Engineered ACE2 receptor traps potently neutralize SARS-CoV-2

Abstract

An essential mechanism for SARS-CoV-1 and -2 infection begins with the viral spike protein binding to the human receptor protein angiotensin-converting enzyme II (ACE2). Here we describe a stepwise engineering approach to generate a set of affinity optimized, enzymatically inactivated ACE2 variants that potently block SARS-CoV-2 infection of cells. These optimized receptor traps tightly bind the receptor binding domain (RBD) of the viral spike protein and prevent entry into host cells. We first computationally designed the ACE2-RBD interface using a two-stage flexible protein backbone design process that improved affinity for the RBD by up to 12-fold. These designed receptor variants were affinity matured an additional 14-fold by random mutagenesis and selection using yeast surface display. The highest affinity variant contained seven amino acid changes and bound to the RBD 170-fold more tightly than wild-type ACE2. With the addition of the natural ACE2 collectrin domain and fusion to a human Fc domain for increased stabilization and avidity, the most optimal ACE2 receptor traps neutralized SARS-CoV-2 pseudotyped lentivirus and authentic SARS-CoV-2 virus with half-maximal inhibitory concentrations (IC50) in the tens of ng/ml range. Engineered ACE2 receptor traps offer a promising route to fighting infections by SARS-CoV-2 and other ACE2-utilizing coronaviruses, with the key advantage that viral resistance would also likely impair viral entry. Moreover, such traps can be pre-designed for viruses with known entry receptors for faster therapeutic response without the need for neutralizing antibodies isolated or generated from convalescent patients.

Endothelium Infection and Dysregulation by SARS-CoV-2: Evidence and Caveats in COVID-19

“…In contrast to previous SARS findings [ 138 ], several reports propose that sACE2 may actually have a protective effect against SARS-CoV-2 infection [ 141 , 142 , 143 ]. Monteil et al [ 141 ] showed by RT-qPCR that clinical-grade human recombinant soluble ACE2 (hrsACE2) reduced SARS-CoV-2 replication by 1000–5000-fold in cell culture, engineered human blood vessels, and kidney organoids.…”

“…It is believed that by binding the SARS-CoV-2 S-protein, sACE2 prevents its association with membrane-bound ACE2 and effectively blocks viral internalization [ 141 ]—a mechanism that previously demonstrated in SARS-CoV-1 [ 41 ]. In fact, in a collaborative study with our group, Glasgow et al [ 143 ] showed by RT-qPCR that highly optimized sACE2 was able to reduce replication of SARS-CoV-2 in Vero E6 cells more than 50,000-fold. Emerging reports of sACE2 neutralization capacity in COVID-19 are promising, although further research is required to elucidate its therapeutic efficacy.…”

Challenges and opportunities for antiviral monoclonal antibodies as COVID-19 therapy

“…Nevertheless, the potency of recombinant WT ACE2 is 2–3 logs worse than the best neutralizing mAbs. To improve the potency of ACE2 decoys, a number of groups have engineered variants of ACE2 using structural knowledge and screening of mutant libraries [ [140] , [141] , [142] ]. Some of these mutants exhibit significantly improved affinity to SARS-CoV-2 RBD, with neutralization potency that rivals some of the human mAbs.…”

Engineered Trimeric ACE2 Binds and Locks “Three-up” Spike Protein to Potently Inhibit SARS-CoVs and Mutants

“…proteins have been shown to increase spike protein binding affinities and virus inhibitory activities, albeit with reduced or loss of catalytic activities 28,47 . ACE2-Ig fusion proteins and neutralizing antibodies potentially could have antibody-dependent enhancement (ADE) effect to facilitate virus infection, although such phenomenon still needs further clinical studies 48,49 .…”