Evaluation of Cryptococcus neoformans galactoxylomannan-protein conjugate as vaccine candidate against murine cryptococcosis

Galactoxylomannan (GaIXM) is a complex polysaccharide produced by the human pathogenic fungus Cryptococcus neoformans that mediates profound immunological derangements in murine models. GaIXM is essentially non-immunogenic and produces immune paralysis in mice. Previous studies have attempted to enhance immunogenicity by conjugating GaIXM to a protein carrier, but only transient antibody responses were elicited. Here we report the generation of two GaIXM conjugates with bovine serum albumin (BSA) and protective antigen (PA) of Bacillus anthracis, respectively, using 1-cyano-4-dimethylaminopyridinium tetrafiuoroborate (CDAP) as the cyanylating reagent. Both conjugates induced potent and sustained antibody responses as detected by both cross antigen-based and CovaLink direct ELISAs. We confirmed the specificity of the response to GaIXM by inhibition ELISA and immunofluorescence. The isotype composition analysis revealed that IgG and IgM were abundant in the immune sera against GaIXM, consistent with the induction of a T cell-dependent response. IgG1 was the predominant IgG subclass against GaIXM, while immunization with Quil A as adjuvant elicited a significantly higher production of IgG2a than with Freund's adjuvant. Immune sera were not opsonic for C neoformans and there was no survival difference between immune and non-immune mice challenged with C neoformans. These results demonstrated the effectiveness of the GaIXM-protein conjugate to induce robust immune responses although no evidence was obtained that such responses contributed to host defense. (C) 2011 Elsevier Ltd. All rights reserved.