Fc engineered ACE2-Fc is a potent multifunctional agent targeting SARS-CoV2

Abstract

Joining a function-enhanced Fc-portion of human IgG to the SARS-CoV-2 entry receptor ACE2 produces an antiviral decoy with strain transcending virus neutralizing activity. SARS-CoV-2 neutralization and Fc-effector functions of ACE2-Fc decoy proteins, formatted with or without the ACE2 collectrin domain, were optimized by Fc-modification. The different Fc-modifications resulted in distinct effects on neutralization and effector functions. H429Y, a point mutation outside the binding sites for FcγRs or complement caused non-covalent oligomerization of the ACE2-Fc decoy proteins, abrogated FcγR interaction and enhanced SARS-CoV-2 neutralization. Another Fc mutation, H429F did not improve virus neutralization but resulted in increased C5b-C9 fixation and transformed ACE2-Fc to a potent mediator of complement-dependent cytotoxicity (CDC) against SARS-CoV-2 spike (S) expressing cells. Furthermore, modification of the Fc-glycan enhanced cell activation via FcγRIIIa. These different immune profiles demonstrate the capacity of Fc-based agents to be engineered to optimize different mechanisms of protection for SARS-CoV-2 and potentially other viral pathogens.

Developing a workflow for the isolation of hybridoma cells producing fully human antigen-specific antibodies using a surface IgG detection method

“…The expression of the hACE2-Fc fusion protein and the associated vector construction methods have been previously published 21 . In brief, a full-length ACE2 extracellular domain coding sequence (amino acids 18-740) was fused to the human IgG1-Fc via a linker with a GGGGS sequence.…”

Targeting SARS-CoV-2 and host cell receptor interactions

“… Wines et al (2022) took further steps to modify the Fc portion of the ACE2 decoy to enable the construct to oligomerise and induce different downstream immunological responses. Trimeric WT ACE2 constructs with superior binding affinity and broad-spectrum neutralization efficacy have also been developed ( Guo et al, 2021 ; Xiao et al, 2021 ).…”

ACE2 decoy Fc-fusions and bi-specific killer engager (BiKEs) require Fc engagement forin vivoefficacy against SARS-CoV-2

“…Furthermore, we utilized known neutralizing and non-neutralizing antibody sequences to assess the contribution of neutralizing and non-neutralizing antibodies and BiKEs in the protection from SARS-CoV-2 58 . Lastly, using sequences previously published, we engineered and tested high affinity (ha) ACE2 mutant decoy molecules that can or cannot bind Fc receptors 33,35,37,46,49,59 K18-hACE mice in the infected group were anesthetized with 3% isoflurane/1.5 L/min oxygen in an induction chamber and inoculated intranasally with 1 x 10 4 plaque forming unit (PFU) of SARS-CoV-2 in a volume of 50 μL DMEM, split equally between each nostril 55,56 . Mice were monitored, and the body weight measured daily for the duration of the experiment.…”

“…Furthermore, we utilized known neutralizing and non-neutralizing antibody sequences to assess the contribution of neutralizing and non-neutralizing antibodies and BiKEs in the protection from SARS-CoV-2 58 . Lastly, using sequences previously published, we engineered and tested high affinity (ha) ACE2 mutant decoy molecules that can or cannot bind Fc receptors 33,35,37,46,49,59 . It was not previously known if haACE2 Fc-fusions or novel haACE2 decoy BiKEs would be effective at clearing SARS-CoV-2 in vivo with or without Fc engagement.…”