Development of a mutant aerosolized ACE2 that neutralizes SARS-CoV-2 in vivo

• Published in: mBio 2024-06, Vol.15 (6), p.e0076824
• Main Authors: Kober, Daniel L, Caballero Van Dyke, Marley C, Eitson, Jennifer L, Boys, Ian N, McDougal, Matthew B, Rosenbaum, Daniel M, Schoggins, John W
• Format: Article
• Language: English
• Subjects: ACE2; Aerosols; Angiotensin-Converting Enzyme 2 - genetics; Angiotensin-Converting Enzyme 2 - metabolism; Animals; antiviral agents; COVID-19 - virology; Female; HEK293 Cells; Humans; Mice; Mutation; mutational studies; receptor-ligand interaction; receptors; Research Article; SARS-CoV-2; SARS-CoV-2 - drug effects; SARS-CoV-2 - genetics; Spike Glycoprotein, Coronavirus - genetics; Spike Glycoprotein, Coronavirus - metabolism; Virology
• ISSN: 2150-7511; 2150-7511
• DOI: 10.1128/mbio.00768-24

The rapid evolution of SARS-CoV-2 variants highlights the need for new therapies to prevent disease spread. SARS-CoV-2, like SARS-CoV-1, uses the human cell surface protein angiotensin-converting enzyme 2 (ACE2) as its native receptor. Here, we design and characterize a mutant ACE2 that enables rapid affinity purification of a dimeric protein by altering the active site to prevent autoproteolytic digestion of a C-terminal His epitope tag. In cultured cells, mutant ACE2 competitively inhibits lentiviral vectors pseudotyped with spikes from multiple SARS-CoV-2 variants and infectious SARS-CoV-2. Moreover, the protein can be nebulized and retains virus-binding properties. We developed a system for the delivery of aerosolized ACE2 to K18-hACE2 mice and demonstrated protection by our modified ACE2 when delivered as a prophylactic agent. These results show proof-of-concept for an aerosolized delivery method to evaluate anti-SARS-CoV-2 agents and suggest a new tool in the ongoing fight against SARS-CoV-2 and other ACE2-dependent viruses. The rapid evolution of SARS-CoV-2 variants poses a challenge for immune recognition and antibody therapies. However, the virus is constrained by the requirement that it recognizes a human host receptor protein. A recombinant ACE2 could protect against SARS-CoV-2 infection by functioning as a soluble decoy receptor. We designed a mutant version of ACE2 with impaired catalytic activity to enable the purification of the protein using a single affinity purification step. This protein can be nebulized and retains the ability to bind the relevant domains from SARS-CoV-1 and SARS-CoV-2. Moreover, this protein inhibits viral infection against a panel of coronaviruses in cells. Finally, we developed an aerosolized delivery system for animal studies and show the modified ACE2 offers protection in an animal model of COVID-19. These results show proof-of-concept for an aerosolized delivery method to evaluate anti-SARS-CoV-2 agents and suggest a new tool in the ongoing fight against SARS-CoV-2.