HUMAN ANTIBODIES TO CRYPTOCOCCUS NEOFORMANS

Cryptococcus neoformans meningitis occurs in 5-13% of individuals with aids in the United States. As an initial infection it is associated with the worst prognosis of all AID-associated infections. This project seeks to establish a potential role for antibodies in immunity to human cryptococcosis by demonstrating that anti-cryptococcal polysaccharide antibodies can be identified in individuals who have been expose to C. Neoformans. Although the traditional view is that immunity to cryptococcosis is cell mediated, numerous studies have demonstrated the importance of antibody-mediated processes. The presence of anti- cryptococcal antibodies in HIV positive individuals is difficult to predict since B cell defects that impair responses to encapsulated bacteria and polyclonal stimulation of B cells have both been noted in HIV infection. The basis of serologic studies to be performed is an antigen-based ELISA. Serotype reactivity of HIV positive and HIV negative individuals will be determined on purified cryptococcal polysaccharide-coated ELISA plates. Associations between HIV status and anti-cryptococcal antibody titer, antibody isotype, IgG subclass, and antibody specificity will be examined in adults and children. Murine anti-cryptococcal monoclonal antibodies will be used in competition experiments with human sera in order to determine whether human anti- cryptococcal antibodies recognize the same epitopes as existing protective murine antibodies. Affinity chromatography and isoelectric focusing will be utilized to further characterize human anti-cryptococcal antibody responses. Preliminary studies of human anti-cryptococcal polysaccharide antibodies with a rabbit anti-idiotypic reagent that recognizes a protective murine anti-cryptococcal monoclonal have demonstrated a cross-reactive idiotype in human and murine anti- cryptococcal polysaccharide antibody response. Human anti-idiotypic reagents will be generated in addition to human monoclonals to further prove the nature of the human anti-cryptococcal antibody response. Two monoclonal antibodies have already been isolated by EBV transformation of human peripheral lymphocytes. Nucleic acid sequences of the monoclonal will be determined to elucidate the molecular genetic structure of human anti-cryptococcal antibodies and to evaluate the nature of variable region gene element utilization in their derivation. Correlates between serologic parameters or antibody structure and the development or prognosis of C. neoformans meningitis in HIV positive individuals would represent a powerful rationale for the use of antibody proposal may provide new, urgently needed reagents for the therapy of cryptococcosis as will as new insights into human C. neoformans immunity.