Background Influenza viruses cause substantial annual morbidity and mortality globally. Current vaccines protect against influenza only when well matched to the circulating strains. However, antigenic drift can cause considerable mismatches between vaccine and circulating strains, substantially reducing vaccine effectiveness. Moreover, current seasonal vaccines are ineffective against pandemic influenza, and production of a vaccine matched to a newly emerging virus strain takes months. Therefore, there is an unmet medical need for a broadly protective influenza virus vaccine. We aimed to test the ability of chimeric H1 haemagglutinin-based universal influenza virus vaccine candidates to induce broadly cross-reactive antibodies targeting the stalk domain of group 1 haemagglutininexpressing influenza viruses. Methods We did a randomised, observer-blinded, phase 1 study in healthy adults in two centres in the USA. Participants were randomly assigned to one of three prime-boost, chimeric haemagglutinin-based vaccine regimens or one of two placebo groups. The vaccine regimens included a chimeric H8/1, intranasal, live-attenuated vaccine on day 1 followed by a non-adjuvanted, chimeric H5/1, intramuscular, inactivated vaccine on day 85; the same regimen but with the inactivated vaccine being adjuvanted with AS03; and an AS03-adjuvanted, chimeric H8/1, intramuscular, inactivated vaccine followed by an AS03-adjuvanted, chimeric H5/1, intramuscular, inactivated vaccine. In this planned interim analysis, the primary endpoints of reactogenicity and safety were assessed by blinded study group. We also assessed anti-H1 haemagglutinin stalk, anti-H2, anti-H9, and anti-H18 IgG antibody titres and plasmablast and memory B-cell responses in peripheral blood. This trial is registered with ClinicalTrials.gov, number NCT03300050.

Summary Background Use of oral live-attenuated polio vaccines (OPV), and injected inactivated polio vaccines (IPV) has almost achieved global eradication of wild polio viruses. To address the goals of achieving and maintaining global eradication and minimising the risk of outbreaks of vaccine-derived polioviruses, we tested novel monovalent oral type-2 poliovirus (OPV2) vaccine candidates that are genetically more stable than existing OPVs, with a lower risk of reversion to neurovirulence. Our study represents the first in-human testing of these two novel OPV2 candidates. We aimed to evaluate the safety and immunogenicity of these vaccines, the presence and extent of faecal shedding, and the neurovirulence of shed virus. Methods In this double-blind, single-centre phase 1 trial, we isolated participants in a purpose-built containment facility at the University of Antwerp Hospital (Antwerp, Belgium), to minimise the risk of environmental release of the novel OPV2 candidates. Participants, who were recruited by local advertising, were adults (aged 18–50 years) in good health who had previously been vaccinated with IPV, and who would not have any contact with immunosuppressed or unvaccinated people for the duration of faecal shedding at the end of the study. The first participant randomly chose an envelope containing the name of a vaccine candidate, and this determined their allocation; the next 14 participants to be enrolled in the study were sequentially allocated to this group and received the same vaccine. The subsequent 15 participants enrolled after this group were allocated to receive the other vaccine. Participants and the study staff were masked to vaccine groups until the end of the study period. Participants each received a single dose of one vaccine candidate (candidate 1, S2/cre5/S15domV/rec1/hifi3; or candidate 2, S2/S15domV/CpG40), and they were monitored for adverse events, immune responses, and faecal shedding of the vaccine virus for 28 days. Shed virus isolates were tested for the genetic stability of attenuation. The primary outcomes were the incidence and type of serious and severe adverse events, the proportion of participants showing viral shedding in their stools, the time to cessation of viral shedding, the cell culture infective dose of shed virus in virus-positive stools, and a combined index of the prevalence, duration, and quantity of viral shedding in all participants. This study is registered with EudraCT, number 2017-000908-21 and ClinicalTrials.gov , number NCT03430349 . Findings Between May 22 and Aug 22, 2017, 48 volunteers were screened, of whom 15 (31%) volunteers were excluded for reasons relating to the inclusion or exclusion criteria, three (6%) volunteers were not treated because of restrictions to the number of participants in each group, and 30 (63%) volunteers were sequentially allocated to groups (15 participants per group). Both no...

Summary The live-attenuated oral poliovirus vaccine (OPV or Sabin vaccine) replicates in gut-associated tissues, eliciting mucosa and systemic immunity. OPV protects from disease and limits poliovirus spread. Accordingly, vaccination with OPV is the primary strategy used to end the circulation of all polioviruses. However, the ability of OPV to regain replication fitness and establish new epidemics represents a significant risk of polio re-emergence should immunization cease. Here, we report the development of a poliovirus type 2 vaccine strain (nOPV2) that is genetically more stable and less likely to regain virulence than the original Sabin2 strain. We introduced modifications within at the 5′ untranslated region of the Sabin2 genome to stabilize attenuation determinants, 2C coding region to prevent recombination, and 3D polymerase to limit viral adaptability. Prior work established that nOPV2 is immunogenic in preclinical and clinical studies, and thus may enable complete poliovirus eradication.

Expression of the poliovirus receptor (PVR) on cells is a major host determinant of infection by poliovirus.Previously, the only immune cell type known to express PVR was the blood-derived monocyte, which is susceptible to infection at very low frequency. We demonstrate that professional antigen-presenting cellsmacrophages and dendritic cells, generated upon differentiation of monocytes-retain expression of PVR and are highly susceptible to infection by type 1 Mahoney strain of poliovirus. Maximal cell-associated titers of virus are obtained within 6 to 8 h postinfection, and cell death and lysis occurs within 24 h postinfection. Similar kinetics are observed in cells infected with the Sabin 1 vaccine strain. Although protein synthesis and receptor-mediated endocytosis are inhibited upon poliovirus infection of these critical antigen-presenting cells, we demonstrate for the first time that functional presentation of antigen occurs in these infected cells via the HLA class II pathway.Poliovirus (PV), a prototypical member of the Picornaviridae family, is a small, nonenveloped, positive-stranded RNA virus whose replication is limited to specific cells and tissues that express the PV receptor (PVR; CD155) (30, 40) on the cell surface. Most cells of the immune system, such as T and B lymphocytes, are not susceptible to PV infection (35, 41), as they do not express PVR. In contrast, Freistadt et al. demonstrated that human peripheral blood monocytes do express PVR (22). However, monocytes are not very permissive to PV infection; viral protein production could only be demonstrated at very low frequency in a subpopulation of monocytes (18,21).Monocytes are precursor cells that are able to differentiate into macrophages or dendritic cells (DCs) (10-12, 36, 44). In response to pathogens, inflammatory cytokines, or necrotic cells, DCs and macrophages play central roles in the induction of immune responses. These antigen-presenting cells (APCs) acquire and process antigens, displaying them in the context of HLA class I and II molecules at the cell surface (9,31,43,45). Subsequent interaction of the HLA-antigen complexes and costimulatory molecules on APCs with T cells in the presence of relevant secreted cytokines induces immune responses (26). DCs are critical APCs that, in contrast to macrophages, are unique in being able to induce primary immune responses from naive T cells to novel antigens in humans (6).To better understand the interaction of PV with the immune system, we characterized the susceptibility of monocyte-derived macrophages and DCs to PV infection. We show here that these in vitro-differentiated macrophages and DCs retain PVR expression and can be productively infected with PV. The kinetics of viral infection in these APCs follows that seen for PV infection with well-characterized laboratory cell lines (7, 38) and results in cell death. Furthermore, PV-induced cytopathic effects typically observed during viral infection of standard cell lines (1, 3, 37, 41, 48) are also seen here upon infection with either...

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The presence of poliovirus (PV)-specific CD4؉ T cells in individuals vaccinated against polio has been shown, but CD8 ؉ T-cell responses have not been described. Here, we functionally characterize the CD4 ؉ T-cell response and show for the first time that dendritic cells and macrophages can stimulate PV-specific CD8 ؉ T-cell responses in vitro from vaccinees. Both CD4؉ T and CD8 ؉ T cells secrete gamma interferon in response to PV antigens and are cytotoxic via the perforin/granzyme B-mediated pathway. Furthermore, the T cells also recognize and kill Sabin 1 vaccine-infected targets. The macrophage-stimulated CD4 ؉ T and CD8 ؉ T cells most likely represent memory T cells that persist for long periods in vaccinated individuals. Thus, immunity to PV vaccination involves not only an effective neutralizing antibody titer but also long-term CD4؉ and CD8؉ cytotoxic T-cell responses.Immunity to poliovirus (PV) as a result of a vaccination regimen with oral Sabin vaccine (OPV) and/or inactivated Salk vaccine (IPV) has been the focus of many studies. The majority of these studies have been directed towards the protective neutralizing antibody response in various populations worldwide (6,10,15,28,45,49); less attention has been given to adaptive T-cell responses and the probable role of these cells in the resolution of PV infection.T-cell responses to PV have been studied primarily in mice. These studies have identified several epitopes in PV structural proteins that are recognized by murine CD4 and CD8 T cells (17,21,23,26). As normal mice lack the human poliovirus receptor (PVR) and are not, therefore, susceptible to poliovirus infection, the role of the PV-specific T cells in protective immunity in these mice could not be determined. However, adoptive transfers of PV-specific CD4 T cells into PVR-transgenic mice that are susceptible to PV infection showed that CD4 T cells do indeed provide protection against infection by lethal doses of PV and that these CD4 T cells provide help to B cells to produce protective antibodies (26). In contrast, studies identifying PV-specific T cells present in humans, the normal host for PV, have been limited. It is clear that PV-specific CD4 T cells are induced in vaccinated individuals, and some CD4 T-cell epitopes have been defined (13, 43). There are no previous reports that a CD8 T-cell-mediated response to PV is initiated in vaccinated individuals, even following recent IPV boosters (20), or in primates that are susceptible to PV infection.The induction of cytotoxic CD8 T cells is a cornerstone of viral immunity, as CD8 T cells are important in viral clearance. Dendritic cells (DCs) and macrophages (M) are central to the initiation of CD4 and CD8 T-cell responses to pathogens. We have recently shown that these critical antigen-presenting cells (APCs) express PVR and can be productively infected with PV, with kinetics and cytopathology similar to those seen during infection of HeLa cells, and that infection results in apoptotic cell death (48). Several processes that are important in...

Novel oral poliovirus vaccine type 2 (nOPV2) is being developed to reduce the rare occurrence of disease and outbreaks associated with the genetic instability of the Sabin vaccine strains. Children aged 1 to 5 years were enrolled in two related clinical studies to assess safety, immunogenicity, shedding rates and properties of the shed virus following vaccination with nOPV2 (two candidates) versus traditional Sabin OPV type 2 (mOPV2). The anticipated pattern of reversion and increased virulence was observed for shed Sabin-2 virus, as assessed using a mouse model of poliovirus neurovirulence. In contrast, there were significantly reduced odds of mouse paralysis for shed virus for both nOPV2 candidates when compared to shed Sabin-2 virus. Next-generation sequencing of shed viral genomes was consistent with and further supportive of the observed neurovirulence associated with shed Sabin-2 virus, as well as the reduced reversion to virulence of shed candidate viruses. While shed Sabin-2 showed anticipated A481G reversion in the primary attenuation site in domain V in the 5’ untranslated region to be associated with increased mouse paralysis, the stabilized domain V in the candidate viruses did not show polymorphisms consistent with reversion to neurovirulence. The available data from a key target age group for outbreak response confirm the superior genetic and phenotypic stability of shed nOPV2 strains compared to shed Sabin-2 and suggest that nOPV2 should be associated with less paralytic disease and potentially a lower risk of seeding new outbreaks.

Sabin-strain oral polio vaccines (OPV) can, in rare instances, cause disease in recipients and susceptible contacts or evolve to become circulating vaccine-derived strains with the potential to cause outbreaks. Two novel type 2 OPV (nOPV2) candidates were designed to stabilize the genome against the rapid reversion that is observed following vaccination with Sabin OPV type 2 (mOPV2). Next-generation sequencing and a modified transgenic mouse neurovirulence test were applied to shed nOPV2 viruses from phase 1 and 2 studies and shed mOPV2 from a phase 4 study. The shed mOPV2 rapidly reverted in the primary attenuation site (domain V) and increased in virulence. In contrast, the shed nOPV2 viruses showed no evidence of reversion in domain V and limited or no increase in neurovirulence in mice. Based on these results and prior published data on safety, immunogenicity, and shedding, the nOPV2 viruses are promising alternatives to mOPV2 for outbreak responses.