. S. sonnei OAg has been identified as a key antigen for protective immunity, and GMMA are able to induce anti-OAg-specific IgG response in animal models and healthy adults. The contribution of protein-specific antibodies induced upon vaccination with GMMA has never been fully elucidated. Anti-protein antibodies are induced in mice upon immunization with either OAg-negative and OAg-positive GMMA. Here we demonstrated that OAg chains shield the bacteria from anti-protein antibody binding and therefore anti-OAg antibodies were the main drivers of bactericidal activity against OAg-positive bacteria. Interestingly, antibodies that are not targeting the OAg are functional against OAg-negative bacteria. The immunodominant protein antigens were identified by proteomic analysis. Our study confirms a critical role of the OAg on the immune response induced by S. sonnei GMMA. However, little is known about OAg length and density regulation during infection and, therefore, protein exposure. Hence, the presence of protein antigens on S. sonnei GMMA represents an added value for GMMA vaccines compared to other OAg-based formulations.
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http://dx.doi.org/10.1038/s41598-020-80421-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7806729PMC
January 2021

Characterization of Competitive ELISA and Formulated Alhydrogel Competitive ELISA (FAcE) for Direct Quantification of Active Ingredients in GMMA-Based Vaccines.

Methods Protoc 2020 Aug 31;3(3). Epub 2020 Aug 31.

GSK Vaccines Institute for Global Health s.r.l (GVGH), 53100 Siena, Italy.

Generalized modules for membrane antigens (GMMA) represent a technology particularly attractive for designing affordable vaccines against Gram-negative bacteria. We explored such technology for the development of O-antigen-based vaccines against and nontyphoidal . Adsorption of GMMA on Alhydrogel was required for abrogation of pyrogenicity in rabbits, and GMMA on Alhydrogel was well tolerated and immunogenic in humans. Quantification of key antigens in formulated vaccines was fundamental for release and to check stability overtime. Traditionally, the direct quantification of antigens adsorbed on aluminum salts has been challenging, and the quantification of each active ingredient in multicomponent formulated vaccines has been even more complicated. To directly quantify each active ingredient and unbound drug substances in formulated vaccines, we developed the Formulated Alhydrogel competitive ELISA (FAcE) and the competitive ELISA method, respectively. The methods were both fully characterized, assessing specificity, repeatability, intermediate precision, and accuracy, for OAg quantification, both in a single component or multicomponent GMMA formulation also containing GMMA. The developed immunological methods allowed us to fully characterize GMMA drug products, supporting their preclinical and clinical development. The same methods, already extended to GMMA from nontyphoidal and , could be potentially extended to any antigen formulated on Alhydrogel.
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http://dx.doi.org/10.3390/mps3030062DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7563494PMC
August 2020

GMMA and Glycoconjugate Approaches Compared in Mice for the Development of a Vaccine against Serotype 6.

Vaccines (Basel) 2020 Apr 3;8(2). Epub 2020 Apr 3.

GSK Vaccines Institute for Global Health (GVGH) S.r.l., via Fiorentina 1, 53100 Siena, Italy.

infections are one of the top causes of diarrhea throughout the world, with being predominant in developing countries. Currently, no vaccines are widely available and increasing levels of multidrug-resistance make a high priority for vaccine development. The serotype-specific O-antigen moiety of lipopolysaccharide has been recognized as a key target for protective immunity, and many O-antigen based candidate vaccines are in development. Recently, the Generalized Modules for Membrane Antigens (GMMA) technology has been proposed as an alternative approach to traditional glycoconjugate vaccines for O-antigen delivery. Here, these two technologies are compared for a vaccine against serotype 6. Genetic strategies for GMMA production, conjugation approaches for linkage of the O-antigen to CRM carrier protein, and a large panel of analytical methods for full vaccine characterization have been put in place. In a head-to-head immunogenicity study in mice, GMMA induced higher anti-O-antigen IgG than glycoconjugate administered without Alhydrogel. When formulated on Alhydrogel, GMMA and glycoconjugate elicited similar levels of persistent anti-O-antigen IgG with bactericidal activity. Glycoconjugates are a well-established bacterial vaccine approach, but can be costly, particularly when multicomponent preparations are required. With similar immunogenicity and a simpler manufacturing process, GMMA are a promising strategy for the development of a vaccine against .
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http://dx.doi.org/10.3390/vaccines8020160DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349896PMC
April 2020

Contribution of factor H-Binding protein sequence to the cross-reactivity of meningococcal native outer membrane vesicle vaccines with over-expressed fHbp variant group 1.

PLoS One 2017 25;12(7):e0181508. Epub 2017 Jul 25.

GSK Vaccines Institute for Global Health (GVGH), Siena, Italy.

Factor H-binding protein (fHbp) is an important meningococcal vaccine antigen. Native outer membrane vesicles with over-expressed fHbp (NOMV OE fHbp) have been shown to induce antibodies with broader functional activity than recombinant fHbp (rfHbp). Improved understanding of this broad coverage would facilitate rational vaccine design. We performed a pair-wise analysis of 48 surface-exposed amino acids involved in interacting with factor H, among 383 fHbp variant group 1 sequences. We generated isogenic NOMV-producing meningococcal strains from an African serogroup W isolate, each over-expressing one of four fHbp variant group 1 sequences (ID 1, 5, 9, or 74), including those most common among invasive African meningococcal isolates. Mice were immunised with each NOMV, and sera tested for IgG levels against each of the rfHbp ID and for ability to kill a panel of heterologous meningococcal isolates. At the fH-binding site, ID pairs differed by a maximum of 13 (27%) amino acids. ID 9 shared an amino acid sequence common to 83 ID types. The selected ID types differed by up to 6 amino acids, in the fH-binding site. All NOMV and rfHbp induced high IgG levels against each rfHbp. Serum killing from mice immunised with rfHbp was generally less efficient and more restricted compared to NOMV, which induced antibodies that killed most meningococci tested, with decreased stringency for ID type differences. Breadth of killing was mostly due to anti-fHbp antibodies, with some restriction according to ID type sequence differences. Nevertheless, under our experimental conditions, no relationship between antibody cross-reactivity and variation fH-binding site sequence was identified. NOMV over-expressing different fHbp IDs belonging to variant group 1 induce antibodies with fine specificities against fHbp, and ability to kill broadly meningococci expressing heterologous fHbp IDs. The work reinforces that meningococcal NOMV with OE fHbp is a promising vaccine strategy, and provides a basis for rational selection of antigen sequence types for over-expression on NOMV.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0181508PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5526518PMC
October 2017

Efficacy of piroxicam plus cisplatin-loaded PLGA nanoparticles in inducing apoptosis in mesothelioma cells.

Pharm Res 2015 Feb 5;32(2):362-74. Epub 2014 Aug 5.

Department of Experimental Medicine, Second University of Naples, Naples, Italy.

Purpose: Combined treatment based on cisplatin-loaded Poly(D,L-lactic-co-glicolic)acid (PLGA) nanoparticles (NP-C) plus the NSAID piroxicam was used as novel treatment for mesothelioma to reduce side effects related to cisplatin toxicity.

Methods: PLGA nanoparticles were prepared by double emulsion solvent evaporation method. Particle size, drug release profile and in vitro cellular uptake were characterized by TEM, DLS, LC/MS and fluorescence microscopy. MSTO-211H cell line was used to analyse NP-C biological efficacy by FACS and protein analysis.

Results: Cisplatin was encapsulated in 197 nm PLGA nanoparticles with 8.2% drug loading efficiency and 47% encapsulation efficiency. Cisplatin delivery from nanoparticles reaches 80% of total encapsulated drug in 14 days following a triphasic trend. PLGA nanoparticles in MSTO-211H cells were localized in the perinuclear space NP-C in combination with piroxicam induced apoptosis using a final cisplatin concentration 1.75 fold less than free drug. Delivered cisplatin cooperated with piroxicam in modulating cell cycle regulators as caspase-3, p53 and p21.

Conclusions: Cisplatin loaded PLGA nanoparticles plus piroxicam showed a good efficacy in exerting cytotoxic activity and inducing the same molecular apoptotic effects of the free drugs. Sustained cisplatin release allowed to use less amount of drug, decreasing toxic side effects. This novel approach could represent a new strategy for mesothelioma treatment.
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http://dx.doi.org/10.1007/s11095-014-1467-3DOI Listing
February 2015