SARS-CoV-2 Binding and Neutralization Properties of Peptides Derived from N-Terminus of Human ACE2.

The binding properties of synthetic and recombinant peptides derived from N-terminal part of ACE2, the main receptor for SARS-CoV-2, were evaluated. Additionally, the ability of these peptides to prevent virus entry in vitro was addressed using both pseudovirus particles decorated with the S protein, as well as through infection of Vero cells with live SARS-CoV-2 virus. Surprisingly, in spite of effective binding to S protein, all linear peptides of various lengths failed to neutralize the viral infection in vitro. However, the P1st peptide that was chemically "stapled" in order to stabilize its alpha-helical structure was able to interfere with virus entry into ACE2-expressing cells. Interestingly, this peptide also neutralized pseudovirus particles decorated with S protein derived from the Omicron BA.1 virus, in spite of variations in key amino acid residues contacting ACE2.

Towards effective COVID­19 vaccines: Updates, perspectives and challenges (Review).

In the current context of the pandemic triggered by SARS-COV-2, the immunization of the population through vaccination is recognized as a public health priority. In the case of SARS­COV­2, the genetic sequencing was done quickly, in one month. Since then, worldwide research has focused on obtaining a vaccine. This has a major economic impact because new technological platforms and advanced genetic engineering procedures are required to obtain a COVID­19 vaccine. The most difficult scientific challenge for this future vaccine obtained in the laboratory is the proof of clinical safety and efficacy. The biggest challenge of manufacturing is the construction and validation of production platforms capable of making the vaccine on a large scale.

5-(Perylen-3-ylethynyl)uracil as an antiviral scaffold: Potent suppression of enveloped virus reproduction by 3-methyl derivatives in vitro.

Amphipathic nucleoside and non-nucleoside derivatives of pentacyclic aromatic hydrocarbon perylene are known as potent non-cytotoxic broad-spectrum antivirals. Here we report 3-methyl-5-(perylen-3-ylethynyl)-uracil-1-acetic acid and its amides, a new series of compounds based on a 5-(perylen-3-ylethynyl)-uracil scaffold. The compounds demonstrate pronounced in vitro activity against arthropod-borne viruses, namely tick-borne encephalitis virus (TBEV) and yellow fever virus (YFV), in plaque reduction assays with EC50 values below 1.9 and 1.3 nM, respectively, and Chikungunya virus (CHIKV) in cytopathic effect inhibition test with EC50 values below 3.2 µM. The compounds are active against respiratory viruses as well: severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) in cytopathic effect inhibition test and influenza A virus (IAV) in virus titer reduction experiments are inhibited - EC50 values below 51 nM and 2.2 µM, respectively. The activity stems from the presence of a hydrophobic perylene core, and all of the synthesized compounds exhibit comparable 1O2 generation rates. Nonetheless, activity can vary by orders of magnitude depending on the hydrophilic part of the molecule, suggesting a complex mode of action. A time-of-addition experiment and fluorescent imaging indicate that the compounds inhibit viral fusion in a dose-dependent manner. The localization of the compound in the lipid bilayers and visible damage to the viral envelope suggest the membrane as the primary target. Dramatic reduction of antiviral activity with limited irradiation or under treatment with antioxidants further cements the idea of photoinduced ROS-mediated viral envelope damage being the mode of antiviral action.

Immunogenicity and protectivity of intranasally delivered vector-based heterologous prime-boost COVID-19 vaccine Sputnik V in mice and non-human primates.

Although unprecedented efforts aiming to stop the COVID-19 pandemic have been made over the past two years, SARSCoV-2 virus still continues to cause intolerable health and economical losses. Vaccines are considered the most effective way to prevent infectious diseases, which has been reaffirmed for COVID-19. However, in the context of the continuing virus spread because of insufficient vaccination coverage and emergence of new variants of concern, there is a high demand for vaccination strategy amendment. The ability to elicit protective immunity at the entry gates of infection provided by mucosal vaccination is key to block virus infection and transmission. Therefore, these mucosal vaccines are believed to be a "silver bullet" that could bring the pandemic to an end. Here, we demonstrate that the intranasally delivered Gam-COVID-Vac (Sputnik V) vaccine induced a robust (no less than 180 days) systemic and local immune response in mice. High immunogenic properties of the vaccine were verified in non-human primates (common marmosets) by marked IgG and neutralizing antibody (NtAb) production in blood serum, antigen-specific Tcell proliferation and cytokine release of peripheral blood mononuclear cells accompanied by formation of IgA antibodies in the nasal mucosa. We also demonstrate that Sputnik V vaccine can provide sterilizing immunity in K18-hACE2 transgenic mice exposed to experimental lethal SARS-CoV-2 infection protecting them against severe lung immunopathology and mortality. We believe that intranasal Sputnik V vaccine is a promising novel needle-free mucosal vaccine candidate for primary immunization as well as for revaccination and is worth further clinical investigation.

Methodology of Purification of Inactivated Cell-Culture-Grown SARS-CoV-2 Using Size-Exclusion Chromatography.

Various types of COVID-19 vaccines, including adenovirus, mRNA, and inactivated ones, have been developed and approved for clinical use worldwide. Inactivated vaccines are produced using a proven technology that is widely used for the production of vaccines for the prevention and control of infectious diseases, including influenza and poliomyelitis. The development of inactivated whole-virion vaccines commonly includes several stages: the production of cellular and viral biomass in cell culture; inactivation of the virus; filtration and ultrafiltration; chromatographic purification of the viral antigen; and formulation with stabilizers and adjuvants. In this study, the suitability of four resins for Size-Exclusion Chromatography was investigated for the purification of a viral antigen for the human COVID-19 vaccine.

Vaccination With Oral Polio Vaccine Reduces COVID-19 Incidence.

Background: Effective response to emerging pandemic threats is complicated by the need to develop specific vaccines and other medical products. The availability of broadly specific countermeasures that could be deployed early in the pandemic could significantly alter its course and save countless lives. Live attenuated vaccines (LAVs) were shown to induce non-specific protection against a broad spectrum of off-target pathogens by stimulating innate immune responses. The purpose of this study was to evaluate the effect of immunization with bivalent Oral Poliovirus Vaccine (bOPV) on the incidence of COVID-19 and other acute respiratory infections (ARIs). Methods and Findings: A randomized parallel-group comparative study was conducted in Kirov Medical University. 1115 healthy volunteers aged 18 to 65 were randomized into two equal groups, one of which was immunized orally with a single dose of bOPV "BiVac Polio" and another with placebo. The study participants were monitored for three months for respiratory illnesses including COVID-19. The endpoint was the incidence of acute respiratory infections and laboratory confirmed COVID-19 in both groups during 3 months after immunization. The number of laboratory-confirmed cases of COVID-19 was significantly lower in the vaccinated group than in placebo (25 cases vs. 44, p=0.036). The difference between the overall number of clinically diagnosed respiratory illnesses in the two groups was not statistically significant. Conclusions: Immunization with bOPV reduced the number of laboratory-confirmed COVID-19 cases, consistent with the original hypothesis that LAVs induce non-specific protection against off-target infections. The findings are in line with previous observations of the protective effects of OPV against seasonal influenza and other viral and bacterial pathogens. The absence of a statistically significant effect on the total number of ARIs may be due to the insufficient number of participants and heterogeneous etiology of ARIs. OPV could be used to complement specific coronavirus vaccines, especially in regions of the world where the vaccines are unavailable, and as a stopgap measure for urgent response to future emerging infections. Clinical trial registration number NCT05083039 at clinicaltrals.gov https://clinicaltrials.gov/ct2/show/NCT05083039?term=NCT05083039&draw=2&rank=1.

Pre-Steady-State Kinetics of the SARS-CoV-2 Main Protease as a Powerful Tool for Antiviral Drug Discovery.

The design of effective target-specific drugs for COVID-19 treatment has become an intriguing challenge for modern science. The SARS-CoV-2 main protease, Mpro, responsible for the processing of SARS-CoV-2 polyproteins and production of individual components of viral replication machinery, is an attractive candidate target for drug discovery. Specific Mpro inhibitors have turned out to be promising anticoronaviral agents. Thus, an effective platform for quantitative screening of Mpro-targeting molecules is urgently needed. Here, we propose a pre-steady-state kinetic analysis of the interaction of Mpro with inhibitors as a basis for such a platform. We examined the kinetic mechanism of peptide substrate binding and cleavage by wild-type Mpro and by its catalytically inactive mutant C145A. The enzyme induces conformational changes of the peptide during the reaction. The inhibition of Mpro by boceprevir, telaprevir, GC-376, PF-00835231, or thimerosal was investigated. Detailed pre-steady-state kinetics of the interaction of the wild-type enzyme with the most potent inhibitor, PF-00835231, revealed a two-step binding mechanism, followed by covalent complex formation. The C145A Mpro mutant interacts with PF-00835231 approximately 100-fold less effectively. Nevertheless, the binding constant of PF-00835231 toward C145A Mpro is still good enough to inhibit the enzyme. Therefore, our results suggest that even noncovalent inhibitor binding due to a fine conformational fit into the active site is sufficient for efficient inhibition. A structure-based virtual screening and a subsequent detailed assessment of inhibition efficacy allowed us to select two compounds as promising noncovalent inhibitor leads of SARS-CoV-2 Mpro.

SARS-CoV-2 infection in children in Moscow in 2020: clinical features and impact on circulation of other respiratory viruses: SARS-CoV-2 infection in children in Moscow in 2020.

OBJECTIVES: This study aimed to estimate the impact of the COVID-19 pandemic on the circulation of non-SARS-CoV-2 respiratory viruses and the clinical characteristics of COVID-19 in hospitalized children. METHODS: A total of 226 and 864 children admitted to the Children's City Clinical Hospital with acute respiratory infection in September to November of 2018 and 2020 in Moscow were tested for respiratory viruses using multiplex polymerase chain reaction (PCR) and Mycoplasma pneumoniae/Chlamydia pneumoniae using enzyme-linked immunosorbent assay. RESULTS: The detection rate of non-SARS-CoV-2 viruses in 2020 was lower than in 2018, 16.9% versus 37.6%. An increase in the median age of children with respiratory viruses was observed during the pandemic (3 years vs 1 year). There was no significant difference in the frequency of intensive care unit (ICU) admission in children with SARS-CoV-2 and other respiratory virus infections (2.7% vs 2.9%). SARS-CoV-2 and human rhinoviruses, human metapneumoviruses, and human adenoviruses showed significantly lower than expected co-detection rates during co-circulation. An increase in body mass index (BMI) or bacterial coinfection leads to an increased risk of ICU admission and a longer duration of COVID-19 in children. CONCLUSIONS: The COVID-19 pandemic led to significant changes in the epidemiological characteristics of non-SARS-CoV-2 respiratory viruses during the autumn peak of the 2020 pandemic, compared with the same period in 2018.

Structural characterization of β-propiolactone inactivated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) particles

Abstract The severe COVID-19 pandemic drives the research toward the SARS-CoV-2 virion structure and the possible therapies against it. Here, we characterized the ?-propiolactone inactivated SARS-CoV-2 virions using transmission electron microscopy (TEM) and atomic force microscopy (AFM). We compared the SARS-CoV-2 samples purified by two consecutive chromatographic procedures (size exclusion chromatography [SEC], followed by ion-exchange chromatography [IEC]) with samples purified by ultracentrifugation. The samples prepared using SEC and IEC retained more spikes on the surface than the ones prepared using ultracentrifugation, as confirmed by TEM and AFM. TEM showed that the spike (S) proteins were in the pre-fusion conformation. Notably, the S proteins could be recognized by specific monoclonal antibodies. Analytical TEM showed that the inactivated virions retained nucleic acid. Altogether, we demonstrated that the inactivated SARS-CoV-2 virions retain the structural features of native viruses and provide a prospective vaccine candidate.

Long-term humoral immunogenicity, safety and protective efficacy of inactivated vaccine against COVID-19 (CoviVac) in preclinical studies.

The unprecedented in recent history global COVID-19 pandemic urged the implementation of all existing vaccine platforms to ensure the availability of the vaccines against COVID-19 to every country in the world. Despite the multitude of high-quality papers describing clinical trials of different vaccine products, basic detailed data on general toxicity, reproductive toxicity, immunogenicity, protective efficacy and durability of immune response in animal models are scarce. Here, we developed a ß-propiolactone-inactivated whole virion vaccine CoviVac and assessed its safety, protective efficacy, immunogenicity and stability of the immune response in rodents and non-human primates. The vaccine showed no signs of acute/chronic, reproductive, embryo- and fetotoxicity, or teratogenic effects, as well as no allergenic properties in studied animal species. The vaccine induced stable and robust humoral immune response both in form of specific anti-SARS-CoV-2 IgG and NAbs in mice, Syrian hamsters, and common marmosets. The NAb levels did not decrease significantly over the course of one year. The course of two immunizations protected Syrian hamsters from severe pneumonia upon intranasal challenge with the live virus. Robustness of the vaccine manufacturing process was demonstrated as well. These data encouraged further evaluation of CoviVac in clinical trials.

EGCG as an anti-SARS-CoV-2 agent: Preventive versus therapeutic potential against original and mutant virus.

In the search for anti-SARS-CoV-2 drugs, much attention is given to safe and widely available native compounds. The green tea component epigallocatechin 3 gallate (EGCG) is particularly promising because it reportedly inhibits viral replication and viral entry in vitro. However, conclusive evidence for its predominant activity is needed. We tested EGCG effects on the native virus isolated from COVID-19 patients in two independent series of experiments using VERO cells and two different treatment schemes in each series. The results confirmed modest cytotoxicity of EGCG and its substantial antiviral activity. The preincubation scheme aimed at infection prevention has proven particularly beneficial. We complemented that finding with a detailed investigation of EGCG interactions with viral S-protein subunits, including S2, RBD, and the RBD mutant harboring the N501Y mutation. Molecular modeling experiments revealed N501Y-specific stacking interactions in the RBD-ACE2 complex and provided insight into EGCG interference with the complex formation. Together, these findings provide a molecular basis for the observed EGCG effects and reinforce its prospects in COVID-19 prevention therapy.

Phenoxazine nucleoside derivatives with a multiple activity against RNA and DNA viruses.

Emerging and re-emerging viruses periodically cause outbreaks and epidemics all over the world, eventually leading to global events such as the current pandemic of the novel SARS-CoV-2 coronavirus infection COVID-19. Therefore, an urgent need for novel antivirals is crystal clear. Here we present the synthesis and evaluation of an antiviral activity of phenoxazine-based nucleoside analogs divided into three groups: (1) 8-alkoxy-substituted, (2) acyclic, and (3) carbocyclic. The antiviral activity was assessed against a structurally and phylogenetically diverse panel of RNA and DNA viruses from 25 species. Four compounds (11a-c, 12c) inhibited 4 DNA/RNA viruses with EC50 ≤ 20 µM. Toxicity of the compounds for the cell lines used for virus cultivation was negligible in most cases. In addition, previously reported and newly synthesized phenoxazine derivatives were evaluated against SARS-CoV-2, and some of them showed promising inhibition of reproduction with EC50 values in low micromolar range, although accompanied by commensurate cytotoxicity.

Isolation and phylogenetic analysis of SARS-CoV-2 variants collected in Russia during the COVID-19 outbreak.

OBJECTIVES: The outbreak of coronavirus disease 2019 (COVID-19) started in December 2019 in China and then spread worldwide over the following months, involving 188 countries. The objective of this study was to determine the molecular epidemiology of the COVID-19 outbreak in Russia. METHODS: In this study, two severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains were isolated and genetically characterized. A phylogenetic analysis of all available Russian sequences was then performed and these were compared to the epidemiological data on COVID-19 incidence to evaluate the molecular epidemiology and pattern of virus spread in the territory of Russia. RESULTS AND CONCLUSIONS: Whole genome analysis of the isolates obtained in this study and 216 others isolated in Russia revealed a set of seven common mutations when compared to the original Wuhan virus, including amino acid substitutions in spike protein S and nucleoprotein N, possibly affecting their properties. Phylogenetic analysis of all Russian sequences and 8717 sequences from other countries showed multiple importations of the virus into Russia, local circulation, and several patterns of virus spread.

A Chemographic Audit of anti-Coronavirus Structure-activity Information from Public Databases (ChEMBL).

Discovery of drugs against newly emerged pathogenic agents like the SARS-CoV-2 coronavirus (CoV) must be based on previous research against related species. Scientists need to get acquainted with and develop a global oversight over so-far tested molecules. Chemography (herein used Generative Topographic Mapping, in particular) places structures on a human-readable 2D map (obtained by dimensionality reduction of the chemical space of molecular descriptors) and is thus well suited for such an audit. The goal is to map medicinal chemistry efforts so far targeted against CoVs. This includes comparing libraries tested against various virus species/genera, predicting their polypharmacological profiles and highlighting often encountered chemotypes. Maps are challenged to provide predictive activity landscapes against viral proteins. Definition of "anti-CoV" map zones led to selection of therein residing 380 potential anti-CoV agents, out of a vast pool of 800 M organic compounds.