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1.
Antiviral Research ; : 105329, 2022.
Article in English | ScienceDirect | ID: covidwho-1819427

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the COVID-19 pandemic, has infected over 260 million people over the past 2 years. Remdesivir (RDV, VEKLURY®) is currently the only antiviral therapy fully approved by the FDA for the treatment of COVID-19. The parent nucleoside of RDV, GS-441524, exhibits antiviral activity against numerous respiratory viruses including SARS-CoV-2, although at reduced in vitro potency compared to RDV in most assays. Here we find in both human alveolar and bronchial primary cells, GS-441524 is metabolized to the pharmacologically active GS-441524 triphosphate (TP) less efficiently than RDV, which correlates with a lower in vitro SARS-CoV-2 antiviral activity. In vivo, African green monkeys (AGM) orally dosed with GS-441524 yielded low plasma levels due to limited oral bioavailability of <10%. When GS-441524 was delivered via intravenous (IV) administration, although plasma concentrations of GS-441524 were significantly higher, lung TP levels were lower than observed from IV RDV. To determine the required systemic exposure of GS-441524 associated with in vivo antiviral efficacy, SARS-CoV-2 infected AGMs were treated with a once-daily IV dose of either 7.5 or 20 mg/kg GS-441524 or IV RDV for 5 days and compared to vehicle control. Despite the reduced lung TP formation compared to IV dosing of RDV, daily treatment with IV GS-441524 resulted in dose-dependent efficacy, with the 20 mg/kg GS-441524 treatment resulting in significant reductions of SARS-CoV-2 replication in the lower respiratory tract of infected animals. These findings demonstrate the in vivo SARS-CoV-2 antiviral efficacy of GS-441524 and support evaluation of its orally bioavailable prodrugs as potential therapies for COVID-19.

2.
Cell Reports ; : 110829, 2022.
Article in English | ScienceDirect | ID: covidwho-1814236

ABSTRACT

Summary We report that SARS-CoV-2 Delta spike mutation P681R plays a key role in the Alpha-to-Delta variant replacement during the COVID-19 pandemic. Delta SARS-CoV-2 efficiently outcompetes the Alpha variant in human lung epithelial cells and primary human airway tissues. The Delta spike mutation P681R is located at a furin cleavage site that separates the spike 1 (S1) and S2 subunits. Reverting the P681R mutation to wild-type P681 significantly reduces the replication of Delta variant, to a level lower than the Alpha variant. Mechanistically, the Delta P681R mutation enhances the cleavage of the full-length spike to S1 and S2, which could improve cell surface-mediated virus entry. In contrast, the Alpha spike also has a mutation at the same amino acid (P681H), but the cleavage of Alpha spike is reduced compared to the Delta spike. Our results suggest P681R as a key mutation in enhancing Delta variant replication via increased S1/S2 cleavage.

3.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-334196

ABSTRACT

Summary The worldwide spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to the repeated emergence of variants of concern. The Omicron variant has two dominant sub-lineages, BA.1 and BA.2, each with unprecedented numbers of nonsynonymous and indel spike protein mutations: 33 and 29, respectively. Some of these mutations individually increase transmissibility and enhance immune evasion, but their interactions within the Omicron mutational background is unknown. We characterize the molecular effects of all Omicron spike mutations on expression, human ACE2 receptor affinity, and neutralizing antibody recognition. We show that key mutations enable escape from neutralizing antibodies at a variety of epitopes. Stabilizing mutations in the N-terminal and S2 domains of the spike protein compensate for destabilizing mutations in the receptor binding domain, thereby enabling the record number of mutations in Omicron sub-lineages. Taken together, our results provide a comprehensive account of the mutational effects in the Omicron spike protein and illuminate previously unknown mechanisms of how the N-terminal domain can compensate for destabilizing mutations within the more evolutionarily constrained RBD.

6.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-333143

ABSTRACT

The continuous emergence of SARS-CoV-2 variants with increased transmission and immune evasion has caused breakthrough infections in vaccinated population. It is important to determine the threshold of neutralizing antibody titers that permit breakthrough infections. Here we tested the neutralization titers of vaccinated patients who contracted Delta variant. All 75 patients with Delta breakthrough infections exhibited neutralization titers (NT50) of less than 70. Among the breakthrough patients, 76%, 18.7%, and 5.3% of them had the NT50 ranges of <20, 20-50, and 50-69, respectively. These clinical laboratory results have implications in vaccine strategy and public health policy.

7.
NPJ Vaccines ; 7(1): 41, 2022 Apr 08.
Article in English | MEDLINE | ID: covidwho-1783982

ABSTRACT

BNT162b2-elicited human sera neutralize the currently dominant Delta SARS-CoV-2 variant. Here, we report the ability of 20 human sera, drawn 2 or 4 weeks after two doses of BNT162b2, to neutralize USA-WA1/2020 SARS-CoV-2 bearing variant spikes from Delta plus (Delta-AY.1, Delta-AY.2), Delta-∆144 (Delta with the Y144 deletion of the Alpha variant), Lambda, B.1.1.519, Theta, and Mu lineage viruses. Geometric mean plaque reduction neutralization titers against Delta-AY.1, Delta-AY.2, and Mu viruses are slightly lower than against USA-WA1/2020, but all sera neutralize the variant viruses to titers of ≥80, and neutralization titers against the Delta-∆144, Lambda, B.1.1.519 and Theta variants not significantly reduced relative to those against USA-WA1/2020. The susceptibility of Delta plus, Lambda, B.1.1.519, Theta, Mu, and other variants to neutralization by the sera indicates that antigenic change has not led to virus escape from vaccine-elicited neutralizing antibodies and supports ongoing mass immunization with BNT162b2 to control the variants and to minimize the emergence of new variants.

8.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-332769

ABSTRACT

The Omicron SARS-CoV-2 has three distinct sublineages, among which sublineage BA.1 is responsible for the initial Omicron surge and is now being replaced by BA.2 world-wide, whereas BA.3 is currently at a low frequency. The ongoing BA.1-to-BA.2 replacement underscores the importance to understand the cross-neutralization among the three Omicron sublineages. Here we tested the neutralization of BA.1-infected human sera against BA.2, BA.3, and USA/WA1-2020 (a strain isolated in late January 2020). The BA.1-infected sera neutralized BA.1, BA.2, BA.3, and USA/WA1-2020 SARS-CoV-2s with geometric mean titers (GMTs) of 445, 107, 102, and 16, respectively. Thus, the neutralizing GMTs against heterologous BA.2, BA.3, and USA/WA1-2020 were 4.2-, 4.4-, and 28.4-fold lower than the GMT against homologous BA.1, respectively. These findings have implications in COVID-19 vaccine strategy.

9.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-331542

ABSTRACT

The newly emerged Omicron SARS-CoV-2 has 3 distinct sublineages: BA.1, BA.2, and BA.3. BA.1 accounts for the initial surge and is being replaced by BA.2, whereas BA.3 is at a low prevalence at this time. Here we report the neutralization of BNT162b2-vaccinated sera (collected at 1 month after dose 3) against the three Omicron sublineages. To facilitate the neutralization testing, we engineered the complete BA.1, BA.2, or BA.3 spike into an mNeonGreen USA-WA1/2020 SRAS-CoV-2. All BNT162b2-vaccinated sera neutralized USA-WA1/2020, BA.1-, BA.2-, and BA.3-spike SARS-CoV-2s with titers of >20;the neutralization geometric mean titers (GMTs) against the four viruses were 1211, 336, 300, and 190, respectively. Thus, the BA.1-, BA.2-, and BA.3-spike SARS-CoV-2s were 3.6-, 4.0-, and 6.4-fold less efficiently neutralized than the USA-WA1/2020, respectively. Our data have implications in vaccine strategy and understanding the biology of Omicron sublineages.

10.
Zoonoses (Burlingt) ; 2(1)2022.
Article in English | MEDLINE | ID: covidwho-1743023

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused global destruction since its emergence in late 2019. Over the past two years, the virus has continuously evolved in human host, leading to emergence of variants with changed viral transmission, disease severity, and evasion of immunity. Although vaccines have been developed for the coronavirus disease 2019 (COVID-19) at an unprecedently pace, the variants have constantly posed threats to the effectiveness of the approved vaccines. In this short communication, we review the key variants and discuss their implications in viral replication, transmission, and immune evasion.

11.
Cell Rep ; 38(10): 110434, 2022 03 08.
Article in English | MEDLINE | ID: covidwho-1729611

ABSTRACT

Type I interferons (IFN-I) are essential to establish antiviral innate immunity. Unanchored (or free) polyubiquitin (poly-Ub) has been shown to regulate IFN-I responses. However, few unanchored poly-Ub interactors are known. To identify factors regulated by unanchored poly-Ub in a physiological setting, we developed an approach to isolate unanchored poly-Ub from lung tissue. We identified the RNA helicase DHX16 as a potential pattern recognition receptor (PRR). Silencing of DHX16 in cells and in vivo diminished IFN-I responses against influenza virus. These effects extended to members of other virus families, including Zika and SARS-CoV-2. DHX16-dependent IFN-I production requires RIG-I and unanchored K48-poly-Ub synthesized by the E3-Ub ligase TRIM6. DHX16 recognizes a signal in influenza RNA segments that undergo splicing and requires its RNA helicase motif for direct, high-affinity interactions with specific viral RNAs. Our study establishes DHX16 as a PRR that partners with RIG-I for optimal activation of antiviral immunity requiring unanchored poly-Ub.


Subject(s)
DEAD Box Protein 58 , Interferon Type I , RNA Helicases , RNA, Viral , Receptors, Immunologic , Zika Virus Infection , Zika Virus , COVID-19 , DEAD Box Protein 58/immunology , Humans , Immunity, Innate , Interferon Type I/immunology , RNA Helicases/immunology , Receptors, Immunologic/immunology , SARS-CoV-2 , Tripartite Motif Proteins , Zika Virus/genetics , Zika Virus Infection/immunology
12.
Cell Host Microbe ; 30(4): 485-488.e3, 2022 Apr 13.
Article in English | MEDLINE | ID: covidwho-1693797

ABSTRACT

Two doses of the BNT162b2 mRNA vaccine are highly effective against SARS-CoV-2. Here, we tested the antibody neutralization against Omicron SARS-CoV-2 after 2 and 3 doses of BNT162b2. Serum from vaccinated individuals was serially tested for its ability to neutralize wild-type SARS-CoV-2 (USA-WA1/2020) and an engineered USA-WA1/2020 bearing the Omicron spike glycoprotein. At 2 or 4 weeks post dose 2, the neutralization geometric mean titers (GMTs) against the wild-type and Omicron-spike viruses were 511 and 20, respectively; at 1 month post dose 3, the neutralization GMTs increased to 1,342 and 336; and at 4 months post dose 3, the neutralization GMTs decreased to 820 and 171. The data support a 3-dose vaccination strategy and provide a glimpse into the durability of the neutralization response against Omicron.

13.
EuropePMC;
Preprint in English | EuropePMC | ID: ppcovidwho-327679

ABSTRACT

We report a live-attenuated SARS-CoV-2 vaccine candidate with (i) re-engineered viral transcriptional regulator sequences and (ii) deleted open-reading-frames (ORF) 3, 6, 7, and 8 (Δ3678). The Δ3678 virus replicates about 7,500-fold lower than wild-type SARS-CoV-2 on primary human airway cultures, but restores its replication on interferon-deficient Vero-E6 cells that are approved for vaccine production. The Δ3678 virus is highly attenuated in both hamster and K18-hACE2 mouse models. A single-dose immunization of the Δ3678 virus protects hamsters from wild-type virus challenge and transmission. Among the deleted ORFs in the Δ3678 virus, ORF3a accounts for the most attenuation through antagonizing STAT1 phosphorylation during type-I interferon signaling. We also developed an mNeonGreen reporter Δ3678 virus for high-throughput neutralization and antiviral testing. Altogether, the results suggest that Δ3678 SARS-CoV-2 may serve as a live-attenuated vaccine candidate and a research tool for potential biosafety level-2 use.

14.
EuropePMC;
Preprint in English | EuropePMC | ID: ppcovidwho-327574

ABSTRACT

Genetic variation of SARS-CoV-2 has resulted in the emergence and rapid spread of multiple variants throughout the pandemic, of which Omicron is currently the predominant variant circulating worldwide. SARS-CoV-2 variants of concern or interest (VOC/VOI) have evidence of increased viral transmission, disease severity, or decreased effectiveness of vaccines and neutralizing antibodies. Remdesivir (RDV, VEKLURY ® ) is a nucleoside analog prodrug and the first FDA-approved antiviral treatment of COVID-19. Here we present a comprehensive antiviral activity assessment of RDV and its parent nucleoside, GS-441524, against 10 current and former SARS-CoV-2 VOC/VOI clinical isolates by nucleoprotein ELISA and plaque reduction assay. Delta and Omicron variants remained susceptible to RDV and GS-441524, with EC 50 values 0.31 to 0.62-fold of those observed against the ancestral WA1 isolate. All other tested variants exhibited EC 50 values ranging from 0.15 to 2.3-fold of the observed EC 50 values against WA1. Analysis of nearly 6 million publicly available variant isolate sequences confirmed that Nsp12, the RNA-dependent RNA polymerase (RdRp) target of RDV and GS-441524, is highly conserved across variants with only 2 prevalent changes (P323L and G671S). Using recombinant viruses, both RDV and GS-441524 retained potency against all viruses containing frequent variant substitutions or their combination. Taken together, these results highlight the conserved nature of SARS-CoV-2 Nsp12 and provide evidence of sustained SARS-CoV-2 antiviral activity of RDV and GS-441524 across the tested variants. The observed pan-variant activity of RDV supports its continued use for the treatment of COVID-19 regardless of the SARS-CoV-2 variant.

15.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-321376

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to evolve around the world, generating new variants that are of concern based on their potential for altered transmissibility, pathogenicity, and coverage by vaccines and therapeutics. Here we report that 20 BNT162b2 vaccine-elicited human sera neutralize engineered SARS-CoV-2 with a USA-WA1/2020 genetic background (a virus strain isolated in January 2020) and spike glycoproteins from the newly emerged B.1.617.1 (first identified in India) or B.1.525 (first identified in Nigeria) lineages. Geometric mean plaque reduction neutralization titers against the variant viruses, particularly the B.1.617.1 variant, are lower than the titer against USA-WA1/2020 virus, but all sera tested neutralize the variant viruses at titers of at least 40. The susceptibility of the newly emerged B.1.617.1 and B.1.525 variants to BNT162b2 vaccine-elicited neutralization supports mass immunization as a central strategy to end the COVID-19 pandemic across geographies.

16.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-309974

ABSTRACT

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) messenger RNA (mRNA)-based vaccines are ~95% effective in preventing coronavirus disease 2019. However, the dynamics of antibody secreting plasmablasts (PBs) and germinal centre (GC) B cells induced by these vaccines in SARS-CoV-2 naïve and antigen-experienced humans remains unclear. Here we examined peripheral blood and/or lymph node (LN) antigen-specific B cell responses in 32 individuals who received two doses of BNT162b2, an mRNA-based vaccine encoding the full-length SARS-CoV-2 spike (S) gene. Circulating IgG- and IgA-secreting PBs targeting the S protein peaked one week after the second immunization then declined and were undetectable three weeks later. PB responses coincided with maximal levels of serum anti-S binding and neutralizing antibodies to a historical strain as well as emerging variants, especially in individuals previously infected with SARS-CoV-2, who produced the most robust serological responses. Fine needle aspirates of draining axillary LNs identified GC B cells that bind S protein in all participants sampled after primary immunization. GC responses increased after boosting and were detectable in two distinct LNs in several participants. Remarkably, high frequencies of S-binding GC B cells and PBs were maintained in draining LNs for up to seven weeks after first immunization, with a substantial fraction of the PB pool class-switched to IgA. GC B cell-derived monoclonal antibodies predominantly targeted the RBD, with fewer clones binding to the N-terminal domain or shared epitopes within the S proteins of human betacoronaviruses OC43 and HKU1. Our studies demonstrate that SARS-CoV-2 mRNA-based vaccination of humans induces a robust and persistent GC B cell response that engages pre-existing as well as new B cell clones, which enables generation of high-affinity, broad, and durable humoral immunity.

17.
Nat Commun ; 13(1): 852, 2022 02 09.
Article in English | MEDLINE | ID: covidwho-1684027

ABSTRACT

The spread of the Omicron SARS-CoV-2 variant underscores the importance of analyzing the cross-protection from previous non-Omicron infection. We have developed a high-throughput neutralization assay for Omicron SARS-CoV-2 by engineering the Omicron spike gene into an mNeonGreen USA-WA1/2020 SARS-CoV-2 (isolated in January 2020). Using this assay, we determine the neutralization titers (defined as the maximal serum dilution that inhibited 50% of infectious virus) of patient sera collected at 1- or 6-months after infection with non-Omicron SARS-CoV-2. From 1- to 6-month post-infection, the neutralization titers against USA-WA1/2020 decrease from 601 to 142 (a 4.2-fold reduction), while the neutralization titers against Omicron-spike SARS-CoV-2 remain low at 38 and 32, respectively. Thus, at 1- and 6-months after non-Omicron SARS-CoV-2 infection, the neutralization titers against Omicron are 15.8- and 4.4-fold lower than those against USA-WA1/2020, respectively. The low cross-neutralization against Omicron from previous non-Omicron infection supports vaccination of formerly infected individuals to mitigate the health impact of the ongoing Omicron surge.


Subject(s)
Antibodies, Viral/immunology , COVID-19/immunology , SARS-CoV-2/immunology , Antibodies, Neutralizing/immunology , COVID-19/blood , COVID-19/virology , Cross Reactions , Humans , Neutralization Tests , Reinfection/blood , Reinfection/immunology , Reinfection/virology , SARS-CoV-2/genetics
18.
mBio ; : e0337721, 2022 Jan 18.
Article in English | MEDLINE | ID: covidwho-1637923

ABSTRACT

Pathogenic coronaviruses are a major threat to global public health. Here, using a recombinant reporter virus-based compound screening approach, we identified small-molecule inhibitors that potently block the replication of severe acute respiratory syndrome virus 2 (SARS-CoV-2). Among them, JIB-04 inhibited SARS-CoV-2 replication in Vero E6 cells with a 50% effective concentration of 695 nM, with a specificity index of greater than 1,000. JIB-04 showed in vitro antiviral activity in multiple cell types, including primary human bronchial epithelial cells, against several DNA and RNA viruses, including porcine coronavirus transmissible gastroenteritis virus. In an in vivo porcine model of coronavirus infection, administration of JIB-04 reduced virus infection and associated tissue pathology, which resulted in improved weight gain and survival. These results highlight the potential utility of JIB-04 as an antiviral agent against SARS-CoV-2 and other viral pathogens. IMPORTANCE The coronavirus disease 2019 (COVID-19), the disease caused by SARS-CoV-2 infection, is an ongoing public health disaster worldwide. Although several vaccines are available as a preventive measure and the FDA approval of an orally bioavailable drug is on the horizon, there remains a need for developing antivirals against SARS-CoV-2 that could work on the early course of infection. By using infectious reporter viruses, we screened small-molecule inhibitors for antiviral activity against SARS-CoV-2. Among the top hits was JIB-04, a compound previously studied for its anticancer activity. Here, we showed that JIB-04 inhibits the replication of SARS-CoV-2 as well as different DNA and RNA viruses. Furthermore, JIB-04 conferred protection in a porcine model of coronavirus infection, although to a lesser extent when given as therapeutic rather than prophylactic doses. Our findings indicate a limited but still promising utility of JIB-04 as an antiviral agent in the combat against COVID-19 and potentially other viral diseases.

19.
Emerg Microbes Infect ; 11(1): 208-211, 2022 Dec.
Article in English | MEDLINE | ID: covidwho-1598042

ABSTRACT

We performed an annotation of 35 mutations in the spike protein of the SARS-CoV-2 Omicron variant. Our analysis of the mutations indicates that Omicron has gained prominent immune evasion and potential for enhanced transmissibility. Previous modeling study has revealed that continued evolution in both immune evasion and enhanced transmissibility by SARS-CoV-2 would compromise vaccines as tools for the pandemic control. To combat the future variants of SARS-CoV-2, the world needs novel antiviral drugs that are effective at curb viral spreading without introducing additional selective pressure towards resistant variants.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , COVID-19/drug therapy , Drug Design/methods , Receptors, Virus/metabolism , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/immunology , Antiviral Agents/chemical synthesis , Antiviral Agents/therapeutic use , Binding Sites , COVID-19/epidemiology , COVID-19/prevention & control , COVID-19/transmission , COVID-19 Vaccines/administration & dosage , COVID-19 Vaccines/adverse effects , Humans , Immune Evasion , Mutation , Protein Binding , Receptors, Virus/genetics , Receptors, Virus/immunology , SARS-CoV-2/drug effects , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism
20.
Mol Ther ; 2022 Jan 03.
Article in English | MEDLINE | ID: covidwho-1586240

ABSTRACT

RNA vaccines have demonstrated efficacy against SARS-CoV-2 in humans, and the technology is being leveraged for rapid emergency response. In this report, we assessed immunogenicity and, for the first time, toxicity, biodistribution, and protective efficacy in preclinical models of a two-dose self-amplifying messenger RNA (SAM) vaccine, encoding a prefusion-stabilized spike antigen of SARS-CoV-2 Wuhan-Hu-1 strain and delivered by lipid nanoparticles (LNPs). In mice, one immunization with the SAM vaccine elicited a robust spike-specific antibody response, which was further boosted by a second immunization, and effectively neutralized the matched SARS-CoV-2 Wuhan strain as well as B.1.1.7 (Alpha), B.1.351 (Beta) and B.1.617.2 (Delta) variants. High frequencies of spike-specific germinal center B, Th0/Th1 CD4, and CD8 T cell responses were observed in mice. Local tolerance, potential systemic toxicity, and biodistribution of the vaccine were characterized in rats. In hamsters, the vaccine candidate was well-tolerated, markedly reduced viral load in the upper and lower airways, and protected animals against disease in a dose-dependent manner, with no evidence of disease enhancement following SARS-CoV-2 challenge. Therefore, the SARS-CoV-2 SAM (LNP) vaccine candidate has a favorable safety profile, elicits robust protective immune responses against multiple SARS-CoV-2 variants, and has been advanced to phase 1 clinical evaluation (NCT04758962).

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