Clofazimine broadly inhibits coronaviruses including SARS-CoV-2.

The COVID-19 pandemic is the third outbreak this century of a zoonotic disease caused by a coronavirus, following the emergence of severe acute respiratory syndrome (SARS) in 20031 and Middle East respiratory syndrome (MERS) in 20122. Treatment options for coronaviruses are limited. Here we show that clofazimine-an anti-leprosy drug with a favourable safety profile3-possesses inhibitory activity against several coronaviruses, and can antagonize the replication of SARS-CoV-2 and MERS-CoV in a range of in vitro systems. We found that this molecule, which has been approved by the US Food and Drug Administration, inhibits cell fusion mediated by the viral spike glycoprotein, as well as activity of the viral helicase. Prophylactic or therapeutic administration of clofazimine in a hamster model of SARS-CoV-2 pathogenesis led to reduced viral loads in the lung and viral shedding in faeces, and also alleviated the inflammation associated with viral infection. Combinations of clofazimine and remdesivir exhibited antiviral synergy in vitro and in vivo, and restricted viral shedding from the upper respiratory tract. Clofazimine, which is orally bioavailable and comparatively cheap to manufacture, is an attractive clinical candidate for the treatment of outpatients and-when combined with remdesivir-in therapy for hospitalized patients with COVID-19, particularly in contexts in which costs are an important factor or specialized medical facilities are limited. Our data provide evidence that clofazimine may have a role in the control of the current pandemic of COVID-19 and-possibly more importantly-in dealing with coronavirus diseases that may emerge in the future.

Integration of metalloproteome and immunoproteome reveals a tight link of iron-related proteins with COVID-19 pathogenesis and immunity.

Increasing clinical data show that the imbalance of host metallome is closely associated with different kinds of disease, however, the intrinsic mechanisms of action of metals in immunity and pathogenesis of disease remain largely undefined. There is lack of multiplexed profiling system to integrate the metalloproteome-immunoproteome information at systemic level for exploring the roles of metals in immunity and disease pathogenesis. In this study, we build up a metal-coding assisted multiplexed proteome assay platform for serum metalloproteomic and immunoproteomic profiling. By taking COVID-19 as a showcase, we unbiasedly uncovered the most evident modulation of iron-related proteins, i.e., Ft and Tf, in serum of severe COVID-19 patients, and the value of Ft/Tf could work as a robust biomarker for COVID-19 severity stratification, which overtakes the well-established clinical risk factors (cytokines). We further uncovered a tight association of transferrin with inflammation mediator IL-10 in COVID-19 patients, which was proved to be mainly governed by the monocyte/macrophage of liver, shedding light on new pathophysiological and immune regulatory mechanisms of COVID-19 disease. We finally validated the beneficial effects of iron chelators as anti-viral agents in SARS-CoV-2-infected K18-hACE2 mice through modulation of iron dyshomeostasis and alleviating inflammation response. Our findings highlight the critical role of liver-mediated iron dysregulation in COVID-19 disease severity, providing solid evidence on the involvement of iron-related proteins in COVID-19 pathophysiology and immunity.

Comparative analysis of SARS-CoV-2 Omicron BA.2.12.1 and BA.5.2 variants.

The initial severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariants, BA.1 and BA.2, are being progressively displaced by BA.5 in many countries. To provide insight on the replacement of BA.2 by BA.5 as the dominant SARS-CoV-2 variant, we performed a comparative analysis of Omicron BA.2.12.1 and BA.5.2 variants in cell culture and hamster models. We found that BA.5.2 exhibited enhanced replicative kinetics over BA.2.12.1 in vitro and in vivo, which is evidenced by the dominant BA.5.2 viral genome detected at different time points, regardless of immune selection pressure with vaccine-induced serum antibodies. Utilizing reverse genetics, we constructed a mutant SARS-CoV-2 carrying spike F486V substitution, which is an uncharacterized mutation that concurrently discriminates Omicron BA.5.2 from BA.2.12.1 variant. We noticed that the 486th residue does not confer viral replication advantage to the virus. We also found that 486V displayed generally reduced immune evasion capacity when compared with its predecessor, 486F. However, the surge of fitness in BA.5.2 over BA.2.12.1 was not due to stand-alone F486V substitution but as a result of the combination of multiple mutations. Our study upholds the urgency for continuous monitoring of SARS-CoV-2 Omicron variants with enhanced replication fitness.

Low Environmental Temperature Exacerbates Severe Acute Respiratory Syndrome Coronavirus 2 Infection in Golden Syrian Hamsters.

BACKGROUND: The effect of low environmental temperature on viral shedding and disease severity of Coronavirus Disease 2019 (COVID-19) is uncertain. METHODS: We investigated the virological, clinical, pathological, and immunological changes in hamsters housed at room (21°C), low (12-15°C), and high (30-33°C) temperature after challenge by 105 plaque-forming units of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). RESULTS: The nasal turbinate, trachea, and lung viral load and live virus titer were significantly higher (~0.5-log10 gene copies/ß-actin, P < .05) in the low-temperature group at 7 days postinfection (dpi). The low-temperature group also demonstrated significantly higher level of tumor necrosis factor-α, interferon-γ (IFN-γ), interleukin-1ß, and C-C motif chemokine ligand 3, and lower level of the antiviral IFN-α in lung tissues at 4 dpi than the other 2 groups. Their lungs were grossly and diffusely hemorrhagic, with more severe and diffuse alveolar and peribronchiolar inflammatory infiltration, bronchial epithelial cell death, and significantly higher mean total lung histology scores. By 7 dpi, the low-temperature group still showed persistent and severe alveolar inflammation and hemorrhage, and little alveolar cell proliferative changes of recovery. The viral loads in the oral swabs of the low-temperature group were significantly higher than those of the other two groups from 10 to 17 dpi by about 0.5-1.0 log10 gene copies/ß-actin. The mean neutralizing antibody titer of the low-temperature group was significantly (P < .05) lower than that of the room temperature group at 7 dpi and 30 dpi. CONCLUSIONS: This study provided in vivo evidence that low environmental temperature exacerbated the degree of virus shedding, disease severity, and tissue proinflammatory cytokines/chemokines expression, and suppressed the neutralizing antibody response of SARS-CoV-2-infected hamsters. Keeping warm in winter may reduce the severity of COVID-19.

Role of the Hippo-YAP/NF-κB signaling pathway crosstalk in regulating biological behaviors of macrophages under titanium ion exposure.

The presence of metal ions, such as titanium (Ti) ions, is toxic to adjacent tissues of implants. Indeed, Ti ions may induce an inflammatory response through the NF-κB pathway, thus causing damage to soft and hard tissues. The involvement of Yes-associated protein (YAP), a key factor of the Hippo pathway, in an immuno-inflammatory response has been confirmed, whereas its role in Ti ion-mediated inflammation has not been elucidated. Therefore, this study aimed to investigate the role of signal crosstalk between the Hippo/YAP and NF-κB signaling pathways in the pro-inflammatory effect of Ti ions on macrophages. In our work, RAW264.7 cells were cocultured with Ti ions. The migration capacity of macrophages under Ti ion exposure was measured by transwell assay. Western blot analysis was used to detect the expressions of related proteins. Polymerase chain reaction was used to evaluate the expression of pro-inflammatory cytokines. The nucleus translocation of YAP and P65 was visualized and analyzed via immunofluorescence staining. The results showed that the migration of macrophages was promoted under Ti ion exposure. Ten parts per million Ti ions induced nuclear expression of YAP and activated the NF-κB pathway, which finally upregulated the expression of pro-inflammatory cytokines in macrophages. Moreover, the inhibition of the NF-κB pathway rescued the reduction of YAP expression under Ti ion exposure. Most importantly, the overexpression of YAP exacerbated the inflammatory response mediated by Ti ions through the NF-κB pathway. In summary, this study explored the mechanism of Hippo-YAP/NF-κB pathway crosstalk involved in the regulation of macrophage behaviors under Ti ion exposure.

Simulation of the Clinical and Pathological Manifestations of Coronavirus Disease 2019 (COVID-19) in a Golden Syrian Hamster Model: Implications for Disease Pathogenesis and Transmissibility.

BACKGROUND: A physiological small-animal model that resembles COVID-19 with low mortality is lacking. METHODS: Molecular docking on the binding between angiotensin-converting enzyme 2 (ACE2) of common laboratory mammals and the receptor-binding domain of the surface spike protein of SARS-CoV-2 suggested that the golden Syrian hamster is an option. Virus challenge, contact transmission, and passive immunoprophylaxis studies were performed. Serial organ tissues and blood were harvested for histopathology, viral load and titer, chemokine/cytokine level, and neutralizing antibody titer. RESULTS: The Syrian hamster could be consistently infected by SARS-CoV-2. Maximal clinical signs of rapid breathing, weight loss, histopathological changes from the initial exudative phase of diffuse alveolar damage with extensive apoptosis to the later proliferative phase of tissue repair, airway and intestinal involvement with viral nucleocapsid protein expression, high lung viral load, and spleen and lymphoid atrophy associated with marked chemokine/cytokine activation were observed within the first week of virus challenge. The mean lung virus titer was between 105 and 107 TCID50/g. Challenged index hamsters consistently infected naive contact hamsters housed within the same cages, resulting in similar pathology but not weight loss. All infected hamsters recovered and developed mean serum neutralizing antibody titers ≥1:427 14 days postchallenge. Immunoprophylaxis with early convalescent serum achieved significant decrease in lung viral load but not in lung pathology. No consistent nonsynonymous adaptive mutation of the spike was found in viruses isolated from the infected hamsters. CONCLUSIONS: Besides satisfying Koch's postulates, this readily available hamster model is an important tool for studying transmission, pathogenesis, treatment, and vaccination against SARS-CoV-2.

Surgical Mask Partition Reduces the Risk of Noncontact Transmission in a Golden Syrian Hamster Model for Coronavirus Disease 2019 (COVID-19).

BACKGROUND: Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is believed to be mostly transmitted by medium- to large-sized respiratory droplets, although airborne transmission may be possible in healthcare settings involving aerosol-generating procedures. Exposure to respiratory droplets can theoretically be reduced by surgical mask usage. However, there is a lack of experimental evidence supporting surgical mask usage for prevention of COVID-19. METHODS: We used a well-established golden Syrian hamster SARS-CoV-2 model. We placed SARS-CoV-2-challenged index hamsters and naive hamsters into closed system units each comprising 2 different cages separated by a polyvinyl chloride air porous partition with unidirectional airflow within the isolator. The effect of a surgical mask partition placed between the cages was investigated. Besides clinical scoring, hamster specimens were tested for viral load, histopathology, and viral nucleocapsid antigen expression. RESULTS: Noncontact transmission was found in 66.7% (10/15) of exposed naive hamsters. Surgical mask partition for challenged index or naive hamsters significantly reduced transmission to 25% (6/24, P = .018). Surgical mask partition for challenged index hamsters significantly reduced transmission to only 16.7% (2/12, P = .019) of exposed naive hamsters. Unlike the severe manifestations of challenged hamsters, infected naive hamsters had lower clinical scores, milder histopathological changes, and lower viral nucleocapsid antigen expression in respiratory tract tissues. CONCLUSIONS: SARS-CoV-2 could be transmitted by respiratory droplets or airborne droplet nuclei which could be reduced by surgical mask partition in the hamster model. This is the first in vivo experimental evidence to support the possible benefit of surgical mask in prevention of COVID-19 transmission, especially when masks were worn by infected individuals.

Discovery of the FDA-approved drugs bexarotene, cetilistat, diiodohydroxyquinoline, and abiraterone as potential COVID-19 treatments with a robust two-tier screening system.

Coronavirus Disease 2019 (COVID-19) caused by the emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is associated with a crude case fatality rate of about 0.5-10 % depending on locality. A few clinically approved drugs, such as remdesivir, chloroquine, hydroxychloroquine, nafamostat, camostat, and ivermectin, exhibited anti-SARS-CoV-2 activity in vitro and/or in a small number of patients. However, their clinical use may be limited by anti-SARS-CoV-2 50 % maximal effective concentrations (EC50) that exceeded their achievable peak serum concentrations (Cmax), side effects, and/or availability. To find more immediately available COVID-19 antivirals, we established a two-tier drug screening system that combines SARS-CoV-2 enzyme-linked immunosorbent assay and cell viability assay, and applied it to screen a library consisting 1528 FDA-approved drugs. Cetilistat (anti-pancreatic lipase), diiodohydroxyquinoline (anti-parasitic), abiraterone acetate (synthetic androstane steroid), and bexarotene (antineoplastic retinoid) exhibited potent in vitro anti-SARS-CoV-2 activity (EC50 1.13-2.01 µM). Bexarotene demonstrated the highest Cmax:EC50 ratio (1.69) which was higher than those of chloroquine, hydroxychloroquine, and ivermectin. These results demonstrated the efficacy of the two-tier screening system and identified potential COVID-19 treatments which can achieve effective levels if given by inhalation or systemically depending on their pharmacokinetics.

Enhanced corrosion resistance of zinc-containing nanowires-modified titanium surface under exposure to oxidizing microenvironment.

Titanium (Ti) and its alloys as bio-implants have excellent biocompatibilities and osteogenic properties after modification of chemical composition and topography via various methods. The corrosion resistance of these modified materials is of great importance for changing oral system, while few researches have reported this point. Recently, oxidative corrosion induced by cellular metabolites has been well concerned. In this study, we explored the corrosion behaviors of four common materials (commercially pure Ti, cp-Ti; Sandblasting and acid etching-modified Ti, Ti-SLA; nanowires-modified Ti, Ti-NW; and zinc-containing nanowires-modified Ti, Ti-NW-Zn) with excellent biocompatibilities and osteogenic capacities under the macrophages induced-oxidizing microenvironment. The results showed that the materials immersed into a high oxidizing environment were more vulnerable to corrode. Meanwhile, different surfaces also showed various corrosion susceptibilities under oxidizing condition. Samples embed with zinc element exhibited more excellent corrosion resistance compared with other three surfaces exposure to excessive H2O2. Besides, we found that zinc-decorated Ti surfaces inhibited the adhesion and proliferation of macrophages on its surface and induced the M2 states of macrophages to better healing and tissue reconstruction. Most importantly, zinc-decorated Ti surfaces markedly increased the expressions of antioxidant enzyme relative genes in macrophages. It improved the oxidation microenvironment around the materials and further protected their properties. In summary, our results demonstrated that Ti-NW-Zn surfaces not only provided excellent corrosion resistance properties, but also inhibited the adhesion of macrophages. These aspects were necessary for maintaining osseointegration capacity and enhancing the corrosion resistance of Ti in numerous medical applications, particularly in dentistry.

An enhanced broad-spectrum peptide inhibits Omicron variants in vivo.

The continual emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) poses a major challenge to vaccines and antiviral therapeutics due to their extensive evasion of immunity. Aiming to develop potent and broad-spectrum anticoronavirus inhibitors, we generated A1-(GGGGS)7-HR2m (A1L35HR2m) by introducing an angiotensin-converting enzyme 2 (ACE2)-derived peptide A1 to the N terminus of the viral HR2-derived peptide HR2m through a long flexible linker, which showed significantly improved antiviral activity. Further cholesterol (Chol) modification at the C terminus of A1L35HR2m greatly enhanced the inhibitory activities against SARS-CoV-2, SARS-CoV-2 VOCs, SARS-CoV, and Middle East respiratory syndrome coronavirus (MERS-CoV) pseudoviruses, with IC50 values ranging from 0.16 to 5.53 nM. A1L35HR2m-Chol also potently inhibits spike-protein-mediated cell-cell fusion and the replication of authentic Omicron BA.2.12.1, BA.5, and EG.5.1. Importantly, A1L35HR2m-Chol distributed widely in respiratory tract tissue and had a long half-life (>10 h) in vivo. Intranasal administration of A1L35HR2m-Chol to K18-hACE2 transgenic mice potently inhibited Omicron BA.5 and EG.5.1 infection both prophylactically and therapeutically.

Mechanistic insights into bismuth(iii) inhibition of SARS-CoV-2 helicase.

The COVID-19 pandemic caused by SARS-CoV-2 resulted in a global public health crisis. In addition to vaccines, the development of effective therapy is highly desirable. Targeting a protein that plays a critical role in virus replication may allow pan-spectrum antiviral drugs to be developed. Among SARS-CoV-2 proteins, helicase (i.e., non-structural protein 13) is considered as a promising antiviral drug target due to its highly conserved sequence, unique structure and function. Herein, we demonstrate SARS-CoV-2 helicase as a target of bismuth-based antivirals in virus-infected mammalian cells by a metal-tagged antibody approach. To search for more potent bismuth-based antivirals, we further screened a panel of bismuth compounds towards inhibition of ATPase and DNA unwinding activity of nsp13 and identified a highly potent bismuth compound Bi(5-aminotropolonate)3, namely Bi(Tro-NH2)3 with an IC50 of 30 nM for ATPase. We show that bismuth-based compounds inhibited nsp13 unwinding activity via disrupting the binding of ATP and the DNA substrate to viral helicase. Binding of Bi(iii) to nsp13 also abolished the interaction between nsp12 and nsp13 as evidenced by immunofluorescence and co-immunoprecipitation assays. Finally, we validate our in vitro data in SARS-CoV-2 infected mammalian cells. Notably, Bi(6-TG)3 exhibited an EC50 of 1.18 ± 0.09 µM with a selective index of 847 in VeroE6-TMPRSS2 infected cells. This study highlights the important role of helicase for the development of more effective antiviral drugs to combat SARS-CoV-2 infection.

PMI-controlled mannose metabolism and glycosylation determines tissue tolerance and virus fitness.

Host survival depends on the elimination of virus and mitigation of tissue damage. Herein, we report the modulation of D-mannose flux rewires the virus-triggered immunometabolic response cascade and reduces tissue damage. Safe and inexpensive D-mannose can compete with glucose for the same transporter and hexokinase. Such competitions suppress glycolysis, reduce mitochondrial reactive-oxygen-species and succinate-mediated hypoxia-inducible factor-1α, and thus reduce virus-induced proinflammatory cytokine production. The combinatorial treatment by D-mannose and antiviral monotherapy exhibits in vivo synergy despite delayed antiviral treatment in mouse model of virus infections. Phosphomannose isomerase (PMI) knockout cells are viable, whereas addition of D-mannose to the PMI knockout cells blocks cell proliferation, indicating that PMI activity determines the beneficial effect of D-mannose. PMI inhibition suppress a panel of virus replication via affecting host and viral surface protein glycosylation. However, D-mannose does not suppress PMI activity or virus fitness. Taken together, PMI-centered therapeutic strategy clears virus infection while D-mannose treatment reprograms glycolysis for control of collateral damage.

SARS-CoV-2 infection activates inflammatory macrophages in vascular immune organoids.

SARS-CoV-2 provokes devastating tissue damage by cytokine release syndrome and leads to multi-organ failure. Modeling the process of immune cell activation and subsequent tissue damage is a significant task. Organoids from human tissues advanced our understanding of SARS-CoV-2 infection mechanisms though, they are missing crucial components: immune cells and endothelial cells. This study aims to generate organoids with these components. We established vascular immune organoids from human pluripotent stem cells and examined the effect of SARS-CoV-2 infection. We demonstrated that infections activated inflammatory macrophages. Notably, the upregulation of interferon signaling supports macrophages' role in cytokine release syndrome. We propose vascular immune organoids are a useful platform to model and discover factors that ameliorate SARS-CoV-2-mediated cytokine release syndrome.

Identification of the SARS-unique domain of SARS-CoV-2 as an antiviral target.

SARS-CoV-2 nsp3 is essential for viral replication and host responses. The SARS-unique domain (SUD) of nsp3 exerts its function through binding to viral and host proteins and RNAs. Herein, we show that SARS-CoV-2 SUD is highly flexible in solution. The intramolecular disulfide bond of SARS-CoV SUD is absent in SARS-CoV-2 SUD. Incorporating this bond in SARS-CoV-2 SUD allowed crystal structure determination to 1.35 Å resolution. However, introducing this bond in SARS-CoV-2 genome was lethal for the virus. Using biolayer interferometry, we screened compounds directly binding to SARS-CoV-2 SUD and identified theaflavin 3,3'-digallate (TF3) as a potent binder, Kd 2.8 µM. TF3 disrupted the SUD-guanine quadruplex interactions and exhibited anti-SARS-CoV-2 activity in Vero E6-TMPRSS2 cells with an EC50 of 5.9 µM and CC50 of 98.5 µM. In this work, we provide evidence that SARS-CoV-2 SUD harbors druggable sites for antiviral development.

Alisol B 23-acetate broadly inhibits coronavirus through blocking virus entry and suppresses proinflammatory T cells responses for the treatment of COVID-19.

INTRODUCTION: Emerging severe acute respiratory syndrome (SARS) coronavirus (CoV)-2 causes a global health disaster and pandemic. Seeking effective anti-pan-CoVs drugs benefit critical illness patients of coronavirus disease 2019 (COVID-19) but also may play a role in emerging CoVs of the future. OBJECTIVES: This study tested the hypothesis that alisol B 23-acetate could be a viral entry inhibitor and would have proinflammatory inhibition for COVID-19 treatment. METHODS: SARS-CoV-2 and its variants infected several cell lines were applied to evaluate the anti-CoVs activities of alisol B 23-aceate in vitro. The effects of alisol B 23-acetate on in vivo models were assessed by using SARS-CoV-2 and its variants challenged hamster and human angiotensin-converting enzyme 2 (ACE2) transgenic mice. The target of alisol B 23-acetate to ACE2 was analyzed using hydrogen/deuterium exchange (HDX) mass spectrometry (MS). RESULTS: Alisol B 23-acetate had inhibitory effects on different species of coronavirus. By using HDX-MS, we found that alisol B 23-acetate had inhibition potency toward ACE2. In vivo experiments showed that alisol B 23-acetate treatment remarkably decreased viral copy, reduced CD4+ T lymphocytes and CD11b+ macrophages infiltration and ameliorated lung damages in the hamster model. In Omicron variant infected human ACE2 transgenic mice, alisol B 23-acetate effectively alleviated viral load in nasal turbinate and reduced proinflammatory cytokines interleukin 17 (IL17) and interferon γ (IFNγ) in peripheral blood. The prophylactic treatment of alisol B 23-acetate by intranasal administration significantly attenuated Omicron viral load in the hamster lung tissues. Moreover, alisol B 23-acetate treatment remarkably inhibited proinflammatory responses through mitigating the secretions of IFNγ and IL17 in the cultured human and mice lymphocytes in vitro. CONCLUSION: Alisol B 23-acetate could be a promising therapeutic agent for COVID-19 treatment and its underlying mechanisms might be attributed to viral entry inhibition and anti-inflammatory activities.

Monovalent vaccination with inactivated SARS-CoV-2 BA.5 protects hamsters against Omicron but not non-Omicron variants.

We compared the protective effects of inactivated SARS-CoV-2 vaccines derived from the ancestral and the currently circulating BA.5.2 strains against infection with multiple variants in Syrian golden hamsters. Vaccination with BA.5.2 effectively protected against infection with the Omicron subvariants including XBB.1, but not the Alpha or Delta variant. In contrast, hamsters vaccinated with the ancestral strain demonstrated decent neutralization activity against both the Omicron and non-Omicron variants. Our findings might instruct future design and formulation of SARS-CoV-2 vaccines.

Broad-spectrum humanized monoclonal neutralizing antibody against SARS-CoV-2 variants, including the Omicron variant.

Introduction: Therapeutic monoclonal antibodies (mAbs) against the SARS-CoV-2 spike protein have been shown to improve the outcome of severe COVID-19 patients in clinical trials. However, novel variants with spike protein mutations can render many currently available mAbs ineffective. Methods: We produced mAbs by using hybridoma cells that generated from mice immunized with spike protein trimer and receptor binding domain (RBD). The panel of mAbs were screened for binding and neutralizing activity against different SARS-CoV-2 variants. The in vivo effectiveness of WKS13 was evaluated in a hamster model. Results: Out of 960 clones, we identified 18 mAbs that could bind spike protein. Ten of the mAbs could attach to RBD, among which five had neutralizing activity against the ancestral strain and could block the binding between the spike protein and human ACE2. One of these mAbs, WKS13, had broad neutralizing activity against all Variants of Concern (VOCs), including the Omicron variant. Both murine or humanized versions of WKS13 could reduce the lung viral load in hamsters infected with the Delta variant. Conclusions: Our data showed that broad-spectrum high potency mAbs can be produced from immunized mice, which can be used in humans after humanization of the Fc region. Our method represents a versatile and rapid strategy for generating therapeutic mAbs for upcoming novel variants.

Metal-coding assisted serological multi-omics profiling deciphers the role of selenium in COVID-19 immunity.

Uncovering how host metal(loid)s mediate the immune response against invading pathogens is critical for better understanding the pathogenesis mechanism of infectious disease. Clinical data show that imbalance of host metal(loid)s is closely associated with the severity and mortality of COVID-19. However, it remains elusive how metal(loid)s, which are essential elements for all forms of life and closely associated with multiple diseases if dysregulated, are involved in COVID-19 pathophysiology and immunopathology. Herein, we built up a metal-coding assisted multiplexed serological metallome and immunoproteome profiling system to characterize the links of metallome with COVID-19 pathogenesis and immunity. We found distinct metallome features in COVID-19 patients compared with non-infected control subjects, which may serve as a biomarker for disease diagnosis. Moreover, we generated the first correlation network between the host metallome and immunity mediators, and unbiasedly uncovered a strong association of selenium with interleukin-10 (IL-10). Supplementation of selenium to immune cells resulted in enhanced IL-10 expression in B cells and reduced induction of proinflammatory cytokines in B and CD4+ T cells. The selenium-enhanced IL-10 production in B cells was confirmed to be attributable to the activation of ERK and Akt pathways. We further validated our cellular data in SARS-CoV-2-infected K18-hACE2 mice, and found that selenium supplementation alleviated SARS-CoV-2-induced lung damage characterized by decreased alveolar inflammatory infiltrates through restoration of virus-repressed selenoproteins to alleviate oxidative stress. Our approach can be readily extended to other diseases to understand how the host defends against invading pathogens through regulation of metallome.

Development of Highly Potent Noncovalent Inhibitors of SARS-CoV-2 3CLpro.

The 3C-like protease (3CLpro) is an essential enzyme for the replication of SARS-CoV-2 and other coronaviruses and thus is a target for coronavirus drug discovery. Nearly all inhibitors of coronavirus 3CLpro reported so far are covalent inhibitors. Here, we report the development of specific, noncovalent inhibitors of 3CLpro. The most potent one, WU-04, effectively blocks SARS-CoV-2 replications in human cells with EC50 values in the 10-nM range. WU-04 also inhibits the 3CLpro of SARS-CoV and MERS-CoV with high potency, indicating that it is a pan-inhibitor of coronavirus 3CLpro. WU-04 showed anti-SARS-CoV-2 activity similar to that of PF-07321332 (Nirmatrelvir) in K18-hACE2 mice when the same dose was administered orally. Thus, WU-04 is a promising drug candidate for coronavirus treatment.

Effect of Different Adjuvants on Immune Responses Elicited by Protein-Based Subunit Vaccines against SARS-CoV-2 and Its Delta Variant.

The global pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become more serious because of the continuous emergence of variants of concern (VOC), thus calling for the development of broad-spectrum vaccines with greater efficacy. Adjuvants play important roles in enhancing the immunogenicity of protein-based subunit vaccines. In this study, we compared the effect of three adjuvants, including aluminum, nanoparticle manganese and MF59, on the immunogenicity of three protein-based COVID-19 vaccine candidates, including RBD-Fc, RBD and S-trimer. We found that the nanoparticle manganese adjuvant elicited the highest titers of SARS-CoV-2 RBD-specific IgG, IgG1 and IgG2a, as well as neutralizing antibodies against infection by pseudotyped SARS-CoV-2 and its Delta variant. What is more, the nanoparticle manganese adjuvant effectively reduced the viral load of the authentic SARS-CoV-2 and Delta variant in the cell culture supernatants. These results suggest that nanoparticle manganese, known to facilitate cGAS-STING activation, is an optimal adjuvant for protein-based COVID-19 subunit vaccines.