Single-domain antibody fragments against poliovirus: mechanisms of neutralization and use in research and eradication

Poliovirus, as the causative agent of poliomyelitis, has plagued mankind throughout history. The development and implementation of two successful vaccines against the virus in the 1950s has since impressively decreased the number of polio-patients. However, to complete the last phases of polio-eradication and to protect people afterwards, the development of multiple anti-polioviral drugs and certain tools (e.g. standards in vaccine-control) is needed. As an interesting approach, single-domain antibody fragments (VHHs or Nanobodies®) can be thought of. Five in vitro neutralizing VHHs against poliovirus type 1 were previously developed at our department. The current work describes the mechanisms of neutralization of these VHHs in detail, as well as the epitopes targeted on the polioviral capsid and the structural changes involved at the VHH-poliovirus interface. Interestingly, the VHHs were found to bind the capsid at sites that extensively overlap the binding site of the natural poliovirus receptor, and at structures that are known to move during the expansion of the metastable capsid during its natural replication cycle. Binding with these residues explains the stabilizing activity of the VHHs against heat-induced degradation of the capsid and the multi-level mechanisms of neutralization of the five VHHs. As a second part of the work, neutralization escape mutants resistant against the VHHs have been selected, identified and characterized. This is important in the context of developing the VHHs as anti-viral drugs and provides knowledge on the implications of these mutants and, concurrently, on strategies to limit therapy-resistance. Additionally, the possibility to format the VHHs into constructs and to use them as probes in research and eradication was explored.