Studies Showing Angiotensin Converting Enzyme 2 Decoy Proteins to be Promising Anti-SARS-CoV-2 Agents

In a recent study posted on Bio Rxiv*Preprint server, Researchers presented a microbody recombinant decoy protein with high affinity for angiotensin-converting enzyme 2 (ACE2) as an option for coronavirus disease 2019 (COVID-19) treatment.

Background

Constant emergence of increasingly immune-evading severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) VOCs (variants of concern) with multiple SARS-CoV-2 spike (S) protein mutations It is an issue. efficacy of Vaccines and monoclonal antibodies (mAbs) in the prevention and control of SARS-CoV-2 infection.

More effective prevention and treatment options are needed to reduce the burden of COVID-19. The authors of this study have previously reported that the SARS-CoV-2 strain, D614G strain and VOCs, were potently neutralized by binding to SARS-CoV-2 S and inhibiting S attachment. We developed the pcACE2.mb receptor decoy protein. Protein containing human ACE2 (hACE2).

About research

In the current study, the researchers extended their previous analysis by modifying pcACE2.mb and testing its efficacy against the SARS-CoV-2 VOC in mice.

The potency of pcACE2.mb was increased by incorporating into the S protein the T27Y/L79T/N330Y mutations that increase S-ACE2 binding. Additionally, her IgG1 CH3 domain in pcACE2.mb was truncated to increase half-life. Decoy proteins were transfected using ExpiCHO cells and purified by size exclusion and affinity type chromatography. A pseudotyped lentivirus neutralization assay was performed to assess the anti-SARS-CoV-2 efficacy of pcACE2.mb.

Decoy binding affinities to SARS-CoV-2 S were determined based on virion binding assays of sACE2 (soluble ACE2)-NLuc (nanoluciferase), ACE2.mb-NLuc, and ACE2.1mb-NLuc decoy:nanoluciferase fusions. It was assessed by assessing binding. Proteins of SARS-CoV-2 S-expressing 293T cells. To assess the tissue localization of pcACE2.mb, proteins ACE2.1mb-NLuc and sACE2-NLuc were injected intraperitoneally (ip) into mice and imaged for 3.0 days.

To assess the effect of delivery route on ACE2.1mb distribution, decoy proteins were administered intraperitoneally, iv (intravenously) or in (intranasally).To evaluate decoy half-life live, luciferase activity in lung tissue and serum was assessed. further away, live To assess the ability of the high-affinity ACE2.1mb decoy to prevent COVID-19, experiments were performed by administering the decoy by either route to hACE2-Tg mice with hACE2 knockins.

Subsequently, mice were challenged with SARS-COV-2 USA-WA1/2020 and, for comparative evaluation, animals were treated with the mAb cocktail REGN-CoV2 followed by SARS-CoV-2 ribonucleic acid (RNA) in the lungs. ) was quantified. 3.0 days after injection (dpi). BALB/c mice were used to assess decoy efficacy against Omicron BA.1 and BA.2 sub-VOCs compared to control mice treated with LY-CoV1404 mAb or REGN-COV2 cocktail .

result

Recombinant decoys significantly reduced viral load in mice when challenged with D614G, Omicron BA.1, and Omicron BA.2, and when administered post-infection, although the Omicron sub-VOC was less potent, SARS-CoV-2 strongly suppressed the replication of The decoy conferred broader protection against SARS-CoV-2 with similar potency compared to REGN-COV2 and better protection than LY-CoV1404 mAb. The ACE2.1mb decoy neutralized D614G with a 10.0-fold higher potency than sACE2, and the high-affinity ACE2.1mb had a 5.0-fold higher neutralizing potency.

In addition, the high-affinity microbody ACE2.1mb decoy was also highly potent against alpha-VOC, beta-VOC, gamma-VOC, and delta-VOC, but not Omicron BA.1 and Omicron BA.2, respectively. In mice injected intraperitoneally and intravenously, the decoy was mainly localized to the serum, liver, and spleen, and, of note, intravenous administration increased localization of the decoy to the lung by 100-fold. did. Intranasal administration localized the decoy to the trachea and lungs.

Decoy injected intravenously or intranasally had a serum half-life of 5 days. In lung tissue, the half-lives of intravenously and intranasally administered decoys were 4 and 8 days, respectively. Intraperitoneal, intravenous, and intranasal decoy administration reduced viral load by 108-fold, 15,700-fold, and 26,500-fold, respectively. Histopathological evaluation of lung tissue showed a pronounced immunological cell infiltration in untreated mice. This was largely absent in sACE2-treated mice and completely absent in ACE2.1mb-treated mice.

SARS-CoV-2 RNA was not detected in the lung during the course of treatment.Intranasal and intravenous administration of 100 μg decoy suppressed SARS-CoV-2 proliferation to undetectable of decoys inhibited SARS-CoV-2 proliferation by 12-fold. Intranasal administration showed slightly higher efficacy compared to intravenous administration at doses of 10 and 50 micrograms.

Decoy reduced the viral load of Omicron BA.1 by 56.0-fold by intravenous administration and Omicron BA.2 by 100.0-fold by intranasal administration. Greater protection was observed when decoy was administered up to 2.0 days before COVID-19, moderate protection was observed when administered at 3.0 dpi (200-fold reduction in viral load), and no protection at 5.0 dpi .

At 1.0 hours after COVID-19, intravenously and intranasally administered decoys reduced viral load by 1,100.0-fold and 27,500.0-fold, respectively. Similar findings were observed when decoys were administered 6 hours after SARS-CoV-2 infection. However, 12 hours after SARS-CoV-2 infection, intravenously injected decoy protein had no significant effect. Consistently maintained.

In conclusion, based on our research results, the high-affinity recombinant ACE2.1mb decoy protein may be a promising preventive and therapeutic option against SARS-CoV-2 VOC and Omicron sub-VOC.

*Important Notices

Bio Rxiv We publish a non-peer-reviewed, preliminary scientific report and should not be viewed as conclusive, to guide clinical practice/health-related actions, or to be treated as established information.