Discovery of broadly‐neutralizing antibodies against brown recluse spider and Gadim scorpion sphingomyelinases using consensus toxins as antigens

Abstract

Broadly‐neutralizing monoclonal antibodies are becoming increasingly important tools for treating infectious diseases and animal envenomings. However, designing and developing broadly‐neutralizing antibodies can be cumbersome using traditional low‐throughput iterative protein engineering methods. Here, we present a new high‐throughput approach for the standardized discovery of broadly‐neutralizing monoclonal antibodies relying on phage display technology and consensus antigens representing average sequences of related proteins. We showcase the utility of this approach by applying it to toxic sphingomyelinases from the venoms of species from very distant orders of the animal kingdom, the recluse spider and Gadim scorpion. First, we designed a consensus sphingomyelinase and performed three rounds of phage display selection, followed by DELFIA‐based screening and ranking, and benchmarked this to a similar campaign involving cross‐panning against recombinant versions of the native toxins. Second, we identified two scFvs that not only bind the consensus toxins, but which can also neutralize sphingomyelinase activity of native whole venom in vitro. Finally, we conclude that the phage display campaign involving the use of the consensus toxin was more successful in yielding cross‐neutralizing scFvs than the phage display campaign involving cross‐panning.

In vivo neutralization of coral snake venoms with an oligoclonal nanobody mixture in a murine challenge model

“…This methodology enables the discovery of specific antibodies against the most medically relevant toxins, regardless of their abundance within the venoms or immunogenicity 20 . Phage display technology can furthermore facilitate the discovery of cross-neutralizing antibodies (antibodies that can neutralize more than one toxin isoform) through cross-panning strategies 17 , 19 , 21 or selections against recombinantly produced consensus antigens (i.e., antigens designed to represent the ‘average’ of several different proteins) 22 . Finally, it has been shown that monoclonal antibodies can be combined as carefully generated oligoclonal mixtures, allowing for the neutralization of multiple toxins by a single cocktail 18 .…”