Soluble Angiotensin-Converting Enzyme 2 Protein Improves Survival and Lowers Viral Titers in Lethal Mouse Model of Severe Acute Respiratory Syndrome Coronavirus Type 2 Infection with the Delta Variant

Abstract

Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) utilizes angiotensin-converting enzyme 2 (ACE2) as its main receptor for cell entry. We bioengineered a soluble ACE2 protein termed ACE2 618-DDC-ABD that has increased binding to SARS-CoV-2 and prolonged duration of action. Here, we investigated the protective effect of this protein when administered intranasally to k18-hACE2 mice infected with the aggressive SARS-CoV-2 Delta variant. k18-hACE2 mice were infected with the SARS-CoV-2 Delta variant by inoculation of a lethal dose (2 × 10⁴ PFU). ACE2 618-DDC-ABD (10 mg/kg) or PBS was administered intranasally six hours prior and 24 and 48 h post-viral inoculation. All animals in the PBS control group succumbed to the disease on day seven post-infection (0% survival), whereas, in contrast, there was only one casualty in the group that received ACE2 618-DDC-ABD (90% survival). Mice in the ACE2 618-DDC-ABD group had minimal disease as assessed using a clinical score and stable weight, and both brain and lung viral titers were markedly reduced. These findings demonstrate the efficacy of a bioengineered soluble ACE2 decoy with an extended duration of action in protecting against the aggressive Delta SARS-CoV-2 variant. Together with previous work, these findings underline the universal protective potential against current and future emerging SARS-CoV-2 variants.

Keywords: ACE2; COVID-19; SARS-CoV-2; angiotensin-converting enzyme 2.

Figure 1

Survival of k18-hACE2 mice infected with the SARS-CoV-2 Delta variant that received either ACE2 618-DDC-ABD or PBS. Infected animals in the PBS control group (black) showed uniform lethality, and survival by day seven post-viral inoculation was 0%. In contrast, in the ACE2 618-DDC-ABD-treated group (purple), survival was 90% by day six post-viral inoculation, with only one mouse having to be humanely euthanized on day six. Five mice from the ACE2 618-DDC-ABD-treated group that were healthy according to their clinical score and weight were sacrificed on day six post-viral inoculation for comparison of viral titers at similar time points. The remaining four mice in the ACE2-618-DDC-ABD-treated group all survived until the end of the study protocol at day 14 post-viral inoculation. The difference in survival was statistically significant (p = 0.0004 using log-rank (Mantel–Cox) test).

Figure 2

Clinical score and body weight of k18-hACE2 mice infected with the SARS-CoV-2 Delta variant that received either ACE2 618-DDC-ABD or PBS. (Panel (A)) Infected animals in the PBS control group (white) developed severe disease, and their clinical scores increased accordingly until days six and seven post-viral inoculation by which time point, all mice had either died or had to be humanely euthanized. In contrast, animals in the ACE2 618-DDC-ABD-treated group (purple) developed less severe disease, and their clinical scores were lower compared to the PBS control group. All remaining animals from the ACE2 618-DDC-ABD-treated group had recovered by the end of the 14-day study period. (Panel (B)) Infected animals in the PBS control group (white) lost up to ~20% of their initial body weight until days six and seven post-viral inoculation by which time point, all mice had either died or had to be humanely euthanized. In contrast, animals in the ACE2 618-DDC-ABD-treated group (purple) lost, on average, less than ~10% of their initial body weight by days six and seven post-viral inoculation. All remaining animals from the ACE2 618-DDC-ABD-treated group recovered stable body weight by the end of the 14-day study period.

Figure 3

Lung, brain and kidney SARS-CoV-2 titers of k18-hACE2 mice infected with the SARS-CoV-2 Delta variant that received either ACE2 618-DDC-ABD or PBS. Organs were collected on day six post-viral inoculation. The data in the upper panels (Panels (A–C)) are also shown on a logarithmic scale in the lower panels (Panels (D–F)). Lung viral titers (Panel (A)) in infected animals from the ACE2 618-DDC-ABD-treated group (purple) were markedly reduced compared to lung viral titers in infected animals that received PBS (white) (2.5 × 10² ± 0.9 × 10² vs. 1.18 × 10⁴ ± 6.7 × 10³ PFU/mL, p = 0.004). Brain viral titers (Panel (B)) were also markedly reduced in mice that received ACE2 618-DDC-ABD (purple) compared to brain viral titers in the PBS control group (white) (1.8 × 10⁶ ± 1.8 × 10⁶ vs. 1.4 × 10⁷ ± 5.0 × 10⁶ PFU/mL, p = 0.008). Kidney viral titers (Panel (C)) could not be detected in any of the animals from either the ACE2 618-DDC-ABD-treated (purple) group or the PBS control group (white). In lower panels, the same differences are shown, but the logarithmic scale allows the visualization of some viral titers that were still detectable in lungs (Panel (D)) and brains (Panel (E)) of a few treated animals. In the kidneys (Panel (F)), no viral titers could be detected. ∗∗ p < 0.01.