Immunogenicity of Low-Dose Prime-Boost Vaccination of mRNA Vaccine CV07050101 in Non-Human Primates

Many different vaccine candidates against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of COVID-19, are currently approved and under development. Vaccine platforms vary from mRNA vaccines to viral-vectored vaccines, and several candidates have been shown to produce humoral and cellular responses in small animal models, non-human primates, and human volunteers. In this study, six non-human primates received a prime-boost intramuscular vaccination with 4 µg of mRNA vaccine candidate CV07050101, which encodes a pre-fusion stabilized spike (S) protein of SARS-CoV-2. Boost vaccination was performed 28 days post prime vaccination. As a control, six animals were similarly injected with PBS. Humoral and cellular immune responses were investigated at time of vaccination, and two weeks afterwards. No antibodies could be detected at two and four weeks after prime vaccination. Two weeks after boost vaccination, binding but no neutralizing antibodies were detected in four out of six non-human primates. SARS-CoV-2 S protein-specific T cell responses were detected in these four animals. In conclusion, prime-boost vaccination with 4 µg of vaccine candidate CV07050101 resulted in limited immune responses in four out of six non-human primates.     

A highly efficacious live attenuated mumps virus–based SARS-CoV-2 vaccine candidate expressing a six-proline stabilized prefusion spike

With the rapid increase in SARS-CoV-2 cases in children, a safe and effective vaccine for this population is urgently needed. The MMR (measles/mumps/rubella) vaccine has been one of the safest and most effective human vaccines used in infants and children since the 1960s. Here, we developed live attenuated recombinant mumps virus (rMuV)–based SARS-CoV-2 vaccine candidates using the MuV Jeryl Lynn (JL2) vaccine strain backbone. The soluble prefusion SARS-CoV-2 spike protein (preS) gene, stablized by two prolines (preS-2P) or six prolines (preS-6P), was inserted into the MuV genome at the P–M or F–SH gene junctions in the MuV genome. preS-6P was more efficiently expressed than preS-2P, and preS-6P expression from the P–M gene junction was more efficient than from the F–SH gene junction. In mice, the rMuV-preS-6P vaccine was more immunogenic than the rMuV-preS-2P vaccine, eliciting stronger neutralizing antibodies and mucosal immunity. Sera raised in response to the rMuV-preS-6P vaccine neutralized SARS-CoV-2 variants of concern, including the Delta variant equivalently. Intranasal and/or subcutaneous immunization of IFNAR1^−/− mice and golden Syrian hamsters with the rMuV-preS-6P vaccine induced high levels of neutralizing antibodies, mucosal immunoglobulin A antibody, and T cell immune responses, and were completely protected from challenge by both SARS-CoV-2 USA-WA1/2020 and Delta variants. Therefore, rMuV-preS-6P is a highly promising COVID-19 vaccine candidate, warranting further development as a tetravalent MMR vaccine, which may include protection against SARS-CoV-2.

The current pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused tremendous damage to all aspects of our society (1–3). As of 1 June 2022, nearly 528 million cases have been reported worldwide, with nearly 6.3 million deaths (∼1.20% mortality). Symptoms of SARS-CoV-2 infection are primarily respiratory although increasing numbers of other syndromes such as cognitive deficits are being reported. As of June 2022, several SARS-CoV-2 vaccines based on messenger RNA (mRNA), inactivated virus, and adenovirus vectors (Ad26.COV2.S and ChAdOx1) have been approved for vaccination in humans over the age of 5 (4). These vaccines are highly efficacious, reaching 70 to 95% effectiveness against SARS-CoV-2 infection (4).Despite the high success of the current SARS-CoV-2 vaccines, there are several limitations. Protection provided by current vaccines begins to decline after 3 mo (5), which has required a third or fourth dose to boost the immune response. Current vaccines are less effective against recently emergent SARS-CoV-2 variants of concern (VoCs) (6–9). More and more evidence has shown that vaccine-induced neutralizing antibodies were significantly weakened or insufficient to neutralize VoCs such as the Delta variant (7–9), which spreads much faster and causes more severe illness than the earlier strains. In addition, the current vaccines neutralize the most recently emerged variant, Omicron, ∼40 times less efficiently compared with early SARS-CoV-2 isolates (10, 11). The mRNA vaccines are expensive to produce, hard to transport internationally, and difficult to store in many countries because of the requirement for expensive −80 °C freezers.A safe and efficacious pediatric SARS-CoV-2 vaccine is needed to halt the current pandemic. Pfizer’s mRNA vaccine is 90.7% effective in preventing COVID-19 symptoms in children 5 to 11 y old (12, 13). On 17 June 2022, Food and Drug Administration (FDA) authorized emergency use of the Moderna and Pfizer mRNA vaccines for children down to 6 mo of age. As of 23 June 2022, a total of 13.7 million COVID-19 cases have occurred in children, representing 18.8% of the total COVID-19 cases in the United States. Notably, COVID-19 cases in children have increased significantly after the reopening of schools. Therefore, development of other vaccine platforms and strategies to enhance durability, reduce cost, and enhance stability are essential for terminating the pandemic.Historically, the MMR (measles/mumps/rubella) vaccine has been one of the safest and most effective human vaccines ever developed (14–16). The application in children started in the 1960s and provides long-lasting protection against these three viruses (14, 16). Among the three MMR components, measles virus (MeV) and mumps virus (MuV) are nonsegmented negative-sense (NNS) RNA viruses, belonging to the family Paramyxoviridae in the order Mononegavirales. The MuV genome is 15,384 nt in length, and it encodes seven structural proteins arranged in the order 3′-leader-N-P-M-F-SH-HN-L-trailer-5′ (17). The limited number of discrete genes of the NNS RNA genome and the intergenic regions available for inserting additional genes facilitates the development of live vectored vaccines. MuV is an excellent viral vector for delivery of vaccines against other highly pathogenic viruses, primarily because of its high safety and efficacy, well-established good manufacturing practices, induction of long-lived immunity, and the potential for the development of a quadrivalent vaccine against four major pediatric diseases (18, 19).In this study, we developed a suite of safe and highly efficacious recombinant MuV (rMuV)–based SARS-CoV-2 vaccine candidates expressing a stabilized prefusion spike with two prolines (preS-2P) or six prolines (preS-6P) at different gene junctions in the MuV genome. Among them, the rMuV-based preS-6P vaccine induces a broad neutralizing antibody against VoCs and T cell immunity, and provides complete protection against SARS-CoV-2 WA1 and the Delta variant challenge in animal models.     

Antibody Response to SARS-CoV-2 Infection and Vaccination in COVID-19-naïve and Experienced Individuals

Understanding the magnitude of responses to vaccination during the ongoing SARS-CoV-2 pandemic is essential for ultimate mitigation of the disease. Here, we describe a cohort of 102 subjects (70 COVID-19-naïve, 32 COVID-19-experienced) who received two doses of one of the mRNA vaccines (BNT162b2 (Pfizer–BioNTech) and mRNA-1273 (Moderna)). We document that a single exposure to antigen via infection or vaccination induces a variable antibody response which is affected by age, gender, race, and co-morbidities. In response to a second antigen dose, both COVID-19-naïve and experienced subjects exhibited elevated levels of anti-spike and SARS-CoV-2 neutralizing activity; however, COVID-19-experienced individuals achieved higher antibody levels and neutralization activity as a group. The COVID-19-experienced subjects exhibited no significant increase in antibody or neutralization titer in response to the second vaccine dose (i.e., third antigen exposure). Finally, we found that COVID-19-naïve individuals who received the Moderna vaccine exhibited a more robust boost response to the second vaccine dose (p = 0.004) as compared to the response to Pfizer–BioNTech. Ongoing studies with this cohort will continue to contribute to our understanding of the range and durability of responses to SARS-CoV-2 mRNA vaccines.     

Characterization of the First SARS-CoV-2 Isolates from Aotearoa New Zealand as Part of a Rapid Response to the COVID-19 Pandemic

SARS-CoV-2, the virus responsible for the COVID-19 pandemic, has wreaked havoc across the globe for the last two years. More than 300 million cases and over 5 million deaths later, we continue battling the first real pandemic of the 21st century. SARS-CoV-2 spread quickly, reaching most countries within the first half of 2020, and New Zealand was not an exception. Here, we describe the first isolation and characterization of SARS-CoV-2 variants during the initial virus outbreak in New Zealand. Patient-derived nasopharyngeal samples were used to inoculate Vero cells and, three to four days later, a cytopathic effect was observed in seven viral cultures. Viral growth kinetics was characterized using Vero and VeroE6/TMPRSS2 cells. The identity of the viruses was verified by RT-qPCR, Western blot, indirect immunofluorescence assays, and electron microscopy. Whole-genome sequences were analyzed using two different yet complementary deep sequencing platforms (MiSeq/Illumina and Ion PGM™/Ion Torrent™), classifying the viruses as SARS-CoV-2 B.55, B.31, B.1, or B.1.369 based on the Pango Lineage nomenclature. All seven SARS-CoV-2 isolates were susceptible to remdesivir (EC₅₀ values from 0.83 to 2.42 µM) and β-D-N⁴-hydroxycytidine (molnupiravir, EC₅₀ values from 0.96 to 1.15 µM) but not to favipiravir (>10 µM). Interestingly, four SARS-CoV-2 isolates, carrying the D614G substitution originally associated with increased transmissibility, were more susceptible (2.4-fold) to a commercial monoclonal antibody targeting the spike glycoprotein than the wild-type viruses. Altogether, this seminal work allowed for early access to SARS-CoV-2 isolates in New Zealand, paving the way for numerous clinical and scientific research projects in the country, including the development and validation of diagnostic assays, antiviral strategies, and a national COVID-19 vaccine development program.     

Highly Thermotolerant SARS-CoV-2 Vaccine Elicits Neutralising Antibodies against Delta and Omicron in Mice

As existing vaccines fail to completely prevent COVID-19 infections or community transmission, there is an unmet need for vaccines that can better combat SARS-CoV-2 variants of concern (VOC). We previously developed highly thermo-tolerant monomeric and trimeric receptor-binding domain derivatives that can withstand 100 °C for 90 min and 37 °C for four weeks and help eliminate cold-chain requirements. We show that mice immunised with these vaccine formulations elicit high titres of antibodies that neutralise SARS-CoV-2 variants VIC31 (with Spike: D614G mutation), Delta and Omicron (BA.1.1) VOC. Compared to VIC31, there was an average 14.4-fold reduction in neutralisation against BA.1.1 for the three monomeric antigen-adjuvant combinations and a 16.5-fold reduction for the three trimeric antigen-adjuvant combinations; the corresponding values against Delta were 2.5 and 3.0. Our findings suggest that monomeric formulations are suitable for upcoming Phase I human clinical trials and that there is potential for increasing the efficacy with vaccine matching to improve the responses against emerging variants. These findings are consistent with in silico modelling and AlphaFold predictions, which show that, while oligomeric presentation can be generally beneficial, it can make important epitopes inaccessible and also carries the risk of eliciting unwanted antibodies against the oligomerisation domain.     

Previous Infection with SARS-CoV-2 Correlates with Increased Protective Humoral Responses after a Single Dose of an Inactivated COVID-19 Vaccine

Previous studies have indicated that antibody responses can be robustly induced after the vaccination in individuals previously infected by SARS-CoV-2. To evaluate anti-SARS-CoV-2 humoral responses in vaccinated individuals with or without a previous history of COVID-19, we compared levels of anti-SARS-CoV-2 antibodies in the sera from 21 vaccinees, including COVID-19-recovered or -naïve individuals in different times, before and after immunization with an inactivated COVID-19 vaccine. Anti-SARS-CoV-2-specific antibodies elicited after COVID-19 and/or immunization with an inactivated vaccine were measured by ELISA and Plaque Reduction Neutralizing assays. Antibody kinetics were consistently different between the two vaccine doses for naïve individuals, contrasting with the SARS-CoV-2-recovered subjects in which we observed no additional increase in antibody levels following the second dose. Sera from SARS-CoV2-naïve individuals had no detectable neutralizing activity against lineage B.1 SARS-CoV-2 or Gamma variant five months after the second vaccine dose. Contrarily, SARS-CoV-2-recovered subjects retained considerable neutralizing activity against both viruses. We conclude that a single inactivated SARS-CoV-2 vaccine dose may be sufficient to induce protective antibody responses in individuals with previous history of SARS-CoV-2 infection.     

Antiviral Activity of Umifenovir In Vitro against a Broad Spectrum of Coronaviruses,Including the Novel SARS-CoV-2 Virus

An escalating pandemic of the novel SARS-CoV-2 virus is impacting global health, and effective antivirals are needed. Umifenovir (Arbidol) is an indole-derivative molecule, licensed in Russia and China for prophylaxis and treatment of influenza and other respiratory viral infections. It has been shown that umifenovir has broad spectrum activity against different viruses. We evaluated the sensitivity of different coronaviruses, including the novel SARS-CoV-2 virus, to umifenovir using in vitro assays. Using a plaque assay, we revealed an antiviral effect of umifenovir against seasonal HCoV-229E and HCoV-OC43 coronaviruses in Vero E6 cells, with estimated 50% effective concentrations (EC50) of 10.0 ± 0.5 µM and 9.0 ± 0.4 µM, respectively. Umifenovir at 90 µM significantly suppressed plaque formation in CMK-AH-1 cells infected with SARS-CoV. Umifenovir also inhibited the replication of SARS-CoV-2 virus, with EC50 values ranging from 15.37 ± 3.6 to 28.0 ± 1.0 µM. In addition, 21–36 µM of umifenovir significantly suppressed SARS-CoV-2 virus titers (≥2 log TCID50/mL) in the first 24 h after infection. Repurposing of antiviral drugs is very helpful in fighting COVID-19. A safe, pan-antiviral drug such as umifenovir could be extremely beneficial in combating the early stages of a viral pandemic.     

Crystal Structure of SARS-CoV-2 Main Protease in Complex with the Non-Covalent Inhibitor ML188

Viral proteases are critical enzymes for the maturation of many human pathogenic viruses and thus are key targets for direct acting antivirals (DAAs). The current viral pandemic caused by SARS-CoV-2 is in dire need of DAAs. The Main protease (M^pro) is the focus of extensive structure-based drug design efforts which are mostly covalent inhibitors targeting the catalytic cysteine. ML188 is a non-covalent inhibitor designed to target SARS-CoV-1 M^pro, and provides an initial scaffold for the creation of effective pan-coronavirus inhibitors. In the current study, we found that ML188 inhibits SARS-CoV-2 M^pro at 2.5 µM, which is more potent than against SAR-CoV-1 M^pro. We determined the crystal structure of ML188 in complex with SARS-CoV-2 M^pro to 2.39 Å resolution. Sharing 96% sequence identity, structural comparison of the two complexes only shows subtle differences. Non-covalent protease inhibitors complement the design of covalent inhibitors against SARS-CoV-2 main protease and are critical initial steps in the design of DAAs to treat CoVID 19.     

Development and Scalable Production of Newcastle Disease Virus-Vectored Vaccines for Human and Veterinary Use

The COVID-19 pandemic has highlighted the need for efficient vaccine platforms that can rapidly be developed and manufactured on a large scale to immunize the population against emerging viruses. Viral-vectored vaccines are prominent vaccine platforms that have been approved for use against the Ebola virus and SARS-CoV-2. The Newcastle Disease Virus is a promising viral vector, as an avian paramyxovirus that infects poultry but is safe for use in humans and other animals. NDV has been extensively studied not only as an oncolytic virus but also a vector for human and veterinary vaccines, with currently ongoing clinical trials for use against SARS-CoV-2. However, there is a gap in NDV research when it comes to process development and scalable manufacturing, which are critical for future approved vaccines. In this review, we summarize the advantages of NDV as a viral vector, describe the steps and limitations to generating recombinant NDV constructs, review the advances in human and veterinary vaccine candidates in pre-clinical and clinical tests, and elaborate on production in embryonated chicken eggs and cell culture. Mainly, we discuss the existing data on NDV propagation from a process development perspective and provide prospects for the next steps necessary to potentially achieve large-scale NDV-vectored vaccine manufacturing.     

Can Individuals with Suboptimal Antibody Responses to Conventional Antiviral Vaccines Acquire Adequate Antibodies from SARS-CoV-2 mRNA Vaccination?

In Japan, healthcare workers (HCWs) are vaccinated against measles, rubella, chickenpox, mumps, and hepatitis B to prevent nosocomial infection; however, some do not produce sufficient antibodies (“suboptimal responders”). This study compared immune responses to a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 mRNA) vaccine among HCWs with normal and suboptimal responses to conventional vaccines. In this prospective cohort study, 50 HCWs received two doses of BNT162b2 mRNA vaccine 3 weeks apart. SARS-CoV-2 anti-spike antibodies were measured 11 times, starting before the first vaccination and ending 5 months after the second vaccination. Antibody titers of four suboptimal and 46 normal responders were compared. SARS-CoV-2 neutralizing antibody activity was measured twice in suboptimal responders, 1 week/1 month and 5 months after the second vaccination. The SARS-CoV-2 anti-spike antibody was detectable in the samples from suboptimal and normal responders at each timepoint after vaccination. Suboptimal responders exhibited SARS-CoV-2 neutralizing antibody activity 1 week/1 month as well as 5 months after the second vaccination; however, activity was slightly reduced at 5 months. Our findings show that suboptimal responders do acquire adequate SARS-CoV-2 anti-spike and SARS-CoV-2 neutralizing antibodies from vaccination to prevent SARS-CoV-2. SARS-CoV-2 mRNA vaccines should thus be recommended for both normal and suboptimal responders to conventional vaccines.     

Immunogenicity and efficacy of the COVID-19 candidate vector vaccine MVA-SARS-2-S in preclinical vaccination

Severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV-2) has emerged as the infectious agent causing the pandemic coronavirus disease 2019 (COVID-19) with dramatic consequences for global human health and economics. Previously, we reached clinical evaluation with our vector vaccine based on modified vaccinia virus Ankara (MVA) against the Middle East respiratory syndrome coronavirus (MERS-CoV), which causes an infection in humans similar to SARS and COVID-19. Here, we describe the construction and preclinical characterization of a recombinant MVA expressing full-length SARS-CoV-2 spike (S) protein (MVA-SARS-2-S). Genetic stability and growth characteristics of MVA-SARS-2-S, plus its robust expression of S protein as antigen, make it a suitable candidate vaccine for industrial-scale production. Vaccinated mice produced S-specific CD8⁺ T cells and serum antibodies binding to S protein that neutralized SARS-CoV-2. Prime-boost vaccination with MVA-SARS-2-S protected mice sensitized with a human ACE2-expressing adenovirus from SARS-CoV-2 infection. MVA-SARS-2-S is currently being investigated in a phase I clinical trial as aspirant for developing a safe and efficacious vaccine against COVID-19.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of coronavirus disease 2019 (COVID-19), first emerged in late 2019 in China (1). SARS-CoV-2 exhibits extremely efficient human-to-human transmission, the new pathogen rapidly spread worldwide, and within months it caused a global pandemic, changing daily life for billions of people. The COVID-19 case fatality rate of ∼2–5% makes the development of countermeasures a global priority. In fact, the development of COVID-19 vaccine candidates is advancing at an international level with unprecedented speed. About 1 y after the first known cases of COVID-19, we can account for >80 SARS-CoV-2–specific vaccines in clinical evaluations and >10 candidate vaccines already in phase III trials (2–4). However, we still lack information on the key immune mechanisms needed for protection against COVID-19. A better understanding of the types of immune response elicited upon natural SARS-CoV-2 infections has become an essential component to assess the promise of various vaccination strategies (5).The SARS-CoV-2 spike (S) protein serves as the most important target antigen for vaccine development based on preclinical research on candidate vaccines against SARS-CoV or Middle East respiratory syndrome coronavirus (MERS-CoV). The trimeric S protein is a prominent structure at the virion surface and essential for SARS-CoV-2 cell entry. As a class I viral fusion protein, it mediates virus interaction with the cellular receptor angiotensin-converting enzyme 2 (ACE2), and fusion with the host cell membrane, both key steps in infection. Thus, infection can be prevented by S-specific antibodies neutralizing the virus (6–9).Among the front-runner vaccines are new technologies such as messenger RNA (mRNA)-based vaccines and nonreplicating adenovirus vector vaccines (10–13). First reports from these SARS-CoV-2-S–specific vaccines in phase 1/2 clinical studies demonstrated acceptable safety and promising immunogenicity profiles, and by now data from large phase 3 clinical trials show promising levels of protective efficacy (4, 12–14). In December 2020, the first mRNA-based COVID-19 vaccines received emergency use authorization or conditional licensing by the US Food and Drug Administration and European Medicines Agency (11, 15, 16). By March 2021, two adenovirus vector-based COVID-10 vaccines had been approved by regulatory authorities (17, 18). This is good news because efficacious vaccines will provide a strategy to change SARS-CoV-2 transmission dynamics. In addition, multiple vaccine types will be advantageous to meet specific demands across different target populations. This includes the possibility of using heterologous immunization strategies depending on an individual’s health status, boosting capacities, and the need for balanced humoral and Th1-directed cellular immune responses.MVA, a highly attenuated strain of vaccinia virus originating from growth selection on chicken embryo tissue cultures, shows a characteristic replication defect in mammalian cells but allows unimpaired production of heterologous proteins (19). At present, MVA serves as an advanced vaccine technology platform for developing new vector vaccines against infectious disease including emerging viruses and cancer (20). In response to the ongoing pandemic, the MVA vector vaccine platform allows rapid generation of experimental SARS-CoV-2–specific vaccines (21). Previous work from our laboratory addressed the development of an MVA candidate vaccine against MERS with immunizations in animal models demonstrating the safety, immunogenicity, and protective efficacy of MVA-induced MERS-CoV S-antigen–specific immunity (22–25). Clinical safety and immunogenicity of the MVA-MERS-S candidate vaccine was established in a first-in-human phase I clinical study under funding from the German Center for Infection Research (DZIF) (26).Here, we show that a recombinant MVA produces the full-length S protein of SARS-CoV-2 as ∼190- to 200-kDa N-glycosylated protein. Our studies confirmed cleavage of the mature full-length S protein into an amino-terminal domain (S1) and a ∼80- to 100-kDa carboxyl-terminal domain (S2) that is anchored to the membrane. When tested as a vaccine in BALB/c mice, recombinant MVA expressing the S protein induced SARS-CoV-2–specific T cells and antibodies, and robustly protected vaccinated animals against lung infection upon SARS-CoV-2 challenge.     

Antibody Response Induced by BNT162b2 and mRNA-1273 Vaccines against the SARS-CoV-2 in a Cohort of Healthcare Workers

The aim of this study was to characterize the antibody response induced by SARS-CoV-2 mRNA vaccines in a cohort of healthcare workers. A total of 2247 serum samples were analyzed using the Elecsys^® Anti-SARS-CoV-2 S-test (Roche Diagnostics International Ltd., Rotkreuz, Switzerland). Sex, age, body mass index (BMI), arterial hypertension, smoking and time between infection and/or vaccination and serology were considered the confounding factors. Regarding the medians, subjects previously infected with SARS-CoV-2 who preserved their response to the nucleocapsid (N) protein showed higher humoral immunogenicity (BNT162b2: 6456.0 U/mL median; mRNA-1273: 2505.0 U/mL) compared with non-infected (BNT162b2: 867.0 U/mL; mRNA-1273: 2300.5 U/mL) and infected subjects with a lost response to N protein (BNT162b2: 2992.0 U/mL). After controlling for the confounders, a higher response was still observed for mRNA-1273 compared with BNT162b2 in uninfected individuals (FC = 2.35, p < 0.0001) but not in previously infected subjects (1.11 FC, p = 0.1862). The lowest levels of antibodies were detected in previously infected non-vaccinated individuals (39.4 U/mL). Clinical variables previously linked to poor prognoses regarding SARS-CoV-2 infection, such as age, BMI and arterial hypertension, were positively associated with increasing levels of anti-S protein antibody exclusively in infected subjects. The mRNA-1273 vaccine generated a higher antibody response to the S protein than BNT162b2 in non-infected subjects only.     

Antibody Response to the BNT162b2 mRNA COVID-19 Vaccine in Subjects with Prior SARS-CoV-2 Infection

Although antibody levels progressively decrease following SARS-CoV-2 infection, the immune memory persists for months. Thus, individuals who naturally contracted SARS-CoV-2 are expected to develop a more rapid and sustained response to COVID-19 vaccines than naïve individuals. In this study, we analyzed the dynamics of the antibody response to the BNT162b2 mRNA COVID-19 vaccine in six healthcare workers who contracted SARS-CoV-2 in March 2020, in comparison to nine control subjects without a previous infection. The vaccine was well tolerated by both groups, with no significant difference in the frequency of vaccine-associated side effects, with the exception of local pain, which was more common in previously infected subjects. Overall, the titers of neutralizing antibodies were markedly higher in response to the vaccine than after natural infection. In all subjects with pre-existing immunity, a rapid increase in anti-spike receptor-binding domain (RBD) IgG antibodies and neutralizing antibody titers was observed one week after the first dose, which seemed to act as a booster. Notably, in previously infected individuals, neutralizing antibody titers 7 days after the first vaccine dose were not significantly different from those observed in naïve subjects 7 days after the second vaccine dose. These results suggest that, in previously infected people, a single dose of the vaccine might be sufficient to induce an effective response.     

Glycosite-deleted mRNA of SARS-CoV-2 spike protein as a broad-spectrum vaccine

Development of the messenger RNA (mRNA) vaccine has emerged as an effective and speedy strategy to control the spread of new pathogens. After vaccination, the mRNA is translated into the real protein vaccine, and there is no need to manufacture the protein in vitro. However, the fate of mRNA and its posttranslational modification inside the cell may affect immune response. Here, we showed that the mRNA vaccine of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein with deletion of glycosites in the receptor-binding domain (RBD) or especially the subunit 2 (S2) domain to expose more conserved epitopes elicited stronger antibody and CD8⁺ T cell responses with broader protection against the alpha, beta, gamma, delta, and omicron variants, compared to the unmodified mRNA. Immunization of such mRNA resulted in accumulation of misfolded spike protein in the endoplasmic reticulum, causing the up-regulation of BiP/GRP78, XBP1, and p-eIF2α to induce cell apoptosis and strong CD8⁺ T cell response. In addition, dendritic cells (DCs) incubated with S2-glysosite deleted mRNA vaccine increased class I major histocompatibility complex (MHC I) expression. This study provides a direction for the development of broad-spectrum mRNA vaccines which may not be achieved with the use of expressed proteins as antigens.

In 1796, Edward Jenner created the first vaccine (cowpox) in the world to protect against smallpox and successfully rescued millions of people, and, since then, vaccination has been recognized as the best way to protect against pathogens (1). Since the outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in December 2019 that caused COVID-19, the virus has spread all over the world and caused more than 200 million infections and 4 million deaths in 20 mo (2–4). Of the various vaccines developed to control the spread of SARS-CoV-2 and variants, the messenger RNA (mRNA) vaccines developed by Moderna and BioNTech/Pfizer have proven highly effective and represent a major breakthrough in vaccine development, mainly due to the speed and convenience. These vaccines were stabilized with new mRNA technology and lipid nanoparticle (LNP) formulation for delivery and translation into the spike (S) protein in vivo to induce immune response (5–7). The nucleoside modification, sequence optimization, synthesis, and high-performance liquid chromatography purification were the major steps used for development of mRNA vaccines. However, after mRNA vaccination, the translational process and posttranslational modification of the expressed immunogen, and how they affect the outcome of immune response, are not well understood (8). Among the many posttranslational modification events, glycosylation is known to play an important role in protein folding, structure, and function (9). Our previous study showed that deletion of certain glycosites in the S protein affected the expression and folding of S protein, making it difficult to prepare pure S proteins with deletion of specific glycosylation sites for the study of their receptor binding and immune response (10). Therefore, in this study, the mRNA of SARS-CoV-2 S protein with deletion of specific glycosites is used to investigate the effect of glycosylation on protein expression and immune response in vivo.The SARS-CoV-2 S protein has three major immunogenic domains: the N-terminal domain, the receptor-binding domain (RBD), and the subunit 2 domain (S2) (11). Previous studies have shown that the neutralizing antibodies that recognize the RBD are highly protective against SARS-CoV-2 and other coronaviruses (12–14). However, the efficacy of antibodies against the emerging variants was found to decrease as the virus continued to mutate. More than 3 million S protein sequences have been reported to the global database Global Initiative on Sharing Avian Influenza Database (GISAID), and all are highly glycosylated, with 22 N-glycosites and 2 O-glycosites per monomer. As part of our efforts to develop broadly protective vaccines and antibodies against SARS-CoV-2 and the emerging variants, we have shown that immunization of wild-type (WT) S protein with all 22 N-glycosites trimmed down to N-acetylglucosamine (GlcNAc) as the mono-GlcNAc decorated S protein (S_mg) elicited broadly protective immune responses, including antibody and CD4⁺ as well as CD8⁺ T cell responses against the alpha, beta, gamma, and delta variants. Further study showed that most of the conserved epitopes on S protein are located in the RBD and the HR2 domain of the S2 subunit, but these conserved epitopes are largely shielded by glycans to escape from the immune response (10). So, removal of the shielded glycans to expose more conserved epitopes induced broader and stronger immune responses. We also used the single B cell technology to screen B cell clones from S_mg immunized mice, and we identified a broadly neutralizing monoclonal antibody targeting a highly conserved epitope in RBD which was not found in S_fg immunized mice, further demonstrating that removal of glycan shields from S protein is an effective strategy for development of a broadly protective vaccine against SARS-CoV-2 variants (10). To translate this finding into the mRNA vaccine design for development of a broad-spectrum mRNA vaccine, here we show the study of S protein mRNA vaccine with deletion of specific glycosites in RBD and S2 with N to Q and S/T to A replacement and investigation of their protein expression and immune response as well as breadth of protection.     

Reduction in Severity of All-Cause Gastroenteritis Requiring Hospitalisation in Children Vaccinated against Rotavirus in Malawi

Rotavirus is the major cause of severe gastroenteritis in children aged <5 years. Introduction of the G1P[8] Rotarix^® rotavirus vaccine in Malawi in 2012 has reduced rotavirus-associated hospitalisations and diarrhoeal mortality. However, the impact of rotavirus vaccine on the severity of gastroenteritis presented in children requiring hospitalisation remains unknown. We conducted a hospital-based surveillance study to assess the impact of Rotarix^® vaccination on the severity of gastroenteritis presented by Malawian children. Stool samples were collected from children aged <5 years who required hospitalisation with acute gastroenteritis from December 2011 to October 2019. Gastroenteritis severity was determined using Ruuska and Vesikari scores. Rotavirus was detected using enzyme immunoassay. Rotavirus genotypes were determined using nested RT-PCR. Associations between Rotarix^® vaccination and gastroenteritis severity were investigated using adjusted linear regression. In total, 3159 children were enrolled. After adjusting for mid-upper arm circumference (MUAC), age, gender and receipt of other vaccines, all-cause gastroenteritis severity scores were 2.21 units lower (p < 0.001) among Rotarix^®-vaccinated (n = 2224) compared to Rotarix^®-unvaccinated children (n = 935). The reduction in severity score was observed against every rotavirus genotype, although the magnitude was smaller among those infected with G12P[6] compared to the remaining genotypes (p = 0.011). Each one-year increment in age was associated with a decrease of 0.43 severity score (p < 0.001). Our findings provide additional evidence on the impact of Rotarix^® in Malawi, lending further support to Malawi’s Rotarix^® programme.     

Lung Transplant Recipients Immunogenicity after Heterologous ChAdOx1 nCoV-19—BNT162b2 mRNA Vaccination

(1) Background: High immunosuppressive regimen in lung transplant recipients (LTRs) hampers the immune response to vaccination. We prospectively investigated the immunogenicity of heterologous ChAdOx1 nCoV-19-BNT162b2 mRNA vaccination in an LTR cohort. (2) Methods: Forty-nine COVID-19 naïve LTRs received a two-dose regimen ChAdOx1 nCoV-19 vaccine. A subset of 32 patients received a booster dose of BNT162b2 mRNA vaccine 18 weeks after the second dose. (3) Results: Two-doses of ChAdOx1 nCoV-19 induced poor immunogenicity with 7.2% seropositivity at day 180 and low neutralizing capacities. The BNT162b2 mRNA vaccine induced significant increases in IgG titers with means of 197.8 binding antibody units per milliliter (BAU/mL) (95% CI 0–491.4) and neutralizing antibodies, with means of 76.6 AU/mL (95% CI 0–159.6). At day 238, 32.2% of LTRs seroconverted after the booster dose. Seroneutralization capacities against Delta and Omicron variants were found in only 13 and 9 LTRs, respectively. Mycophenolate mofetil and high-dose corticosteroids were associated with a weak serological response. (4) Conclusions: The immunogenicity of a two-dose ChAdOx1 nCoV-19 vaccine regimen was very poor in LTRs, but was significantly enhanced after the booster dose in one-third of LTRs. In immunocompromised individuals, the administration of a fourth dose may be considered to increase the immune response against SARS-CoV-2.     

Elicitation of Broadly Neutralizing Antibodies against B.1.1.7, B.1.351, and B.1.617.1 SARS-CoV-2 Variants by Three Prototype Strain-Derived Recombinant Protein Vaccines

The ongoing coronavirus disease 2019 (COVID-19) pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Most of the currently approved SARS-CoV-2 vaccines use the prototype strain-derived spike (S) protein or its receptor-binding domain (RBD) as the vaccine antigen. The emergence of several novel SARS-CoV-2 variants has raised concerns about potential immune escape. In this study, we performed an immunogenicity comparison of prototype strain-derived RBD, S1, and S ectodomain trimer (S-trimer) antigens and evaluated their induction of neutralizing antibodies against three circulating SARS-CoV-2 variants, including B.1.1.7, B.1.351, and B.1.617.1. We found that, at the same antigen dose, the RBD and S-trimer vaccines were more potent than the S1 vaccine in eliciting long-lasting, high-titer broadly neutralizing antibodies in mice. The RBD immune sera remained highly effective against the B.1.1.7, B.1.351, and B.1.617.1 variants despite the corresponding neutralizing titers decreasing by 1.2-, 2.8-, and 3.5-fold relative to that against the wild-type strain. Significantly, the S-trimer immune sera exhibited comparable neutralization potency (less than twofold variation in neutralizing GMTs) towards the prototype strain and all three variants tested. These findings provide valuable information for further development of recombinant protein-based SARS-CoV-2 vaccines and support the continued use of currently approved SARS-CoV-2 vaccines in the regions/countries where variant viruses circulate.     

Efficacy and effectiveness of COVID-19 vaccines against SARS-CoV-2 infection: interim results of a living systematic review, 1 January to 14 May 2021

Evidence on COVID-19 vaccine efficacy/effectiveness (VE) in preventing asymptomatic SARS-CoV-2 infections is needed to guide public health recommendations for vaccinated people. We report interim results of a living systematic review. We identified a total of 30 studies that investigated VE against symptomatic and/or asymptomatic infection. In fully vaccinated individuals, VE against symptomatic and asymptomatic infections was 80–90% in nearly all studies. Fully vaccinated persons are less likely to become infected and contribute to transmission.Key words: SARS-CoV-2, systematic review, vaccine effectiveness, vaccination, COVID-19

Vaccination against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection plays a key role in the containment of the coronavirus disease (COVID-19) pandemic. All vaccines approved by the European Medical Agency (EMA) at the time of writing demonstrated high vaccine efficacy/effectiveness (VE) against severe COVID-19. With vaccination programmes being implemented in most European countries, it becomes urgent to assess the extent to which these vaccines are also able to prevent symptomatic and asymptomatic infections to guide public health recommendations and develop strategies for fully vaccinated people. In December 2020, the Robert Koch Institute (RKI), in collaboration with the National Immunisation Technical Advisory Groups (NITAGs) network coordinated by the European Centre for Disease Prevention and Control (ECDC) initiated a living systematic review on the VE and safety of European Union (EU)-licensed COVID-19 vaccines (PROSPERO registration: CRD42020208935). In this paper, only efficacy and effectiveness data but not those on safety will be covered. In detail, we report the interim results of the review focusing on two research questions:

• What is the efficacy/effectiveness of COVID-19 vaccines in preventing SARS-CoV-2 infections (irrespective of whether those infected were symptomatic or asymptomatic)?
• What is the efficacy/effectiveness of COVID-19 vaccines in preventing asymptomatic SARS-CoV-2 infections?

     

Anti-SU Antibody Responses in Client-Owned Cats Following Vaccination against Feline Leukaemia Virus with Two Inactivated Whole-Virus Vaccines (Fel-O-Vax® Lv-K and Fel-O-Vax® 5)

A field study undertaken in Australia compared the antibody responses induced in client-owned cats that had been vaccinated using two inactivated whole feline leukaemia virus (FeLV) vaccines, the monovalent vaccine Fel-O-Vax^® Lv-K and the polyvalent vaccine Fel-O-Vax^® 5. Serum samples from 428 FeLV-uninfected cats (118 FeLV-vaccinated and 310 FeLV-unvaccinated) were tested for anti-FeLV neutralising antibodies (NAb) using a live virus neutralisation assay to identify 378 FeLV-unexposed (NAb-negative) and 50 FeLV-exposed (NAb-positive; abortive infections) cats, following by anti-surface unit (SU) FeLV-A and FeLV-B antibody ELISA testing. An additional 42 FeLV-infected cats (28 presumptively regressively infected, 14 presumptively progressively infected) were also tested for anti-SU antibodies. NAb-positive cats displayed significantly higher anti-SU antibody ELISA responses compared to NAb-negative cats (p < 0.001). FeLV-unexposed cats (NAb-negative) that had been vaccinated less than 18 months after a previous FeLV vaccination using the monovalent vaccine (Fel-O-Vax^® Lv-K) displayed higher anti-SU antibody ELISA responses than a comparable group vaccinated with the polyvalent vaccine (Fel-O-Vax^® 5) (p < 0.001 for both anti-FeLV-A and FeLV-B SU antibody responses). This difference in anti-SU antibody responses between cats vaccinated with the monovalent or polyvalent vaccine, however, was not observed in cats that had been naturally exposed to FeLV (NAb-positive) (p = 0.33). It was postulated that vaccination with Fel-O-Vax^® 5 primed the humoral response prior to FeLV exposure, such that antibody production increased when the animal was challenged, while vaccination with Fel-O-Vax^® Lv-K induced an immediate preparatory antibody response that did not quantitatively increase after FeLV exposure. These results raise questions about the comparable vaccine efficacy of the different FeLV vaccine formulations and correlates of protection.     

Antiviral Effect of Budesonide against SARS-CoV-2

Treatment options for COVID-19, a disease caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection, are currently severely limited. Therefore, antiviral drugs that efficiently reduce SARS-CoV-2 replication or alleviate COVID-19 symptoms are urgently needed. Inhaled glucocorticoids are currently being discussed in the context of treatment for COVID-19, partly based on a previous study that reported reduced recovery times in cases of mild COVID-19 after inhalative administration of the glucocorticoid budesonide. Given various reports that describe the potential antiviral activity of glucocorticoids against respiratory viruses, we aimed to analyze a potential antiviral activity of budesonide against SARS-CoV-2 and circulating variants of concern (VOC) B.1.1.7 (alpha) and B.1.351 (beta). We demonstrate a dose-dependent inhibition of SARS-CoV-2 that was comparable between all viral variants tested while cell viability remains unaffected. Our results are encouraging as they could indicate a multimodal mode of action of budesonide against SARS-CoV-2 and COVID-19, which could contribute to an improved clinical performance.