Specht, C. A. | bioRxiv Science

bioRxiv2026

Identification of Cryptococcus neoformans capsule-associated protein antigens that mediate B cell-dependent vaccine protection in mice and elicit antibody responses in humans with cryptococcosis

Infection with the encapsulated fungal pathogen Cryptococcus neoformans causes substantial global disease burden in persons with compromised T cell function. Previously, we demonstrated that vaccines containing adjuvanted recombinant cryptococcal proteins protected mice against lethal infection by mechanisms dependent on CD4+ T cells. For the present study, we hypothesized that vaccines containing protein antigens naturally located on the surface of the cryptococcal capsule would contribute protective B cell responses. We found that B cell-deficient mice exhibited reduced survival following vaccination with any of four capsule-associated antigens: Blp4, Cpd1, Lhc1, or Krp1. To further elucidate the role of antibodies in protection, we tested serum from mice vaccinated with a live-attenuated strain (cda123) and then challenged with a virulent C. neoformans strain for IgG responses to a panel of 26 cryptococcal recombinant proteins. Strong reactivity was detected against Blp4, which we focused on because it is capsule-associated and protects mice when administered in adjuvanted subunit vaccines, by mechanisms partly dependent on B cells. Blp4 antibodies were consistently present in surviving mice, including those vaccinated with unrelated proteins or those challenged with diverse clinical isolates. Additionally, serum from Blp4-vaccinated mice promoted macrophage opsonophagocytosis of C. neoformans. Finally, IgG antibodies against Blp4 were detected in 20 of 21 serum samples from patients who survived cryptococcosis, compared with only 4 of 14 samples from healthy controls. Together, these data identify Blp4 as an antigen that elicits protective B cell responses and is a strong candidate for inclusion in subunit cryptococcal vaccines.

immunology↗

bioRxiv2024

Protection against experimental cryptococcosis elicited by Cationic Adjuvant Formulation 01-adjuvanted subunit vaccines

The fungal infection, cryptococcosis, is responsible for >100,000 deaths annually. No licensed vaccines are available. We explored the efficacy and immune responses of subunit cryptococcal vaccines adjuvanted with Cationic Adjuvant Formulation 01 (CAF01). CAF01 promotes humoral and T helper (Th) 1 and Th17 immune responses and has been safely used in human vaccine trials. Four subcutaneous vaccines, each containing single recombinant Cryptococcus neoformans protein antigens, partially protected mice from experimental cryptococcosis. Protection increased, up to 100%, in mice that received bivalent and quadrivalent vaccine formulations. Vaccinated mice that received a pulmonary challenge with C. neoformans had an influx of leukocytes into the lung including robust numbers of polyfunctional CD4+ T cells which produced Interferon gamma (IFN{gamma}), tumor necrosis factor alpha (TNF), and interleukin (IL)-17 upon ex vivo antigenic stimulation. Cytokine-producing lung CD8+ T cells were also found, albeit in lesser numbers. A significant, durable IFN{gamma} response was observed in the lungs, spleen, and blood. Moreover, IFN{gamma} secretion following ex vivo stimulation directly correlated with fungal clearance in the lungs. Thus, we have developed multivalent cryptococcal vaccines which protect mice from experimental cryptococcosis using an adjuvant which has been safely tested in humans. These preclinical studies suggest a path towards human cryptococcal vaccine trials. Author summaryCryptococcosis is a fungal infection that poses great challenges to public health, especially in resource-limited regions with high HIV prevalence. Despite the urgent need, no licensed vaccines are currently available. In this study, we used a lethal mouse model of cryptococcosis to explore protection and immune responses elicited by vaccines consisting of recombinant cryptococcal proteins formulated with CAF01, an adjuvant that has an established safety and immunogenicity profile in human clinical vaccine trials. We discovered that while vaccines containing a single protein partially protected mouse strains, the protection was greatly augmented when the mice received vaccines formulated with multiple antigens. The lungs of vaccinated and infected mice had a robust influx of CD4+ T cells, many of which made the cytokines IFN{gamma} and IL-17 when stimulated ex vivo. Moreover, we found the production of IFN{gamma} directly correlated with clearance of fungi from the lungs. Cytotoxic CD8+ T cell responses were also observed, albeit in lesser numbers. Our promising findings from this preclinical research paves the way for future human cryptococcal vaccine trials.

immunology↗