Human Antibody Cross-Reactivity in Non-Polio Enteroviruses

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Project Summary Manifestations of enterovirus infections are variable, with several severe illnesses such as meningitis, acute flaccid myelitis (AFM), myocarditis, and neonatal sepsis. Poliovirus vaccines are near perfect at preventing severe illness by generating strong, circulating antibody responses. However, the polioviruses represent only three of the more than 100 enteroviruses capable of causing human disease. Recently approved vaccines for enterovirus A71 are demonstrating that other enterovirus diseases, in this case hand, foot, and mouth disease, can be prevented by vaccination as well. Our previous work shows that monoclonal antibodies that cross-react to all clades of enterovirus D68 can protect infected mice from AFM-like disease, and even treat disease after onset. Rather than try to make 100 vaccines to cover each enterovirus, we propose to understand the qualities of the human antibody response that could provide cross-reactive immunity across many enteroviruses. With this knowledge, immunogens can be designed that preserve the B cell epitopes that stimulate cross-reactive antibodies. We first aim to determine the degree of cross-reactivity of enterovirus antibody responses at a B cell level. We will complement seroepidemiology studies of hundreds of healthy adults, looking to see what proportion of them have naturally made antibody responses to the four species of enterovirus, with studies of stimulated memory B cells from the peripheral blood mononuclear cell compartment. The latter will allow us to see how many individual B cells in healthy adults produce antibodies that cross-react between enterovirus species, a knowledge gap with no existing data. Second, we aim to determine the molecular details of enterovirus species cross-reactivity using human mAbs. We will create human hybridomas that produce monoclonal antibodies (mAbs), screening and selecting for mAbs that bind to multiple enteroviruses. Antigen-binding fragments of the mAbs, in complex with virions from all immune-reactive enteroviruses, will be used for cryo-electron microscopy studies to obtain atomic resolution maps of antibody epitopes, detailing at a molecular level the determinants of cross-reactivity. Third, we aim to determine mechanisms of antibody action utilizing human primary cell infection models. As we investigate which stage of the viral life cycle different mAbs disrupt to exert function, using ex vivo differentiated human epithelial cell cultures will preserve the natural cell surface receptors used by viruses during in vivo human infection. The knowledge gained from these studies will allow detailed understanding of how cross-reactive antibodies bind to enteroviruses and how they function. In turn, the mAbs can be used to functionally validate that candidate immunogens preserve these cross-reactive B cell epitopes. Finally, some mAbs could later have potential for direct use in humans, either as prophylactic or therapeutic agents. This is an especially important consideration for populations such as the immunocompromised, who may not be able to generate strong antibody responses upon vaccination.