Angiotensin-converting enzyme 2 (ACE2) is a receptor for cell entry of SARS-CoV-2, and recombinant soluble ACE2 protein inhibits SARS-CoV-2 infection as a decoy. ACE2 is a carboxypeptidase that degrades angiotensin II, thereby improving the pathologies of cardiovascular disease or acute lung injury. Here we show that B38-CAP, an ACE2-like enzyme, is protective against SARS-CoV-2-induced lung injury. Endogenous ACE2 expression is downregulated in the lungs of SARS-CoV-2-infected hamsters, leading to elevation of angiotensin II levels. Recombinant Spike also downregulates ACE2 expression and worsens the symptoms of acid-induced lung injury. B38-CAP does not neutralize cell entry of SARS-CoV-2. However, B38-CAP treatment improves the pathologies of Spike-augmented acid-induced lung injury. In SARS-CoV-2-infected hamsters or human ACE2 transgenic mice, B38-CAP significantly improves lung edema and pathologies of lung injury. These results provide the first in vivo evidence that increasing ACE2-like enzymatic activity is a potential therapeutic strategy to alleviate lung pathologies in COVID-19 patients.

Significance: COVID-19 has shown severe pathogenicity in people with underlying diseases and in elderly people. It is necessary to elucidate the pathological conditions in patients with underlying diseases and in elderly patients. The use of appropriate animal models of COVID-19 is required for the development of pharmaceutical products; however, usually healthy young animals are used as experimental animals. Cynomolgus macaques with various clinical conditions and ages were infected with severe acute respiratory syndrome coronavirus 2 and there was a difference in pathological condition between young individuals and old-aged individuals with underlying diseases; therefore, the COVID-19 cynomolgus monkey model reflecting the pathophysiology of humans would be useful for elucidating the pathophysiology and developing therapeutic and prophylactic agents.

The Omicron variant continuously evolves under the humoral immune pressure exerted by vaccination and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, and the resulting Omicron subvariants display further immune evasion and antibody escape. An engineered angiotensin-converting enzyme 2 (ACE2) decoy composed of high-affinity ACE2 and an IgG1 Fc domain could offer an alternative modality to neutralize SARS-CoV-2. We previously reported its broad spectrum and therapeutic potential in rodent models. Here, we demonstrate that the engineered ACE2 decoy retains neutralization activity against Omicron subvariants, including the currently emerging XBB and BQ.1 strains, which completely evade antibodies currently in clinical use. SARS-CoV-2, under the suboptimal concentration of neutralizing drugs, generated SARS-CoV-2 mutants escaping wild-type ACE2 decoy and monoclonal antibodies, whereas no escape mutant emerged against the engineered ACE2 decoy. Furthermore, inhalation of aerosolized decoys improved the outcomes of rodents infected with SARS-CoV-2 at a 20-fold lower dose than that of intravenous administration. Last, the engineered ACE2 decoy exhibited therapeutic efficacy for cynomolgus macaques infected with SARS-CoV-2. These results indicate that this engineered ACE2 decoy represents a promising therapeutic strategy to overcome immune-evading SARS-CoV-2 variants and that liquid aerosol inhalation could be considered as a noninvasive approach to enhance the efficacy of COVID-19 treatments.

COVID-19, caused by SARS-CoV-2, has spread worldwide with dire consequences. To urgently investigate the pathogenicity of COVID-19 and develop vaccines and therapeutics, animal models that are highly susceptible to SARS-CoV-2 infection are needed. In the present study, we established an animal model highly susceptible to SARS-CoV-2 via the intratracheal tract infection in CAG promoter–driven human angiotensin-converting enzyme 2–transgenic (CAG-hACE2) mice. The CAG-hACE2 mice showed several severe symptoms of SARS-CoV-2 infection, with definitive weight loss and subsequent death. Acute lung injury with elevated cytokine and chemokine levels was observed at an early stage of infection in CAG-hACE2 mice infected with SARS-CoV-2. Analysis of the hACE2 gene in CAG-hACE2 mice revealed that more than 15 copies of hACE2 genes were integrated in tandem into the mouse genome, supporting the high susceptibility to SARS-CoV-2. In the developed model, immunization with viral antigen or injection of plasma from immunized mice prevented body weight loss and lethality due to infection with SARS-CoV-2. These results indicate that a highly susceptible model of SARS-CoV-2 infection in CAG-hACE2 mice via the intratracheal tract is suitable for evaluating vaccines and therapeutic medicines.

An infiltration of CD8+ lymphocytes in the dermis and epidermis underlies the skin rash that commonly occurs as a primary manifestation of an AIDS virus infection. These cutaneous lymphocytes were characterized in simian immunodeficiency virus of macaques (SIVmac)-infected rhesus monkeys. Skin rash-associated lymphocytes exhibited greater lysis of SIVmac-expressing target cells and a higher cloning efficiency for SIVmac-specific effector T cells than PBL. Moreover, both SIVmac envelope- and gag-specific CTL could be readily cloned from these skin rash-associated lymphocytes. In fact, the skin rash-associated CTL exhibited the same MHC restriction and epitope specificity as those CTL derived from PBL. These studies, therefore, demonstrate that the cutaneous infiltrating CD8+ lymphocytes in SIVmac-infected rhesus monkeys include SIVmac-specific CTL. Thus, whereas virus-specific CTL are likely to represent an important mechanism for controlling AIDS virus infections, they also may play a role in the pathogenesis of the skin lesions that occur after this infection.

Background: SARS-CoV-2 Omicron variants are highly resistant to vaccine-induced immunity and human monoclonal antibodies. Methods: We previously reported that two nanobodies, P17 and P86, potently neutralize SARS-CoV-2 VOCs. In this study, we modified these nanobodies into trimers, called TP17 and TP86 and tested their neutralization activities against Omicron BA.1 and subvariant BA.2 using pseudovirus assays. Next, we used TP17 and TP86 nanobody cocktail to treat ACE2 transgenic mice infected with lethal dose of SARS-CoV-2 strains, original, Delta and Omicron BA.1. Results: Here, we demonstrate that a novel nanobody TP86 potently neutralizes both BA.1 and BA.2 Omicron variants, and that the TP17 and TP86 nanobody cocktail broadly neutralizes in vitro all VOCs as well as original strain. Furthermore, intratracheal administration of this nanobody cocktail suppresses weight loss and prolongs survival of human ACE2 transgenic mice infected with SARS-CoV-2 strains, original, Delta and Omicron BA.1. Conclusions: Intratracheal trimerized nanobody cocktail administration suppresses weight loss and prolongs survival of SARS-CoV-2 infected mice.