Biological and Clinical Impact of Cryptococcal Extralcellular Vesicles

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Cryptococcus neoformans is a major fungal pathogen for individuals with impaired immunity, including those with advanced HIV infection, organ transplants, and on immunosuppressive regimens. Furthermore, there is increasing evidence that this fungus can establish latent infection in humans that could have profound consequences for the development of other chronic diseases, including possibly asthma. C, neoformans has several well-characterized virulence factors, among which a polysaccharide capsule is considered to be the most important. The capsule is composed of at least three components known as Glucuronoxylomannan (GXM), galactoxylomannan (GalXM) and highly mannosylated proteins known as mannoproteins. Antibody responses to the capsular GXM elicit protective and non-protedive antibodies. Given the seriousness of cryptococcal infections there has been great interest in harnessing humoral immunity for therapy and prevention of disease. A monoclonal antibody (mAb) has completed preliminary clinical studies and continues in development. Immunization with GXM conjugated to protein carriers elicits protective antibodies. The mechanism of antibody action Is multifactorial and includes opsonization, modulation ofthe inflammatory response, and abrogation of GXM release from yeast cells. Remari^ably, protective and non- protective mAbs can be distinguished by their ability to affect GXM release and block biofilm formation in vitro. Although much is now known about the mechanism of antibody action in vivo, and in vitro, the molecular nature ofthe GXM epitopes recognized by protective and non-protective mAbs is unknown. Furthermore, the mechanisms of action of antibodies at the level of the yeast cell that abrogate polysaccharide release are not understood. This application proposes to continue the study ofthe interaction of antibodies with the capsule of C. neofomnans. In contrast to the prior funding cycles when the effort was focused on molecular analysis of the antibody molecule, this proposal refocuses the research program on the polysaccharide antigen and the capsule. In addition to continuing to study antibodies to GXM this application proposes to explore the role of GalXM in capsule structure. Three specific Aims are proposed: 1) To define the polysaccharide molecular structure(s) that bind protective and non-protective mAbs; 2) To determine the mechanism and consequences of antibody-mediated changes in C. neoformans metabolism: 3) To determine the protective efficacy of GalXM-congugate vaccines and GatXM-binding antibodies in host defense. RELEVANCE (See instructions): The research program is focused on a fungal pathogen known as Cryptococcus neoformans that is a major problem for patients with AIDS. Over a million people woridwide suffer life-threatening infections with this fungus and as many as 600,000 die each year in Africa alone. This fungus has a capsule that allows it to cause disease. Hence, understanding this structure is important for knowing how the fungus protects itself against the immune system. We hope that this information will lead to new immune therapies and vaccines.