Evaluation of Potency and Duration of Immunity Elicited by a Multivalent FMD Vaccine for Use in South Africa

South Africa (SA) experiences sporadic foot and mouth disease (FMD) outbreaks irrespective of routine prophylactic vaccinations of cattle using imported commercial vaccines. The problem could be mitigated by preparation of vaccines from local virus strains related to those circulating in the endemically infected buffalo populations in the Kruger National Park (KNP). This study demonstrates the individual number of protective doses (PD) of five vaccine candidate strains after homologous virus challenge, as well as the vaccines safety and onset of humoral immunity in naïve cattle. Furthermore, the duration of post-vaccination immunity over a 12-month period is shown, when a multivalent vaccine prepared from the five strains is administered as a primary dose with or without booster vaccinations. The five monovalent vaccines were shown to contain a 50% PD between 4 and 32, elicit humoral immunity with antibody titers ≥2.0 log10 from day 7 post-vaccination, and cause no adverse reactions. Meanwhile, the multivalent vaccine elicited antibody titers ≥2.0 log10 and clinical protection up to 12 months when one or two booster vaccinations were administered within 6 months of the primary vaccination. An insignificant difference between the application of one or two booster vaccinations was revealed. Owing to the number of PDs, we anticipate that the multivalent vaccine could be used successfully for prophylactic and emergency vaccinations without adjustment of the antigen payloads. Furthermore, a prophylactic vaccination regimen comprising primary vaccination of naïve cattle followed by two booster vaccinations 1.5 and 6 months later could potentially maintain herd immunity over a period of 12 months.

Matrix M Adjuvanted H5N1 Vaccine Elicits Broadly Neutralizing Antibodies and Neuraminidase Inhibiting Antibodies in Humans That Correlate With In Vivo Protection

The highly pathogenic avian influenza H5N1 viruses constantly evolve and give rise to novel variants that have caused widespread zoonotic outbreaks and sporadic human infections. Therefore, vaccines capable of eliciting broadly protective antibody responses are desired and under development. We here investigated the magnitude, kinetics and protective efficacy of the multi-faceted humoral immunity induced by vaccination in healthy adult volunteers with a Matrix M adjuvanted virosomal H5N1 vaccine. Vaccinees were given escalating doses of adjuvanted vaccine (1.5μg, 7.5μg, or 30μg), or a non-adjuvanted vaccine (30μg). An evaluation of sera from vaccinees against pseudotyped viruses covering all (sub)clades isolated from human H5N1 infections demonstrated that the adjuvanted vaccines (7.5μg and 30μg) could elicit rapid and robust increases of broadly cross-neutralizing antibodies against all clades. In addition, the adjuvanted vaccines also induced multifaceted antibody responses including hemagglutinin stalk domain specific, neuraminidase inhibiting, and antibody-dependent cellular cytotoxicity inducing antibodies. The lower adjuvanted dose (1.5µg) showed delayed kinetics, whilst the non-adjuvanted vaccine induced overall lower levels of antibody responses. Importantly, we demonstrate that human sera post vaccination with the adjuvanted (30μg) vaccine provided full protection against a lethal homologous virus challenge in mice. Of note, when combining our data from mice and humans we identified the neutralizing and neuraminidase inhibiting antibody titers as correlates of in vivo protection.

Development of molecular clamp stabilized hemagglutinin vaccines for Influenza A viruses

Influenza viruses cause a significant number of infections and deaths annually. In addition to seasonal infections, the risk of an influenza virus pandemic emerging is extremely high owing to the large reservoir of diverse influenza viruses found in animals and the co-circulation of many influenza subtypes which can reassort into novel strains. Development of a universal influenza vaccine has proven extremely challenging. In the absence of such a vaccine, rapid response technologies provide the best potential to counter a novel influenza outbreak. Here, we demonstrate that a modular trimerization domain known as the molecular clamp allows the efficient production and purification of conformationally stabilised prefusion hemagglutinin (HA) from a diverse range of influenza A subtypes. These clamp-stabilised HA proteins provided robust protection from homologous virus challenge in mouse and ferret models and some cross protection against heterologous virus challenge. This work provides a proof-of-concept for clamp-stabilised HA vaccines as a tool for rapid response vaccine development against future influenza A virus pandemics.

Development of the Inactivated QazCovid-in Vaccine: Protective Efficacy of the Vaccine in Syrian Hamsters

In March 2020, the first cases of the human coronavirus disease COVID-19 were registered in Kazakhstan. We isolated the SARS-CoV-2 virus from clinical materials from some of these patients. Subsequently, a whole virion inactivated candidate vaccine, QazCovid-in, was developed based on this virus. To develop the vaccine, a virus grown in Vero cell culture was used, which was inactivated with formaldehyde, purified, concentrated, sterilized by filtration, and then adsorbed on aluminum hydroxide gel particles. The formula virus and adjuvant in buffer saline solution were used as the vaccine. The safety and protective effectiveness of the developed vaccine were studied in Syrian hamsters. The results of the studies showed the absolute safety of the candidate vaccine in the Syrian hamsters. When studying the protective effectiveness, the developed vaccine with an immunizing dose of 5 μg/dose specific antigen protected animals from a wild homologous virus at a dose of 104.5 TCID50/mL. The candidate vaccine induced the formation of virus-neutralizing antibodies in vaccinated hamsters at titers of 3.3 ± 1.45 log2 to 7.25 ± 0.78 log2, and these antibodies were retained for 6 months (observation period) for the indicated titers. No viral replication was detected in vaccinated hamsters, protected against the development of acute pneumonia, and ensured 100% survival of the animals. Further, no replicative virus was isolated from the lungs of vaccinated animals. However, a virulent virus was isolated from the lungs of unvaccinated animals at relatively high titers, reaching 4.5 ± 0.7 log TCID50/mL. After challenge infection, 100% of unvaccinated hamsters showed clinical symptoms (stress state, passivity, tousled coat, decreased body temperature, and body weight, and the development of acute pneumonia), with 25 ± 5% dying. These findings pave the way for testing the candidate vaccine in clinical human trials.

Development of a Novel Live Attenuated Influenza A Virus Vaccine Encoding the IgA-Inducing Protein

Live attenuated influenza virus (LAIV) vaccines elicit a combination of systemic and mucosal immunity by mimicking a natural infection. To further enhance protective mucosal responses, we incorporated the gene encoding the IgA-inducing protein (IGIP) into the LAIV genomes of the cold-adapted A/Leningrad/134/17/57 (H2N2) strain (caLen) and the experimental attenuated backbone A/turkey/Ohio/313053/04 (H3N2) (OH/04att). Incorporation of IGIP into the caLen background led to a virus that grew poorly in prototypical substrates. In contrast, IGIP in the OH/04att background (IGIP-H1att) virus grew to titers comparable to the isogenic backbone H1att (H1N1) without IGIP. IGIP-H1att- and H1caLen-vaccinated mice were protected against lethal challenge with a homologous virus. The IGIP-H1att vaccine generated robust serum HAI responses in naïve mice against the homologous virus, equal or better than those obtained with the H1caLen vaccine. Analyses of IgG and IgA responses using a protein microarray revealed qualitative differences in humoral and mucosal responses between vaccine groups. Overall, serum and bronchoalveolar lavage samples from the IGIP-H1att group showed trends towards increased stimulation of IgG and IgA responses compared to H1caLen samples. In summary, the introduction of genes encoding immunomodulatory functions into a candidate LAIV can serve as natural adjuvants to improve overall vaccine safety and efficacy.

Virus and helper component interactions favour the transmission of recombinant potato virus Y strains

In recent years, several recombinant strains of potato virus Y, notably PVYNTN and PVYN:O have displaced the ordinary strain, PVYO, and emerged as the predominant strains affecting the USA potato crop. Previously we reported that recombinant strains were transmitted more efficiently than PVYO when they were acquired sequentially, regardless of acquisition order. In another recent study, we showed that PVYNTN binds preferentially to the aphid stylet over PVYO when aphids feed on a mixture of PVYO and PVYNTN. To understand the mechanism of this transmission bias as well as preferential virus binding, we separated virus and active helper component proteins (HC), mixed them in homologous and heterologous combinations, and then fed them to aphids using Parafilm sachets. Mixtures of PVYO HC with either PVYN:O or PVYNTN resulted in efficient transmission. PVYN:O HC also facilitated the transmission of PVYO and PVYNTN, albeit with reduced efficiency. PVYNTN HC failed to facilitate transmission of either PVYO or PVYN:O. When PVYO HC or PVYN:O HC was mixed with equal amounts of the two viruses, both viruses in all combinations were transmitted at high efficiencies. In contrast, no transmission occurred when combinations of viruses were mixed with PVYNTN HC. Further study evaluated transmission using serial dilutions of purified virus mixed with HCs. While PVYNTN HC only facilitated the transmission of the homologous virus, the HCs of PVYO and PVYN:O facilitated the transmission of all strains tested. This phenomenon has likely contributed to the increase in the recombinant strains affecting the USA potato crop.

Development of a novel Live Attenuated Influenza A Virus vaccine encoding the IgA-inducing protein

Live attenuated influenza virus (LAIV) vaccines elicit a combination of systemic and mucosal immunity by mimicking a natural infection. To further enhance protective mucosal responses, we incorporated the gene encoding the IgA-inducing protein (IGIP) into the LAIV genomes of the cold-adapted A/Leningrad/134/17/57 (H2N2) strain (caLen) and the experimental attenuated backbone A/turkey/Ohio/313053/04 (H3N2) (OH/04att). Incorporation of IGIP into the caLen background led to a virus that grew poorly in prototypical substrates. In contrast, IGIP in the OH/04att background (IGIP-H1att) virus grew to titers comparable to the isogenic backbone H1att (H1N1) without IGIP. IGIP-H1att- and H1caLen-vaccinated mice were protected against lethal challenge with a homologous virus. The IGIP-H1att vaccine generated robust serum HAI responses in naïve mice against the homologous virus, equal or better than those obtained with the H1caLen vaccine. Analyses of IgG and IgA responses using a protein microarray revealed qualitative differences in humoral and mucosal responses between vaccine groups. Overall, serum and bronchoalveolar lavage samples from the IGIP-H1att group showed trends towards increased stimulation of IgG and IgA responses compared to H1caLen samples. In summary, introduction of genes encoding immunomodulatory functions into a candidate LAIV that can serve as natural adjuvants to improve overall vaccine safety and efficacy.

Immunization by exposure to live virus (SIVmne/HIV-2287) during antiretroviral drug prophylaxis may reduce risk of subsequent viral challenge

Rationale/Study design A major challenge in the development of HIV vaccines is finding immunogens that elicit protection against a broad range of viral strains. Immunity to a narrow range of viral strains may protect infants of HIV-infected women or partners discordant for HIV. We hypothesized that immunization to the relevant viral variants could be achieved by exposure to infectious virus during prophylaxis with antiretroviral drugs. To explore this approach in an animal model, macaques were exposed to live virus (SIVmne or HIV-2287) during prophylaxis with parenteral tenofovir and humoral and cellular immune responses were quantified. Subsequently, experimental animals were challenged with homologous virus to evaluate protection from infection, and if infection occurred, the course of disease was compared to control animals. Experimental animals uninfected with SIVmne were challenged with heterologous HIV-2287 to assess resistance to retroviral infection. Methodology/Principal findings Juvenile female Macaca nemestrina (N = 8) were given ten weekly intravaginal exposures with either moderately (SIVmne) or highly (HIV-2287) pathogenic virus during tenofovir prophylaxis. Tenofovir protected all 8 experimental animals from infection, while all untreated control animals became infected. Specific non-neutralizing antibodies were elicited in blood and vaginal secretions of experimental animals, but no ELISPOT responses were detected. Six weeks following the cessation of tenofovir, intravaginal challenge with homologous virus infected 2/4 (50%) of the SIVmne-immunized animals and 4/4 (100%) of the HIV-2287-immunized animals. The two SIVmne-infected and 3 (75%) HIV-2287-infected had attenuated disease, suggesting partial protection. Conclusions/Significance Repeated exposure to SIVmne or HIV-2287, during antiretroviral prophylaxis that blocked infection, induced binding antibodies in the blood and mucosa, but not neutralizing antibodies or specific cellular immune responses. Studies to determine whether antibodies are similarly induced in breastfeeding infants and sexual partners discordant for HIV infection and receiving pre-exposure antiretroviral prophylaxis are warranted, including whether these antibodies appear to confer partial or complete protection from infection.

Insights into SARS-CoV-2: Medicinal Chemistry Approaches to Combat Its Structural and Functional Biology

Coronavirus disease 2019, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is still a pandemic around the world. Currently, specific antiviral drugs to control the epidemic remain deficient. Understanding the details of SARS-CoV-2 structural biology is extremely important for development of antiviral agents that will enable regulation of its life cycle. This review focuses on the structural biology and medicinal chemistry of various key proteins (Spike, ACE2, TMPRSS2, RdRp and Mpro) in the life cycle of SARS-CoV-2, as well as their inhibitors/drug candidates. Representative broad-spectrum antiviral drugs, especially those against the homologous virus SARS-CoV, are summarized with the expectation they will drive the development of effective, broad-spectrum inhibitors against coronaviruses. We are hopeful that this review will be a useful aid for discovery of novel, potent anti-SARS-CoV-2 drugs with excellent therapeutic results in the near future.

Persistently high antibody responses after AS03-adjuvanted H1N1pdm09 vaccine: Dissecting the HA specific antibody response

Current influenza vaccines have a suboptimal effectiveness. The introduction of a novel A/H1N1 influenza virus in 2009 (H1N1pdm09) provided a unique opportunity to study the humoral response to the AS03-adjuvanted H1N1pdm09 vaccine and repeated annual vaccination with the homologous virus in subsequent influenza seasons. Thirty-two HCWs immunized with the AS03-adjuvanted H1N1pdm09 vaccine in 2009 were divided into four groups based on the longevity of their antibody responses (persistently high or transient), and whether they were repeatedly annually vaccinated in the subsequent four influenza seasons or not. Serological assays were utilized to measure the quantity, quality and functionality of antibodies targeting the major surface glycoprotein hemagglutinin (HA). Persistent high responders (hemagglutination inhibition (HI) titre ≥ 80 at 12 months after H1N1pdm09 vaccination) had protective levels of HI antibodies throughout the study period. In addition, the quality and functionality of these antibodies were greater than the individuals who had a transient antibody response to the pandemic vaccine (HI titre < 40 at 12 months after H1N1pdm09 vaccination). All groups had similar levels of antibodies towards the conserved HA stalk domain. The level of HA head-specific antibodies gradually increased over time with annual vaccination in the transient responders. The AS03-adjuvanted H1N1pdm09 vaccine elicited a robust humoral response that persisted up to 5 years in some individuals. Seasonal annual vaccination boosted the HA-antibodies over time in individuals with a transient response to the pandemic H1N1pdm09 vaccine.