Abstract
Passive immunization is an effective option for treatment against hand, foot and mouth disease caused by EV71, especially with cross-neutralizing IgG monoclonal antibodies. In this study, an EV71-specific IgG2a antibody designated 5H7 was identified and characterized. 5H7 efficiently neutralizes the major EV71 genogroups (A, B4, C2, C4). The conformational epitope of 5H7 was mapped to the highly conserved amino acid position 74 on VP3 capsid protein using escape mutants. Neutralization with 5H7 is mediated by the inhibition of viral attachment, as revealed by virus-binding and post-attachment assays. In a competitive pull-down assay with SCARB2, 5H7 blocks the receptor-binding site on EV71 for virus neutralization. Passive immunization of chimeric 5H7 protected 100% of two-week-old AG129 mice from lethal challenge with an EV71 B4 strain for both prophylactic and therapeutic treatments. In contrast, 10D3, a previously reported neutralizing antibody that takes effect after virus attachment, could only confer prophylactic protection. These results indicate that efficient interruption of viral attachment is critical for effective therapeutic activity with 5H7. This report documents a novel universal neutralizing IgG antibody for EV71 therapeutics and reveals the underlying mechanism.
Highlights
Over the last decade, frequent epidemic outbreaks of hand, foot and mouth disease (HFMD) have been observed in the Asia-Pacific region
The 5H7 immunoglobulin isotype was determined as IgG2a. 5H7 was selected based on its broad reactivity to all 11 enterovirus 71 (EV71) subgenogroups by Immunofluorescence assay (IFA) with Vero cells, without cross-reactivity to coxsackievirus A16 (CVA16) (Fig. 1)
Figure 7. 5H7 prevents EV71 from binding to SCARB2 receptor. (A): A pull-down assay was performed with SCARB2, B4 virus, and different antibodies
Summary
Frequent epidemic outbreaks of hand, foot and mouth disease (HFMD) have been observed in the Asia-Pacific region. 2 ml of EV71 B4 virus (108 TCID50/ml) mixed with either 2 ml of antibodies or medium were added to each T25 flask with Vero cells, and incubated at 37 °C for 1 hour. No neutralization activity by Mab 5H7 was detected against the identified escape mutants (Table 1 and Fig. 2).
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