Engineered ACE2 receptor therapy overcomes mutational escape of SARS-CoV-2

Yusuke Higuchi,Eriko Ohgitani,Toru Okamoto,Yusuke Sakai,Atsushi Hoshino,Nariko Ikemura,Fuminori Sugihara,Yoshiharu Matsuura,Osam Mazda,Daisuke Motooka,Takao Arimori,Shota Nakamura,Tatsuya Suzuki,Satoaki Matoba,Yuhei Kirita,Emiko Mihara,Junichi Takagi,Yumi Itoh

doi:10.1038/s41467-021-24013-y

Abstract

SARS-CoV-2 has mutated during the global pandemic leading to viral adaptation to medications and vaccinations. Here we describe an engineered human virus receptor, ACE2, by mutagenesis and screening for binding to the receptor binding domain (RBD). Three cycles of random mutagenesis and cell sorting achieved sub-nanomolar affinity to RBD. Our structural data show that the enhanced affinity comes from better hydrophobic packing and hydrogen-bonding geometry at the interface. Additional disulfide mutations caused the fixing of a closed ACE2 conformation to avoid off-target effects of protease activity, and also improved structural stability. Our engineered ACE2 neutralized SARS-CoV-2 at a 100-fold lower concentration than wild type; we also report that no escape mutants emerged in the co-incubation after 15 passages. Therapeutic administration of engineered ACE2 protected hamsters from SARS-CoV-2 infection, decreased lung virus titers and pathology. Our results provide evidence of a therapeutic potential of engineered ACE2.

Highlights

SARS-CoV-2 has mutated during the global pandemic leading to viral adaptation to medications and vaccinations
The protease domain (PD) of Angiotensin-converting enzyme 2 (ACE2) is known to harbor the interface to viral spike protein, located in the top-middle part of ACE2 ectodomain
Mutant plasmid library was packaged into lentivirus, followed by expression in human 293T cells in less than 0.3 multiplicity of infection (MOI) to yield no more than one mutant ACE2 per cell

Summary

Introduction

SARS-CoV-2 has mutated during the global pandemic leading to viral adaptation to medications and vaccinations. The mutant escaping from sACE2 should have limited binding affinity to cell surface native ACE2 receptors, leading to a diminished or eliminated infectivity. Engineered ACE2 based on high affinity ACE2 mutant and human IgG1-Fc effectively neutralizes SARS-CoV-2 and exhibits the therapeutic effect in a COVID-19 model hamster. It prevents the emergence of escape mutation and effectively neutralizes current N501Y and E484K-based mutants as well as escape mutants from COVID-19 convalescent plasma. Engineering ACE2 decoy receptors with human cell-based directed evolution is a promising approach to develop a SARS-CoV-2 neutralizing drug that has affinity comparable to monoclonal antibodies yet displaying resistance to escape mutations of virus

Methods

Results

Conclusion