Longitudinal analyses reveal distinct immune response landscapes in lung and intestinal tissues from SARS-CoV-2-infected rhesus macaques.

The pathological and immune response of individuals with COVID-19 display different dynamics in lung and intestine. Here, we depict the single-cell transcriptional atlas of longitudinally collected lung and intestinal tissue samples from SARS-CoV-2-infected monkeys at 3 to 10 dpi. We find that intestinal enterocytes are degraded at 3 days post-infection but recovered rapidly, revealing that infection has mild effects on the intestine. Crucially, we observe suppression of the inflammatory response and tissue damage related to B-cell and Paneth cell accumulation in the intestines, although T cells are activated in the SARS-CoV-2 infection. Compared with that in the lung, the expression of interferon response-related genes is inhibited, and inflammatory factor secretion is reduced in the intestines. Our findings indicate an imbalance of immune dynamic in intestinal mucosa during SARS-CoV-2 infection, which may underlie ongoing rectal viral shedding and mild tissue damage.

Three doses of prototypic SARS-CoV-2 inactivated vaccine induce cross-protection against its variants of concern.

Variants are globally emerging very quickly following pandemic prototypic SARS-CoV-2. To evaluate the cross-protection of prototypic SARS-CoV-2 vaccine against its variants, we vaccinated rhesus monkeys with three doses of prototypic SARS-CoV-2 inactivated vaccine, followed by challenging with emerging SARS-CoV-2 variants of concern (VOCs). These vaccinated animals produced neutralizing antibodies against Alpha, Beta, Delta, and Omicron variants, although there were certain declinations of geometric mean titer (GMT) as compared with prototypic SARS-CoV-2. Of note, in vivo this prototypic vaccine not only reduced the viral loads in nasal, throat and anal swabs, pulmonary tissues, but also improved the pathological changes in the lung infected by variants of Alpha, Beta, and Delta. In summary, the prototypic SARS-CoV-2 inactivated vaccine in this study protected against VOCs to certain extension, which is of great significance for prevention and control of COVID-19.

Immunogenicity and protective efficacy of a recombinant protein subunit vaccine and an inactivated vaccine against SARS-CoV-2 variants in non-human primates.

Emerging SARS-CoV-2 variants and the gradually decreasing neutralizing antibodies over time post vaccination have led to an increase in incidents of breakthrough infection across the world. To investigate the potential protective effect of the recombinant protein subunit COVID-19 vaccine targeting receptor-binding domain (RBD) (PS-RBD) and whole inactivated virus particle vaccine (IV) against the variant strains, in this study, rhesus macaques were immunized with PS-RBD or IV vaccine, followed by a Beta variant (B.1.351) challenge. Although neutralizing activity against the Beta variant was reduced compared with that against the prototype, the decreased viral load in both upper and lower respiratory tracts, milder pathological changes, and downregulated inflammatory cytokine levels in lung tissues after challenge demonstrated that PS-RBD and IV still provided effective protection against the Beta variant in the macaque model. Furthermore, PS-RBD-induced macaque sera possessed general binding and neutralizing activity to Alpha, Beta, Delta, and Omicron variants in our study, though the neutralizing antibody (NAb) titers declined by varying degrees, demonstrating potential protection of PS-RBD against current circulating variants of concern (VOCs). Interestingly, although the IV vaccine-induced extremely low neutralizing antibody titers against the Beta variant, it still showed reduction for viral load and significantly alleviated pathological change. Other correlates of vaccine-induced protection (CoP) like antibody-dependent cellular cytotoxicity (ADCC) and immune memory were both confirmed to be existing in IV vaccinated group and possibly be involved in the protective mechanism.

An engineered bispecific human monoclonal antibody against SARS-CoV-2.

The global severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic requires effective therapies against coronavirus disease 2019 (COVID-19), and neutralizing antibodies are a promising therapy. A noncompeting pair of human neutralizing antibodies (B38 and H4) blocking SARS-CoV-2 binding to its receptor, ACE2, have been described previously. Here, we develop bsAb15, a bispecific monoclonal antibody (bsAb) based on B38 and H4. bsAb15 has greater neutralizing efficiency than these parental antibodies, results in less selective pressure and retains neutralizing ability to most SARS-CoV-2 variants of concern (with more potent neutralizing activity against the Delta variant). We also selected for escape mutants of the two parental mAbs, a mAb cocktail and bsAb15, demonstrating that bsAb15 can efficiently neutralize all single-mAb escape mutants. Furthermore, prophylactic and therapeutic application of bsAb15 reduced the viral titer in infected nonhuman primates and human ACE2 transgenic mice. Therefore, this bsAb is a feasible and effective strategy to treat and prevent severe COVID-19.

Preclinical immunological evaluation of an intradermal heterologous vaccine against SARS-CoV-2 variants.

The recent emergence of COVID-19 variants has necessitated the development of new vaccines that stimulate the formation of high levels of neutralizing antibodies against S antigen variants. A new strategy involves the intradermal administration of heterologous vaccines composed of one or two doses of inactivated vaccine and a booster dose with the mutated S1 protein (K-S). Such vaccines improve the immune efficacy by increasing the neutralizing antibody titers and promoting specific T cell responses against five variants of the RBD protein. A viral challenge test with the B.1.617.2 (Delta) variant confirmed that both administration schedules (i.e. "1 + 1" and "2 + 1") ensured protection against this strain. These results suggest that the aforementioned strategy is effective for protecting against new variants and enhances the anamnestic immune response in the immunized population.

Randomized, Double-Blinded, Placebo-Controlled Phase 2 Trial of an Inactivated Severe Acute Respiratory Syndrome Coronavirus 2 Vaccine in Healthy Adults.

BACKGROUND: We evaluated an inactivated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine for immunogenicity and safety in adults aged 18-59 years. METHODS: In this randomized, double-blinded, controlled trial, healthy adults received a medium dose (MD) or a high dose (HD) of the vaccine at an interval of either 14 days or 28 days. Neutralizing antibody (NAb) and anti-S and anti-N antibodies were detected at different times, and adverse reactions were monitored for 28 days after full immunization. RESULTS: A total of 742 adults were enrolled in the immunogenicity and safety analysis. Among subjects in the 0, 14 procedure, the seroconversion rates of NAb in MD and HD groups were 89% and 96% with geometric mean titers (GMTs) of 23 and 30, respectively, at day 14 and 92% and 96% with GMTs of 19 and 21, respectively, at day 28 after immunization. Anti-S antibodies had GMTs of 1883 and 2370 in the MD group and 2295 and 2432 in the HD group. Anti-N antibodies had GMTs of 387 and 434 in the MD group and 342 and 380 in the HD group. Among subjects in the 0, 28 procedure, seroconversion rates for NAb at both doses were both 95% with GMTs of 19 at day 28 after immunization. Anti-S antibodies had GMTs of 937 and 929 for the MD and HD groups, and anti-N antibodies had GMTs of 570 and 494 for the MD and HD groups, respectively. No serious adverse events were observed during the study period. CONCLUSIONS: Adults vaccinated with inactivated SARS-CoV-2 vaccine had NAb as well as anti-S/N antibody and had a low rate of adverse reactions. CLINICAL TRIALS REGISTRATION: NCT04412538.

Immunological evaluation of an inactivated SARS-CoV-2 vaccine in rhesus macaques.

Because of the relatively limited understanding of coronavirus disease 2019 (COVID-19) pathogenesis, immunological analysis for vaccine development is needed. Mice and macaques were immunized with an inactivated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine prepared by two inactivators. Various immunological indexes were tested, and viral challenges were performed on day 7 or 150 after booster immunization in monkeys. This inactivated SARS-CoV-2 vaccine was produced by sequential inactivation with formaldehyde followed by propiolactone. The various antibody responses and specific T cell responses to different viral antigens elicited in immunized animals were maintained for longer than 150 days. This comprehensive immune response could effectively protect vaccinated macaques by inhibiting viral replication in macaques and substantially alleviating immunopathological damage, and no clinical manifestation of immunopathogenicity was observed in immunized individuals during viral challenge. This candidate inactivated vaccine was identified as being effective against SARS-CoV-2 challenge in rhesus macaques.

Identification of T Cell Epitopes in the Spike Glycoprotein of Severe Acute Respiratory Syndrome Coronavirus 2 in Rhesus Macaques.

The T cell response is an important detection index in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine development. The present study was undertaken to determine the T cell epitopes in the spike (S) protein of SARS-CoV-2 that dominate the T cell responses in SARS-CoV-2-infected patients. PBMCs from rhesus macaques vaccinated with a DNA vaccine encoding the full-length S protein were isolated, and an ELISPOT assay was used to identify the recognized T cell epitopes among a total of 158 18-mer and 10-aa-overlapping peptides spanning the full-length S protein. Six multipeptide-based epitopes located in the S1 region, with four of the six located in the receptor-binding domain, were defined as the most frequently recognized epitopes in macaques. The conservation of the epitopes across species was also verified, and peptide mixtures for T cell response detection were established. Six newly defined T cell epitopes were found in the current study, which may provide a novel potential target for T cell response detection and the diagnosis and vaccine design of SARS-CoV-2 based on multipeptide subunit-based epitopes.

Self-Assembling Nanoparticle Vaccines Displaying the Receptor Binding Domain of SARS-CoV-2 Elicit Robust Protective Immune Responses in Rhesus Monkeys.

SARS-CoV-2 caused the COVID-19 pandemic that lasted for more than a year. Globally, there is an urgent need to use safe and effective vaccines for immunization to achieve comprehensive protection against SARS-CoV-2 infection. Focusing on developing a rapid vaccine platform with significant immunogenicity as well as broad and high protection efficiency, we designed a SARS-CoV-2 spike protein receptor-binding domain (RBD) displayed on self-assembled ferritin nanoparticles. In a 293i cells eukaryotic expression system, this candidate vaccine was prepared and purified. After rhesus monkeys are immunized with 20 µg of RBD-ferritin nanoparticles three times, the vaccine can elicit specific humoral immunity and T cell immune response, and the neutralizing antibodies can cross-neutralize four SARS-CoV-2 strains from different sources. In the challenge protection test, after nasal infection with 2 × 105 CCID50 SARS-CoV-2 virus, compared with unimmunized control animals, virus replication in the vaccine-immunized rhesus monkeys was significantly inhibited, and respiratory pathology observations also showed only slight pathological damage. These analyses will benefit the immunization program of the RBD-ferritin nanoparticle vaccine in the clinical trial design and the platform construction to present a specific antigen domain in the self-assembling nanoparticle in a short time to harvest stable, safe, and effective vaccine candidates for new SARS-CoV-2 isolates.

The safety and immunogenicity of an inactivated SARS-CoV-2 vaccine in Chinese adults aged 18-59 years: A phase I randomized, double-blinded, controlled trial.

BACKGROUND: This study examined the safety and immunogenicity of an inactivated SARS-CoV-2 vaccine. METHOD: In a phase I randomized, double-blinded, placebo-controlled trial involving 192 healthy adults 18-59 years old, two injections of three doses (50 EU, 100 EU, 150 EU) of an inactivated SARS-CoV-2 vaccine or placebo were administered intramuscularly at a 2- or 4-week interval. The safety and immunogenicity of the vaccine were evaluated. RESULTS: Vaccination was completed in 191 subjects. Forty-four adverse reactions occurred within 28 days, most commonly mild pain and redness at the injection site or slight fatigue. At days 14 and 28, the seroconversion rates were 87.5% and 79.2% (50 EU), 100% and 95.8% (100 EU), and 95.8% and 87.5% (150 EU), respectively, with geometric mean titers (GMTs) of 18.1 and 10.6, 54.5 and 15.4, and 37.1 and 18.5, respectively, for the schedules with 2-week and 4-week intervals. Seroconversion was associated with synchronous upregulation of antibodies against the S protein, N protein and virion and a cytotoxic T lymphocyte (CTL) response. No cytokines and immune cells related to immunopathology were observed. Transcriptome analysis revealed the genetic diversity of immune responses induced by the vaccine. INTERPRETATION: In a population aged 18-59 years in this trial, this inactivated SARS-CoV-2 vaccine was safe and immunogenic. TRIAL REGISTRATION: CTR20200943 and NCT04412538.

A novel DNA and protein combination COVID-19 vaccine formulation provides full protection against SARS-CoV-2 in rhesus macaques.

The current study aims to develop a safe and highly immunogenic COVID-19 vaccine. The novel combination of a DNA vaccine encoding the full-length Spike (S) protein of SARS-CoV-2 and a recombinant S1 protein vaccine induced high level neutralizing antibody and T cell immune responses in both small and large animal models. More significantly, the co-delivery of DNA and protein components at the same time elicited full protection against intratracheal challenge of SARS-CoV-2 viruses in immunized rhesus macaques. As both DNA and protein vaccines have been proven safe in previous human studies, and DNA vaccines are capable of eliciting germinal center B cell development, which is critical for high-affinity memory B cell responses, the DNA and protein co-delivery vaccine approach has great potential to serve as a safe and effective approach to develop COVID-19 vaccines that provide long-term protection.

Virulence and pathogenesis of SARS-CoV-2 infection in rhesus macaques: A nonhuman primate model of COVID-19 progression.

The COVID-19 has emerged as an epidemic, causing severe pneumonia with a high infection rate globally. To better understand the pathogenesis caused by SARS-CoV-2, we developed a rhesus macaque model to mimic natural infection via the nasal route, resulting in the SARS-CoV-2 virus shedding in the nose and stool up to 27 days. Importantly, we observed the pathological progression of marked interstitial pneumonia in the infected animals on 5-7 dpi, with virus dissemination widely occurring in the lower respiratory tract and lymph nodes, and viral RNA was consistently detected from 5 to 21 dpi. During the infection period, the kinetics response of T cells was revealed to contribute to COVID-19 progression. Our findings implied that the antiviral response of T cells was suppressed after 3 days post infection, which might be related to increases in the Treg cell population in PBMCs. Moreover, two waves of the enhanced production of cytokines (TGF-α, IL-4, IL-6, GM-CSF, IL-10, IL-15, IL-1ß), chemokines (MCP-1/CCL2, IL-8/CXCL8, and MIP-1ß/CCL4) were detected in lung tissue. Our data collected from this model suggested that T cell response and cytokine/chemokine changes in lung should be considered as evaluation parameters for COVID-19 treatment and vaccine development, besides of observation of virus shedding and pathological analysis.

An Antioxidant Enzyme Therapeutic for COVID-19.

The COVID-19 pandemic has taken a significant toll on people worldwide, and there are currently no specific antivirus drugs or vaccines. Herein it is a therapeutic based on catalase, an antioxidant enzyme that can effectively breakdown hydrogen peroxide and minimize the downstream reactive oxygen species, which are excessively produced resulting from the infection and inflammatory process, is reported. Catalase assists to regulate production of cytokines, protect oxidative injury, and repress replication of SARS-CoV-2, as demonstrated in human leukocytes and alveolar epithelial cells, and rhesus macaques, without noticeable toxicity. Such a therapeutic can be readily manufactured at low cost as a potential treatment for COVID-19.