Proteome-wide Discovery of Diagnostic Targets for Coccidioidomycosis

PROJECT SUMMARY Antibody mediated immunity has long been considered irrelevant to the disease coccidioidomycosis (CM) – an endemic fungal infection in the southwestern United States that can cause severe morbidity and mortality. However, new evidence has revealed an important and previously unappreciated role for antibodies in other fungal infections, suggesting that the prevailing paradigm is incomplete. Since antibodies are an attractive target for diagnostics, an improved understanding of their potential in the setting of CM is crucial; but this remains a significantly under-developed area of investigation. We hypothesize that antibodies play a role in CM disease course and can inform the development of improved and rapid diagnostic assays. In this proposal, we will test the following subsidiary hypotheses: 1), that there are undiscovered immunogenic protein targets within the Coccidioides proteome; 2), that these antigenic targets will group into conserved antigen recognition patterns across patients; and 3), that genomic analysis of B cells isolated from CM patients will identify novel monoclonal antibodies which target known and previously unknown Coccidioides proteins. To test these hypotheses, we propose to apply two innovative technologies; first we will identify novel CM antigens at proteome-wide scale using PepSeq, a highly-multiplexed, epitope-resolved serological assay. Secondly, we will combine newly-identified antigens with known antigens to build probesets for the multiplexed analysis of 100,000s of single B cells from CM patients. This will allow us to identify antigen/epitope-specific IgH/L chain sequences and generate and validate a diverse panel of novel monoclonal antibodies. Such antibodies will have potential as therapeutic and/or diagnostic tools, to be developed in future studies.