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Glucosylceramide Administration as a Vaccination Strategy in Mouse Models of Cryptococcosis.

ABSTRACT: Cryptococcus neoformans is an opportunistic fungal pathogen and the causative agent of the disease cryptococcosis. Cryptococcosis is initiated as a pulmonary infection and in conditions of immune deficiency disseminates to the blood stream and central nervous system, resulting in life-threatening meningoencephalitis. A number of studies have focused on the development of a vaccine against Cryptococcus, primarily utilizing protein-conjugated components of the Cryptococcus polysaccharide capsule as antigen. However, there is currently no vaccine against Cryptococcus in the clinic. Previous studies have shown that the glycosphingolipid, glucosylceramide (GlcCer), is a virulence factor in C. neoformans and antibodies against this lipid inhibit fungal growth and cell division. In the present study, we have investigated the possibility of using GlcCer as a therapeutic agent against C. neoformans infections in mouse models of cryptococcosis. GlcCer purified from a non-pathogenic fungus, Candida utilis, was administered intraperitoneally, prior to infecting mice with a lethal dose of C. neoformans. GlcCer administration prevented the dissemination of C. neoformans from the lungs to the brain and led to 60% mouse survival. GlcCer administration did not cause hepatic injury and elicited an anti-GlcCer antibody response, which was observed independent of the route of administration and the strains of mouse. Taken together, our results suggest that fungal GlcCer can protect mice against lethal doses of C. neoformans infection and can provide a viable vaccination strategy against Cryptococcus.

SUBMITTER: Mor V

PROVIDER: S-EPMC4833283 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Glucosylceramide Administration as a Vaccination Strategy in Mouse Models of Cryptococcosis.

Mor Visesato V Farnoud Amir M AM Singh Ashutosh A Rella Antonella A Tanno Hiromasa H Ishii Keiko K Kawakami Kazuyoshi K Sato Toshiya T Del Poeta Maurizio M

PloS one 20160415 4

Cryptococcus neoformans is an opportunistic fungal pathogen and the causative agent of the disease cryptococcosis. Cryptococcosis is initiated as a pulmonary infection and in conditions of immune deficiency disseminates to the blood stream and central nervous system, resulting in life-threatening meningoencephalitis. A number of studies have focused on the development of a vaccine against Cryptococcus, primarily utilizing protein-conjugated components of the Cryptococcus polysaccharide capsule a ...[more]

PMID: 27082428

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Project description:UnlabelledA vaccine capable of protecting at-risk persons against infections due to Cryptococcus neoformans and Cryptococcus gattii could reduce the substantial global burden of human cryptococcosis. Vaccine development has been hampered though, by lack of knowledge as to which antigens are immunoprotective and the need for an effective vaccine delivery system. We made alkaline extracts from mutant cryptococcal strains that lacked capsule or chitosan. The extracts were then packaged into glucan particles (GPs), which are purified Saccharomyces cerevisiae cell walls composed primarily of β-1,3-glucans. Subcutaneous vaccination with the GP-based vaccines provided significant protection against subsequent pulmonary infection with highly virulent strains of C. neoformans and C. gattii. The alkaline extract derived from the acapsular strain was analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS), and the most abundant proteins were identified. Separation of the alkaline extract by size exclusion chromatography revealed fractions that conferred protection when loaded in GP-based vaccines. Robust Th1- and Th17-biased CD4(+) T cell recall responses were observed in the lungs of vaccinated and infected mice. Thus, our preclinical studies have indicated promising cryptococcal vaccine candidates in alkaline extracts delivered in GPs. Ongoing studies are directed at identifying the individual components of the extracts that confer protection and thus would be promising candidates for a human vaccine.ImportanceThe encapsulated yeast Cryptococcus neoformans and its closely related sister species, Cryptococcus gattii, are major causes of morbidity and mortality, particularly in immunocompromised persons. This study reports on the preclinical development of vaccines to protect at-risk populations from cryptococcosis. Antigens were extracted from Cryptococcus by treatment with an alkaline solution. The extracted antigens were then packaged into glucan particles, which are hollow yeast cell walls composed mainly of β-glucans. The glucan particle-based vaccines elicited robust T cell immune responses and protected mice from otherwise-lethal challenge with virulent strains of C. neoformans and C. gattii. The technology used for antigen extraction and subsequent loading into the glucan particle delivery system is relatively simple and can be applied to vaccine development against other pathogens.

Project description:Development of a vaccine to protect against cryptococcosis is a priority given the enormous global burden of disease in at-risk individuals. Using glucan particles (GPs) as a delivery system, we previously demonstrated that mice vaccinated with crude Cryptococcus-derived alkaline extracts were protected against lethal challenge with Cryptococcus neoformans and Cryptococcus gattii The goal of the present study was to identify protective protein antigens that could be used in a subunit vaccine. Using biased and unbiased approaches, six candidate antigens (Cda1, Cda2, Cda3, Fpd1, MP88, and Sod1) were selected, recombinantly expressed in Escherichia coli, purified, and loaded into GPs. Three mouse strains (C57BL/6, BALB/c, and DR4) were then vaccinated with the antigen-laden GPs, following which they received a pulmonary challenge with virulent C. neoformans and C. gattii strains. Four candidate vaccines (GP-Cda1, GP-Cda2, GP-Cda3, and GP-Sod1) afforded a significant survival advantage in at least one mouse model; some vaccine combinations provided added protection over that seen with either antigen alone. Vaccine-mediated protection against C. neoformans did not necessarily predict protection against C. gattii Vaccinated mice developed pulmonary inflammatory responses that effectively contained the infection; many surviving mice developed sterilizing immunity. Predicted T helper cell epitopes differed between mouse strains and in the degree to which they matched epitopes predicted in humans. Thus, we have discovered cryptococcal proteins that make promising candidate vaccine antigens. Protection varied depending on the mouse strain and cryptococcal species, suggesting that a successful human subunit vaccine will need to contain multiple antigens, including ones that are species specific.IMPORTANCE The encapsulated fungi Cryptococcus neoformans and Cryptococcus gattii are responsible for nearly 200,000 deaths annually, mostly in immunocompromised individuals. An effective vaccine could substantially reduce the burden of cryptococcosis. However, a major gap in cryptococcal vaccine development has been the discovery of protective antigens to use in vaccines. Here, six cryptococcal proteins with potential as vaccine antigens were expressed recombinantly and purified. Mice were then vaccinated with glucan particle preparations containing each antigen. Of the six candidate vaccines, four protected mice from a lethal cryptococcal challenge. However, the degree of protection varied as a function of mouse strain and cryptococcal species. These preclinical studies identify cryptococcal proteins that could serve as candidate vaccine antigens and provide a proof of principle regarding the feasibility of protein antigen-based vaccines to protect against cryptococcosis.

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Glucosylceramide Administration as a Vaccination Strategy in Mouse Models of Cryptococcosis.

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Glucosylceramide Administration as a Vaccination Strategy in Mouse Models of Cryptococcosis.

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