Domain Of Spike Protein Research Articles

Development and Evaluation of In‐House ELISAs for the Detection of SARS‐CoV‐2‐Specific Antibodies in COVID‐19 Patients in Sri Lanka

COVID‐19 serological tests complement the molecular diagnostics and can be used as important tool for serosurveillance and vaccine efficiency evaluation. The aim of this study was to develop and evaluate the diagnostic performance of an in‐house ELISA for retrospective serosurveillance of SARS‐CoV‐2. Total IgG and IgM levels in sera of PCR positive SARS‐CoV‐2 patients (n = 50) from North Colombo Teaching Hospital were evaluated and compared with sera (n = 50) collected from prepandemic healthy individuals as controls. Patient sample collection was initiated before vaccination programme was widely started within the country. Seropositivity of 94.0% (n = 47/50) was observed for either IgG or IgM anti‐SARS‐CoV‐2 antibodies against receptor binding domain of spike protein or nucleocapsid protein in confirmed cases while none of controls were seropositive. In contrast, the seropositivity of only 48.0% (n = 24/50) was demonstrated with commercial ELISA kits for detection of IgG or IgM. All samples detected seropositive by commercially available kits remained seropositive with either in‐house IgM or IgG ELISA. Significant correlations (p ≤ 0.001) were observed between Ab levels and day of sampling from the onset of illness. The overall sensitivity values of the in‐house assays were 66.7%, 96.9%, and 100.0% for the first, second, and third week or longer after onset of symptoms for either in‐house IgM or IgG ELISAs. Majority of the patients (>80.0%) were seropositive, regardless of age (<60 vs. >60 years), gender (male vs. female), or clinical severity (mild vs. moderate/severe). These data suggest that the developed in‐house ELISAs can be applied to assess anti‐SARS‐CoV‐2 antibody levels induced by either natural infections or vaccination.

Bond strength between receptor binding domain of spike protein and human angiotensin converting enzyme-2 using machine learning

Bond strength between receptor binding domain of spike protein and human angiotensin converting enzyme-2 using machine learning

Boosting with variant-matched adenovirus-based vaccines promotes neutralizing antibody responses against SARS-CoV-2 Omicron sublineages in mice

Global spreading of SARS-CoV-2 Omicron subvariants, such as BA.4, BA.5, and XBB.1.5, has been leading the recent wave of COVID-19. Unique mutations in the spike proteins in these emerging Omicron subvariants caused immune evasion from the pre-existing protective immunity induced by vaccination or natural infection. We previously developed AdCLD-CoV19-1, a non-replicating recombinant adenoviral vector that encode receptor binding domain of spike protein of ancestral SARS-CoV-2 strain. Based on the same recombinant adenoviral vector platform, we constructed updated vaccines that cover unique mutations found in each Omicron subvariants including BA.1, BA.2, BA.4.1, and BA.5. Preclinical studies reveal that each updated vaccine as a booster shot following primary vaccination targeting ancestral strain most effectively improved neutralizing antibody responses against pseudo-virus of its respective strain. Of note, boosting with a vaccine targeting the BA.1 or BA.2 Omicron subvariant was most effective in neutralization against the pseudo-virus of BA.2.75 strain, whereas BA.4.1/5-adapted booster shots were most effective in neutralization against BQ.1, BQ1.1, and BF.7. Therefore, it is imperative to develop a vaccination strategy that can cover the unique spike mutations of currently circulating Omicron subvariants to prevent the next wave of COVID-19.

Defining a de novo non-RBM antibody as RBD-8 and its synergistic rescue of immune-evaded antibodies to neutralize Omicron SARS-CoV-2.

Currently, monoclonal antibodies (MAbs) targeting the SARS-CoV-2 receptor binding domain (RBD) of spike (S) protein are classified into seven classes based on their binding epitopes. However, most of these antibodies are seriously impaired by SARS-CoV-2 Omicron and its subvariants, especially the recent BQ.1.1, XBB and its derivatives. Identification of broadly neutralizing MAbs against currently circulating variants is imperative. In this study, we identified a "breathing" cryptic epitope in the S protein, named as RBD-8. Two human MAbs, BIOLS56 and IMCAS74, were isolated recognizing this epitope with broad neutralization abilities against tested sarbecoviruses, including SARS-CoV, pangolin-origin coronaviruses, and all the SARS-CoV-2 variants tested (Omicron BA.4/BA.5, BQ.1.1, and XBB subvariants). Searching through the literature, some more RBD-8 MAbs were defined. More importantly, BIOLS56 rescues the immune-evaded antibody, RBD-5 MAb IMCAS-L4.65, by making a bispecific MAb, to neutralize BQ.1 and BQ.1.1, thereby producing an MAb to cover all the currently circulating Omicron subvariants. Structural analysis reveals that the neutralization effect of RBD-8 antibodies depends on the extent of epitope exposure, which is affected by the angle of antibody binding and the number of up-RBDs induced by angiotensin-converting enzyme 2 binding. This cryptic epitope which recognizes non- receptor binding motif (non-RBM) provides guidance for the development of universal therapeutic antibodies and vaccines against COVID-19.

Lasioglossin-1 peptide inhibits binding of spike protein of SARS-CoV-2 to ACE2 receptor: an in silico approach of some bee venom peptides

ABSTRACT The coronavirus prevalent, initiated by the SARS-CoV-2 virus, has induced a global health crisis, recently. Coronavirus can bind to ACE2 receptors on the surface of human cells and infect them through the RBD domain of spike protein. Bee venom peptides have anti-inflammatory effects and were proposed for coronavirus cure. To investigate this proposal, 11 peptides from bee venom were selected and 3D structures of them were prepared. Each peptide was simulated for 100 ns molecular dynamics simulation and 22 conformations were extracted from the last 20 ns of MD simulation. Then All 220 conformations were docked to the RBD domain of spike protein by haddock server and the best complexes for each peptide were chosen for another 100 ns MD simulation. Then MM/PBSA binding free energy calculations were done. The results revealed that the Lasioglossin-1 peptide (VNWKKVLGKIIKVAK) had better interaction with the RBD domain of spike protein than other peptides and can reduce virus entrance to human cells. This peptide can be a potential drug for coronavirus. Also, to find a probable better peptide, two mutations were created in this peptide, and previous procedures were repeated and it determined that none of the mutated peptides were better than native Lasioglossin-1.

Neutralizing Antibody Responses in Patients with Hematological Malignancies after Omicron Infection: A Multicenter, Prospective Cohort Study in China

BACKGROUND The mortality rate of hospitalized hematological malignancies (HM) patients with COVID-19 has significantly decreased from 40.7% in the pre-Omicron era (before December 2021) to 16.5% in the Omicron era (Blennow O, et al. AJH 2022; Pagano L, et al. JHO 2021). However, the mortality rate of Omicron infection in the general population is approximately 1%, which is comparable to influenza (Wang B, et al. J Infect 2023). Therefore, COVID-19 continues to pose a substantial threat to HM patients, even under the Omicron era. Neutralizing antibodies (NAbs) play a crucial role in preventing breakthrough infections and reinfections by blocking the entry of SARS-CoV-2 into the body (Nordstrom P, et al. Lancet Infect Dis 2022). However, little is known about neutralizing antibody (NAb) response development after natural Omicron infection and the risk factors for poor responders in HM patients. METHODS A registered multi-center, prospective, observational study was conducted during the latest Omicron wave in Chongqing, China (November 2022 to January 2023). NAb titers (measured by serum IgG anti-receptor binding domain of spike protein [anti-S-RBD]) were assessed 3-6 weeks after infection in the HM cohort (HMC), healthy control cohort (HCC), and solid cancer cohort (SCC). The primary outcomes compared the anti-S-RBD IgG titer and seroconversion rate between the HMC and HCC or SCC. The exploratory outcome investigated risk factors for poor responders, defined as patients with non-seroconversion or low anti-S-RBD IgG titer, in the HMC. RESULTS Among the 466 HM patients in this cohort, the seroconversion rate was 92.7%, no statistically difference compared with HCC (98.2%, P=0.0513) or SCC (100%, P=0.1363); the median anti-S-RBD IgG titer was 29.9 ng/mL, significantly lower than HCC (46.9 ng/mL, P<0.0001) or SCC (46.2 ng/mL, P<0.0001). Risk factors associated with non-seroconversion included no COVID-19 vaccination history (OR=4.58, 95% CI: 1.75-12.00, P=0.002), clinical course of COVID-19 ≤ 7 days (OR=2.86, 95% CI: 1.31-6.25, P=0.008) and severe B-cell reduction (0-10/μL) (OR=3.22, 95% CI: 1.32-7.88, P=0.010); Risk factors associated with low anti-S-RBD IgG titer were clinical course of COVID-19 ≤ 7 days (OR=2.58, 95% CI: 1.59-4.18, P<0.001) and severe B-cell reduction (0-10/μL) (OR=2.87, 95% CI: 1.57-5.24, P<0.001). CONCLUSIONS This study reveals a poor NAb response in HM patients after Omicron (BA.5.2.48) infection and identifies risk factors for poor responders. Highlights that HM patients, especially those with these risk factors, may be susceptible to SARS-CoV-2 reinfection; and the post-infection vaccination strategies for these patients should be tailored.

Mutation Interest in the Receptor-Binding Domain of Spike Protein of Severe Acute Respiratory Syndrome Coronavirus 2 in the Northwest of Iran

Background: The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been undergoing variation. Most of the variants cause no concern for human health. Some others have had worse outcomes in terms of transmissibility, vaccination resistance, and, generally, the survival of patients infected with SARS-CoV-2. Objectives: This study investigated the mutation of interest in the receptor-binding domain (RBD) of SARS-CoV-2. Methods: Ribonucleic acid (RNA) was extracted from 40 swap samples. Next, the polymerase chain reaction (PCR) assay was carried out to detect the RBD. Investigation of SARS-COV-2 RBD was performed completely by phylogenetic and tree alignment. The multiple sequence alignment (MSA) of the biological sequence of RBD was created by BioEdit, Snap Gene, and MEGA software and was then compared to sequences of different variants of SARS-COV-2 in the GenBank (National Center for Biotechnology Information). The Influenza Surveillance and Response System (GSAID) was used to detect mutations in the RBD sequence. Results: Multiple sequence alignment (MSA) RBD domain showed that the RBD domain sequence obtained from the Iranian patients' highest identity by severe acute respiratory syndrome coronavirus 2 isolate MZ907347. Several mutations of interest, including A475V, L452R, V483A, and F490L, were detected in the RBD region. However, the dN/dS analysis detected positive selection in RBD regions. Conclusions: Amino acid changes in the surface protein can significantly alter the viral function and/or interactions with neutralizing antibodies. Most of the nucleotide changes in the spike gene reduce infectivity. The V503W and P521Q mutations reduce infectivity, the A522Q mutation increases sensitivity to neutralizing antibodies, and the H519T mutation decreases susceptibility to convalescent sera.

Non-synonymous Single Nucleotide Polymorphisms in Human ACE2 Gene May Affect the Infectivity of SARS-CoV-2 Omicron Subvariants.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the coronavirus disease 2019 (COVID-19), which first appeared in December 2019. Angiotensin I converting enzyme 2 (ACE2) receptor, present on the host cells, interacts with the receptor binding domain (RBD) of spike (S) protein of SARS-CoV-2 and facilitates the viral entry into host cells. Non-synonymous single nucleotide polymorphisms (nsSNPs) in the ACE2 gene may have an impact on the protein's stability and its function. The deleterious or harmful nsSNPs of the ACE2 gene that can change the strength as well as the pattern of interaction with the RBD of S protein were selected for this study. The ACE2:RBD interactions were analyzed by protein-protein docking study. The missense mutations A242V, R708W, G405E, D292N, Y633C, F308L, and G405E in ACE2 receptor were found to interact with RBD of Omicron subvariants with stronger binding affinity. Among the other selected nsSNPs of human ACE2 (hACE2), R768W, Y654S, F588S, R710C, R710C, A191P, and R710C were found to have lower binding affinity for RBD of Omicron subvariants. The findings of this study suggest that the nsSNPs present in the human ACE2 gene alter the structure and function of the protein and, consequently, the susceptibility to Omicron subvariants.

Comparison of Anti-SARS-CoV-2-Specific Antibody Signatures in Maternal and Infant Blood after COVID-19 Infection versus COVID-19 Vaccination during Pregnancy.

The Advisory Committee on Immunization Practices and The American College of Obstetricians and Gynecologists recommend coronavirus disease 2019 (COVID-19) vaccine for pregnant persons to prevent severe illness and death. The objective was to examine levels of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) IgG, IgM, and IgA against spike protein receptor binding domain (RBD) and nucleocapsid protein (NCP) in maternal and infant/cord blood at delivery after COVID 19 vaccination compared with SARS-CoV-2 infection at in mother-infant dyads at specified time points. Mothers with SARS-CoV-2 infection (n = 31) or COVID-19 vaccination (n = 25) during pregnancy were enrolled between July 2020 and November 2021. Samples were collected at delivery and IgG, IgM, and IgA to RBD of spike and NCPs compared in the infected and vaccinated groups. Timing of infection/vaccination prior to delivery and correlation with antibody levels was performed. The majority of participants received vaccination within 90 days of delivery and over half received the Pfizer BioNTech vaccine. There were no significant correlations between antibody levels and timing of infection or vaccination. Infant IgG levels to the RBD domain of spike protein were higher in the vaccinated group (n = 25) as compared with the infants born to mothers with infection (n = 31). Vaccination against COVID-19 during pregnancy was associated with detectable maternal and infant anti-RBD IgG levels at delivery irrespective of the timing of vaccination. Timing of vaccination had no correlation to the antibody levels suggesting that the timing of maternal vaccination in the cohort did not matter. There was no IgM detected in infants from vaccinated mothers. Infants from vaccinated mothers had robust IgG titers to RBD, which have a lasting protective effect in infants. · COVID-19 vaccination during pregnancy had detectable antibody.. · No correlation between antibody levels and timing of vaccination.. · Infants from vaccinated mothers had robust IgG titers to RBD..

Biophysical evolution of the receptor-binding domains of SARS-CoVs

With hundreds of coronaviruses (CoVs) identified in bats that can infect humans, it is essential to understand how CoVs that affected the human population have evolved. Seven known CoVs have infected humans, of which three CoVs caused severe disease with high mortalities: severe acute respiratory syndrome (SARS)-CoV emerged in 2002, Middle East respiratory syndrome-CoV in 2012, and SARS-CoV-2 in 2019. SARS-CoV and SARS-CoV-2 belong to the same family, follow the same receptor pathway, and use their receptor-binding domain (RBD) of spike protein to bind to the angiotensin-converting enzyme 2 (ACE2) receptor on the human epithelial cell surface. The sequence of the two RBDs is divergent, especially in the receptor-binding motif that directly interacts with ACE2. We probed the biophysical differences between the two RBDs in terms of their structure, stability, aggregation, and function. Since RBD is being explored as an antigen in protein subunit vaccines against CoVs, determining these biophysical properties will also aid in developing stable protein subunit vaccines. Our results show that, despite RBDs having a similar three-dimensional structure, they differ in their thermodynamic stability. RBD of SARS-CoV-2 is significantly less stable than that of SARS-CoV. Correspondingly, SARS-CoV-2 RBD shows a higher aggregation propensity. Regarding binding to ACE2, less stable SARS-CoV-2 RBD binds with a higher affinity than more stable SARS-CoV RBD. In addition, SARS-CoV-2 RBD is more homogenous in terms of its binding stoichiometry toward ACE2 compared to SARS-CoV RBD. These results indicate that SARS-CoV-2 RBD differs from SARS-CoV RBD in terms of its stability, aggregation, and function, possibly originating from the diverse receptor-binding motifs. Higher aggregation propensity and decreased stability of SARS-CoV-2 RBD warrant further optimization of protein subunit vaccines that use RBD as an antigen by inserting stabilizing mutations or formulation screening.

A novel recombinant S-based subunit vaccine induces protective immunity against porcine deltacoronavirus challenge in piglets.

As an emerging porcine enteropathogenic coronavirus that has the potential to infect humans, porcine deltacoronavirus (PDCoV) is receiving increasing attention. However, no effective commercially available vaccines against this virus are available. In this work, we designed a spike (S) protein and receptor-binding domain (RBD) trimer as a candidate PDCoV subunit vaccine. We demonstrated that S protein induced more robust humoral and cellular immune responses than the RBD trimer in mice. Furthermore, the protective efficacy of the S protein was compared with that of inactivated PDCoV vaccines in piglets and sows. Of note, the immunized piglets and suckling pig showed a high level of NAbs and were associated with reduced virus shedding and mild diarrhea, and the high level of NAbs was maintained for at least 4 months. Importantly, we demonstrated that S protein-based subunit vaccines conferred significant protection against PDCoV infection.

Classification of five SARS-CoV-2 serotypes based on RBD antigenicities

The continuous evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in a significant number of variants, particularly with the emergence of Omicron with many sub-variants. These variants have exhibited increased immune escape, leading to reduced efficacy of existing vaccines and therapeutic antibodies. Given the diminished cross-neutralization observed among these variants, it is plausible that SARS-CoV-2 has developed multiple serotypes. As the major antigenic site, the receptor-binding domain (RBD) of viral spike (S) protein was chosen for serotyping. We selected 23 representative variants, including pre-Omicron variants and Omicron sub-variants, and classified them into five serotypes based on systematic evaluation of the antigenicities of their RBDs. Each serotype includes several genetically distinct variants. Serotype-I encompasses all pre-Omicron variants (with two subtypes), while the remaining four serotypes are all comprised of Omicron sub-variants at different stages of evolution. We propose that these serotypes can serve as a foundation for rapid classification of newly emerging SARS-CoV-2 variants, and guide the development of future broad-spectrum vaccines and neutralizing antibodies against the coronavirus disease 2019 (COVID-19).

Synthesis, docking studies and biological evaluation of 1H-1,2,3-triazole-7-chloroquinoline derivatives against SARS-CoV-2

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a single-stranded enveloped positive RNA virus and the cause of the ongoing coronavirus disease 2019 (COVID-19) pandemic. Chloroquine (CQ), an antimalarial drug, was reported to be active against several viruses including coronaviruses. The mechanism of host cell invasion by SARS-CoV-2 involves the interaction of angiotensin-converting enzyme (ACE2) with receptor-binding domain (RBD) of spike protein (S). The main protease (Mpro/3CLpro) is an attractive drug target due to its vital function in regulation of polyprotein translated from viral RNA. In this study, a series of novel quinoline-triazole hybrid compounds was synthesized and subjected to evaluations on their cytotoxicity, interactions with different variants of RBD in SARS-CoV-2 and with 3CLpro enzyme by experimental and computational techniques to identify their ability of counteracting viral infection. The results of bio-layer interferometry showed that quinoline derivative 11 has good interaction with delta plus and omicron RBD variants (KD = 3.46 × 10−5 and 6.38 × 10−5 M) while derivative 1 is the best binder for recent variant omicron (KD = 26.9 µM) among the series. Potent compounds 1–4 and 11 also demonstrated a suppressive effect on 3CLpro activity in a non-dose-dependent manner. Further docking study revealed that these compounds interacted within the same area of RBD, while no correlation was found for 3CLpro. Furthermore, the molecular dynamics simulations were carried out to assess the conformational stability of docked complexes for preliminary verification.

B-089 Development of a Secondary Standard for Quantitation of IgG Antibodies to SARS-CoV-2 Proteins in a Multiplex Assay

Abstract Background Serological surveillance is essential for monitoring disease burden and vaccine uptake at an individual and population level. During the pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), several serological assays were developed and obtained emergency use authorization to detect antibodies after infection. At that time, the need was to differentiate the infected from the naïve population, and these qualitative assays played a critical role in disease management. In the wake of vaccinations, there is a need for the development of quantitative assays. The world health organization (WHO) developed an international standard for quantitation. We performed this study to develop and validate an in-house secondary standard for our multiplex assay based on this WHO standard. Methods We developed a novel multiplex assay for quantifying human IgG antibodies to four different SARS-CoV-2 antigens in human serum or plasma. We have used Luminex® xMAP® technology for this assay and immobilized four recombinant proteins, namely, the receptor-binding domain of S1(RBD), nucleocapsid protein (NP), S1 protein (S1), and trimeric Spike protein (trimer) on internally coded magnetic microspheres. We procured lyophilized WHO quantification standard 20/136 from the National Institute for Biological Standards and Control (NIBSC), which has the arbitrary assignment of 1000 BAU/mL after reconstitution in 0.25 mL. We tested this standard in the range of 10 to 0.01 BAU/mL for the standard curve. We tested NIBSC 20/162 control reagent using this assay for the linearity of dilution. Thirty-seven samples from the NIBSC verification panel were also tested for this evaluation. We also quantified IgG antibodies in 25 samples from vaccinated individuals, ten samples from vaccine breakthrough individuals, 25 samples from PCR-positive individuals, and 75 samples collected before 2019. Results The secondary standard was calibrated using the 20/136 standard. The NIBSC 20/136 and the secondary standard displayed a wide dynamic range of quantitation. The difference in the expected and actual concentrations was less than 20%. Multiple dilutions of the serum or plasma samples were used to get readings within the standard curve range. Excellent linearity was noted at various sample dilutions to get the correct quantification. We observed that vaccinated individuals had a higher concentration of binding antibodies to S1, Trimer, and RBD and were negative for NP antibodies. Conversely, the acute samples from infected individuals collected less than ten days post-positive PCR showed higher antibodies to NP than different spike proteins. The vaccine break-through infection samples showed lower NP antibodies than unvaccinated but infected individuals. Conclusion Our development of a secondary standard calibrated against WHO international standard is vital for quantitatively measuring different SARS-CoV-2 antibodies. A secondary standard can help manufacturers and users to normalize results from various assays and lots over a prolonged period.

Analyses of spike protein from first deposited sequences of SARS-CoV2 from West Bengal, India.

India has recently started sequencing SARS-CoV2 genome from clinical isolates. Currently only few sequences are available from three states in India. Kerala was the first state to deposit complete sequence from two isolates followed by one from Gujarat. On April 27, 2020, the first five sequences from the state of West Bengal (Eastern India) were deposited on GISAID, a global initiative for sharing avian flu data. In this study, we have analysed the spike protein sequences from all five isolates and also compared their similarities or differences with other sequences reported in India and with isolates of Wuhan origin. We report one unique mutation at position 723 and another at 1124 in the S2 domain of spike protein of the isolates from West Bengal only. There was one mutation downstream of the receptor binding domain at position 614 in S1 domain which was common with the sequence from Gujarat (a state of western India). Mutation in the S2 domain showed changes in the secondary structure of the spike protein at region of the mutation. We also studied molecular dynamics using normal mode analyses and found that this mutation decreases the flexibility of S2 domain. Since both S1 and S2 are important in receptor binding followed by entry in the host cells, such mutations may define the affinity or avidity of receptor binding.

Intranasally inoculated bacterium-like particles displaying porcine epidemic diarrhea virus S1 protein induced intestinal mucosal immune response in mice.

Porcine epidemic diarrhea virus (PEDV) causes acute watery diarrhea and high mortality in newborn piglets. Activation of intestinal mucosal immunity is crucial to anti-PEDV infection. To develop a vaccine capable of stimulating intestinal mucosal immunity, we prepared a bacterium (Lactococcus lactis)-like particle (BLP) vaccine (S1-BLPs) displaying the S1 protein, a domain of PEDV spike protein (S), based on gram-positive enhancer matrix (GEM) particle display technology. We further compared the effects of different vaccination routes on mucosal immune responses in mice induced by S1-BLPs. The specific IgG titer in serum of intramuscularly immunized mice with S1-BLPs was significantly higher than that of the intranasally administered. The specific IgA antibody was found in the serum and intestinal lavage fluid of mice vaccinated intranasally, but not intramuscularly. Moreover, the intranasally inoculated S1-BLPs induced higher levels of IFN-γ and IL-4 in serum than the intramuscularly inoculated. In addition, the ratio of serum IgG2a/IgG1 of mice inoculated intramuscularly was significantly higher with S1-BLPs compared to that of with S1 protein, suggesting that the immune responses induced by S1-BLPs was characterized by helper T (Th) cell type 1 immunity. The results indicated that S1-BLPs induced systemic and local immunity, and the immunization routes significantly affected the specific antibody classes and Th immune response types. The intranasally administered S1-BLPs could effectively stimulate intestinal mucosal specific secretory IgA response. S1-BLPs have the potential to be developed as PEDV mucosal vaccine.

Antiviral Drugs and Vaccines for Omicron Variant: A Focused Review.

The Omicron variant of concern (VOC) replaced the delta variant rapidly and became the predominant strain due to more mutations in spike protein and receptor-binding domain (RBD) enhancing its infectivity and binding affinity. The severity of the illness is less than that of the delta variant. Omicron is nonsusceptible to REGEN-COV™ and bamlanivimab with etesevimab. Drugs that are effective against the Omicron variant are oral antiviral drugs such as Paxlovid (nirmatrelvir/ritonavir), remdesivir, sotrovimab, and molnupiravir. The potency of sotrovimab is reduced to 3-fold against Omicron, and 8-fold reduction in potency with sotrovimab is found in a particular variant of Omicron with a R346K substitution in spike protein. There are neither clinical trials comparing the efficacy of these 4 therapies with each other nor any data on a combination of two or more therapies. The current recommendation for mild-moderate, nonhospitalized patients who are at a high risk of disease progression is to use Paxlovid as the first-line option. If Paxlovid is not available or cannot be administered due to drug interactions, then the next best choice is sotrovimab. The third choice is remdesivir if sotrovimab is also not available and molnupiravir is to be given if the other three options are not available or cannot be administered. For prevention, 2130 (cilgavimab) in combination with COV2-2196 (tixagevimab) has been effective against BA.2 only. LY-CoV1404 (bebtelovimab) is recently authorized as it is effective against all sublineages of the Omicron variant. Regarding vaccine efficacy (VE), the 3-dose VE with mRNA vaccines at 14-60 days was found to be 71.6%, and after 60 days, it is 47.4%. There is a 34-38-fold reduction of neutralizing activity with prebooster sera and a 19-fold reduction with booster sera for the Omicron variant. This probably explains the reason for worldwide breakthrough infections with the Omicron variant with waning immunity. The neutralizing antibody response against Omicron elicited by the bivalent vaccine is superior to that of the ancestral Wuhan strain, without any safety concerns. For future advances, the ribosome display technology can be applied for the generation of human single-chain fragment variable (scFv) antibodies from B cells of recovered patients against Omicron and other Coronavirus variants as they are easier and faster to produce and have high affinity and high specificity.

In silico prediction of immune-escaping hot spots for future COVID-19 vaccine design

The COVID-19 pandemic has had a widespread impact on a global scale, and the evolution of considerable dominants has already taken place. Some variants contained certain key mutations located on the receptor binding domain (RBD) of spike protein, such as E484K and N501Y. It is increasingly worrying that these variants could impair the efficacy of current vaccines or therapies. Therefore, analyzing and predicting the high-risk mutations of SARS-CoV-2 spike glycoprotein is crucial to design future vaccines against the different variants. In this work, we proposed an in silico approach, immune-escaping score (IES), to predict high-risk immune-escaping hot spots on the receptor-binding domain (RBD), implemented through integrated delta binding free energy measured by computational mutagenesis of spike-antibody complexes and mutation frequency calculated from viral genome sequencing data. We identified 23 potentially immune-escaping mutations on the RBD by using IES, nine of which occurred in omicron variants (R346K, K417N, N440K, L452Q, L452R, S477N, T478K, F490S, and N501Y), despite our dataset being curated before the omicron first appeared. The highest immune-escaping score (IES = 1) was found for E484K, which agrees with recent studies stating that the mutation significantly reduced the efficacy of neutralization antibodies. Furthermore, our predicted delta binding free energy and IES show a high correlation with high-throughput deep mutational scanning data (Pearson’s r = 0.70) and experimentally measured neutralization titers data (mean Pearson’s r = −0.80). In summary, our work presents a new method to identify the potentially immune-escaping mutations on the RBD and provides valuable insights into future COVID-19 vaccine design.

Comparative Longitudinal Serological Study of Anti-SARS-CoV-2 Antibody Profiles in People with COVID-19.

Serological diagnostic assays are essential tools for determining an individual's protection against viruses like SARS-CoV-2, tracking the spread of the virus in the community, and evaluating population immunity. To assess the diversity and quality of the anti-SARS-CoV-2 antibody response, we have compared the antibody profiles of people with mild, moderate, and severe COVID-19 using a dot blot assay. The test targeted the four major structural proteins of SARS-CoV-2, namely the nucleocapsid (N), spike (S) protein domains S1 and S2, and receptor-binding domain (RBD). Serum samples were collected from 63 participants at various time points for up to 300 days after disease onset. The dot blot assay revealed patient-specific differences in the anti-SARS-CoV-2 antibody profiles. Out of the 63 participants with confirmed SARS-CoV-2 infections and clinical COVID-19, 35/63 participants exhibited diverse and robust responses against the tested antigens, while 14/63 participants displayed either limited responses to a subset of antigens or no detectable antibody response to any of the antigens. Anti-N-specific antibody levels decreased within 300 days after disease onset, whereas anti-S-specific antibodies persisted. The dynamics of the antibody response did not change during the test period, indicating stable antibody profiles. Among the participants, 28/63 patients with restricted anti-S antibody profiles or undetectable anti-S antibody levels in the dot blot assay also exhibited weak neutralization activity, as measured by a surrogate virus neutralization test (sVNT) and a microneutralization test. These results indicate that in some cases, natural infections do not lead to the production of neutralizing antibodies. Furthermore, the study revealed significant serological variability among patients, regardless of the severity of their COVID-19 illness. These differences need to be carefully considered when evaluating the protective antibody status of individuals who have experienced primary SARS-CoV-2 infections.

Computational analysis of spike protein of SARS-CoV-2 (Omicron variant) for development of peptide-based therapeutics and diagnostics

In the last few years, the worldwide population has suffered from the SARS-CoV-2 pandemic. The WHO dashboard indicated that around 504,079,039 people were infected and 6,204,155 died from COVID-19 caused by different variants of SARS-CoV-2. Recently, a new variant of SARS-CoV-2 (B.1.1.529) was reported by South Africa known as Omicron. The high transmissibility rate and resistance towards available anti-SARS-CoV-2 drugs/vaccines/monoclonal antibodies, make Omicron a variant of concern. Because of various mutations in spike protein, available diagnostic and therapeutic treatments are not reliable. Therefore, the present study explored the development of some therapeutic peptides that can inhibit the SARS-CoV-2 virus interaction with host ACE2 receptors and can also be used for diagnostic purposes. The screened linear B cell epitopes derived from receptor-binding domain of spike protein of Omicron variant were evaluated as peptide inhibitor/vaccine candidates through different bioinformatics tools including molecular docking and simulation to analyze the interaction between Omicron peptide and human ACE2 receptor. Overall, in-silico studies revealed that Omicron peptides OP1-P12, OP14, OP20, OP23, OP24, OP25, OP26, OP27, OP28, OP29, and OP30 have the potential to inhibit Omicron interaction with ACE2 receptor. Moreover, Omicron peptides OP20, OP22, OP23, OP24, OP25, OP26, OP27, and OP30 have shown potential antigenic and immunogenic properties that can be used in design and development vaccines against Omicron. Although the in-silico validation was performed by comparative analysis with the control peptide inhibitor, further validation through wet lab experimentation is required before its use as therapeutic peptides. Communicated by Ramaswamy H. Sarma