Functional Antibody Repertoire Against S. aureus Leukocidins after Invasive Human Infection

PROJECT SUMMARY / ABSTRACTStaphylococcus aureus is now the most common invasive bacterial pathogen in children in the US, andantibiotic resistance rates continue to increase. Novel targets and approaches are needed if a safe andeffective S. aureus vaccine is to become a reality. Optimal targets of intervention against S. aureus, a highlyhuman-specific pathogen, may be identified most effectively by defining the antigens that contribute to invasivehuman infections. LukAB, a secreted leukotoxin, is produced by S. aureus in the setting of invasive humandisease and is essential for immune evasion by the pathogen in a variety of models. We have purified a seriesof potently neutralizing human monoclonal antibodies (mAbs) targeting LukAB, which have broad neutralizingcapacity, distinct binding sites and mechanisms of activity, and efficacy in a murine model of sepsis. Theoverall goals of this proposal are twofold: first, to test the hypothesis that key aspects of the humananti-leukocidin adaptive response include both mechanisms of toxin neutralization and the function ofnon-neutralizing antibodies targeting these toxins; second, to test the hypothesis that a selectedcombination of mAbs with distinct functions will most potently inhibit toxin-mediated staphylococcalimmune evasion. Careful validation of the anti-leukocidin antibody response in rigorous models will allow forthe evaluation of human mAbs as candidate agents of intervention against S. aureus. The proposed studieswill define the diversity of the naturally occurring antibody repertoire to S. aureus leukocidins following naturalhuman infection, given their importance in virulence and promise as targets of intervention. This will beaccomplished by: 1) Characterizing the diversity and functional capacity of naturally occurring humanantibodies targeting the staphylococcal leukocidins following invasive pediatric infection; 2) Elucidating the roleof non-neutralizing antibodies specific to S. aureus leukocidins, including antibody-dependent complementdeposition and phagocytosis; and 3) Determining the critical components of the anti-leukocidin adaptiveresponse against S. aureus in human blood. We will leverage our existing workflow for the purification andcharacterization of neutralizing mAbs from prospectively enrolled children with invasive and non-invasive S.aureus infections at a major tertiary care children?s hospital. A panel of human mAbs will be created to allowthe investigation of diverse mechanisms of neutralization, cross-toxin activity, and other important functions.The proposed aims will define, for the first time, the breadth of mechanisms by which the human host responsetargets and neutralizes S. aureus leukocidins during invasive human disease. Together these studies addressa critical need for novel anti-S. aureus strategies. This work is designed to produce rationally selectedcandidates for anti-staphylococcal interventions based on our knowledge of important antigens expressedduring human disease, obtained from children with S. aureus infections, to be tested for efficacy in humanizedanimal models in immediately subsequent work.