The Experts below are selected from a list of 59187 Experts worldwide ranked by ideXlab platform
Florian Krammer - One of the best experts on this subject based on the ideXlab platform.
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A mosaic Hemagglutinin-based influenza virus vaccine candidate protects mice from challenge with divergent H3N2 strains
npj Vaccines, 2019Co-Authors: Felix Broecker, Raffael Nachbagauer, Florian Krammer, Weina Sun, Sean T. H. Liu, Nungruthai Suntronwong, Mark J. Bailey, Peter PaleseAbstract:Current seasonal influenza virus vaccines only provide limited, short-lived protection, and antigenic drift in the Hemagglutinin surface glycoprotein necessitates their annual re-formulation and re-administration. To overcome these limitations, universal vaccine strategies that aim at eliciting broadly protective antibodies to conserved epitopes of the Hemagglutinin show promise for protecting against diverse and drifted influenza viruses. Here a vaccination strategy that focuses antibody responses to conserved epitopes of the H3 Hemagglutinin is described. The approach is based on antigenic silencing of the immunodominant major antigenic sites of an H3 protein from 2014 by replacing them with corresponding sequences of exotic avian Hemagglutinins, yielding “mosaic” Hemagglutinins. In mice, vaccination with inactivated viruses expressing mosaic Hemagglutinins induced highly cross-reactive antibodies against the H3 stalk domain that elicited Fc-mediated effector functions in vitro. In addition, the mosaic viruses elicited head-specific antibodies with neutralizing and hemagglutination-inhibiting activity against recent H3N2 viruses in vitro. Immune sera protected mice from heterologous challenge with viruses carrying H3 proteins from 1968 and 1982, whereas immune sera generated with a seasonal vaccine did not protect. Consequently, the mosaic vaccination approach provides a promising avenue toward a universal influenza virus vaccine. Flu vaccines that target the head domain of the viral Hemagglutinin (HA) surface glycoprotein are highly immunogenic, yet the variable nature of this domain across viral strains and its continuous antigenic drift limit vaccine efficacy. As the HA stalk domain is conserved across influenza viruses, targeting this region is a promising strategy to generate universal flu vaccines. However, eliciting high antibody levels against the HA stalk domain and conserved regions in the head domain remains challenging. Here Peter Palese and colleagues at Mount Sinai develop “mosaic” HAs (mHAs) aimed at eliciting antibodies targeting both the stalk domain and epitopes in the head domain outside of the major antigenic sites. These mHAs are based on antigenic silencing of the immunodominant antigenic sites of an H3 protein, which are exchanged with exotic HA sequences from avian influenza viruses. Vaccination of mice with inactivated viruses expressing mHAs induced both stalk- and head-specific antibodies active in vitro, and transfer of sera from mice immunized with mHAs was better than sera from mice immunized with a seasonal vaccine at protecting mice from heterologous challenge with viruses carrying different H3 proteins, demonstrating the potential of mHAs as universal influenza virus vaccine candidates.
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Antibody Responses toward the Major Antigenic Sites of Influenza B Virus Hemagglutinin in Mice, Ferrets, and Humans.
Journal of Virology, 2018Co-Authors: Weina Sun, Florian Krammer, Davina S. Kang, Allen Zheng, Sean T. H. Liu, Felix Broecker, Viviana Simon, Peter PaleseAbstract:ABSTRACT The influenza B virus Hemagglutinin contains four major antigenic sites (the 120 loop, the 150 loop, the 160 loop, and the 190 helix) within the head domain. These immunodominant antigenic sites are the main targets of neutralizing antibodies and are subject to antigenic drift. Yet little is known about the specific antibody responses toward each site in terms of antibody prevalence and hemagglutination inhibition activity. In this study, we used modified Hemagglutinins of influenza B virus which display only one or none of the major antigenic sites to measure antibody responses toward the classical as well as the noncanonical epitopes in mice, ferrets, and humans. With our novel reagents, we found that both hemagglutination inhibition antibodies and total IgGs were mostly induced by the major antigenic sites. However, in human adults, we observed high hemagglutination inhibition antibody responses toward the noncanonical epitopes. By stratifying the human samples into age groups, we found that the noncanonical antibody responses appeared to increase with age. IMPORTANCE This study dissected the specific antibody responses toward the major antigenic sites and the noncanonical epitopes of influenza B virus Hemagglutinin in animals and humans using novel reagents. These findings will guide the design of the next generation of influenza virus vaccines.
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immunodominance of antigenic site b in the Hemagglutinin of the current h3n2 influenza virus in humans and mice
Journal of Virology, 2018Co-Authors: Felix Broecker, Florian Krammer, Viviana Simon, Peter PaleseAbstract:The Hemagglutinin protein of H3N2 influenza viruses is the major target of neutralizing antibodies induced by infection and vaccination. However, the virus frequently escapes antibody-mediated neutralization due to mutations in the globular head domain. Five topologically distinct antigenic sites in the head domain of H3 Hemagglutinin, A to E, have been previously described by mapping the binding sites of monoclonal antibodies, yet little is known about the contribution of each site to the immunogenicity of modern H3 Hemagglutinins, as measured by hemagglutination inhibition activity, which is known to correlate with protection. To investigate the hierarchy of antibody immunodominance, five Δ1 recombinant influenza viruses expressing Hemagglutinin of the A/Hong Kong/4801/2014 (H3N2) strain with mutations in single antigenic sites were generated. Next, the Δ1 viruses were used to determine the hierarchy of immunodominance by measuring the hemagglutination inhibition reactivity of mouse antisera and plasma from 18 human subjects before and after seasonal influenza vaccination in 2017-2018. In both mice and humans, mutations in antigenic site B caused the most significant decrease in hemagglutination inhibition titers compared to wild-type Hemagglutinin. This study revealed that antigenic site B is immunodominant in the H3N2 influenza virus strain included in the current vaccine preparations. IMPORTANCE Influenza viruses rapidly evade humoral immunity through antigenic drift, making current vaccines poorly effective and antibody-mediated protection short-lived. The majority of neutralizing antibodies target five antigenic sites in the head domain of the Hemagglutinin protein that are also the most sequence-variable regions. A better understanding of the contribution of each antigenic site to the overall antibody response to Hemagglutinin may help in the design of improved influenza virus vaccines.
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Universal influenza virus vaccines: what can we learn from the human immune response following exposure to H7 subtype viruses?
Frontiers of Medicine, 2017Co-Authors: Daniel Stadlbauer, Philip Meade, Raffael Nachbagauer, Florian KrammerAbstract:Several universal influenza virus vaccine candidates based on eliciting antibodies against the Hemagglutinin stalk domain are in development. Typically, these vaccines induce responses that target group 1 or group 2 Hemagglutinins with little to no cross-group reactivity and protection. Similarly, the majority of human anti-stalk monoclonal antibodies that have been isolated are directed against group 1 or group 2 Hemagglutinins with very few that bind to Hemagglutinins of both groups. Here we review what is known about the human humoral immune response to vaccination and infection with H7 subtype influenza viruses on a polyclonal and monoclonal level. It seems that unlike vaccination with H5 Hemagglutinin, which induces antibody responses mostly restricted to the group 1 stalk domain, H7 exposure induces both group 2 and cross-group antibody responses. A better understanding of this phenomenon and the underlying mechanisms might help to develop future universal influenza virus vaccine candidates.
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age dependence and isotype specificity of influenza virus Hemagglutinin stalk reactive antibodies in humans
Mbio, 2016Co-Authors: Raffael Nachbagauer, Peter Palese, Angela Choi, Ruvim Izikson, Manon M J Cox, Florian KrammerAbstract:ABSTRACT Influenza remains a major global health burden. Seasonal vaccines offer protection but can be rendered less effective when the virus undergoes extensive antigenic drift. Antibodies that target the highly conserved Hemagglutinin stalk can protect against drifted viruses, and vaccine constructs designed to induce such antibodies form the basis for a universal influenza virus vaccine approach. In this study, we analyzed baseline and postvaccination serum samples of children (6 to 59 months), adults (18 to 49 years), and elderly individuals (≥65 years) who participated in clinical trials with a recombinant Hemagglutinin-based vaccine. We found that baseline IgG and IgA antibodies against the H1 stalk domain correlated with the ages of patients. Children generally had very low baseline titers and did not respond well to the vaccine in terms of making stalk-specific antibodies. Adults showed the highest induction of stalk-specific antibodies, but the elderly had the highest absolute antibody titers against the stalk. Importantly, the stalk antibodies measured by enzyme-linked immunosorbent assay (ELISA) showed neutralizing activity in neutralization assays and protected mice in a passive-transfer model in a stalk titer-dependent manner. Finally, we found similar patterns of stalk-specific antibodies directed against the H3 and influenza B virus Hemagglutinins, albeit at lower levels than those measured against the H1 stalk. The relatively high levels of stalk-specific antibodies in the elderly patients may explain the previously reported low influenza virus infection rates in this age group. (This study has been registered at ClinicalTrials.gov under registration no. NCT00336453, NCT00539981, and NCT00395174.) IMPORTANCE The present study provides evidence that titers of broadly neutralizing Hemagglutinin stalk-reactive antibodies increase with age, possibly due to repeated exposure to divergent influenza viruses. These relatively high levels of antistalk titers may be responsible for lower circulation rates of influenza viruses in older individuals. Our findings suggest that the level of antistalk antibodies is a good surrogate marker for protection against influenza virus infection. In addition, the levels of antistalk antibodies might determine the breadth of protection against different drifted strains.
Peter Palese - One of the best experts on this subject based on the ideXlab platform.
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A mosaic Hemagglutinin-based influenza virus vaccine candidate protects mice from challenge with divergent H3N2 strains
npj Vaccines, 2019Co-Authors: Felix Broecker, Raffael Nachbagauer, Florian Krammer, Weina Sun, Sean T. H. Liu, Nungruthai Suntronwong, Mark J. Bailey, Peter PaleseAbstract:Current seasonal influenza virus vaccines only provide limited, short-lived protection, and antigenic drift in the Hemagglutinin surface glycoprotein necessitates their annual re-formulation and re-administration. To overcome these limitations, universal vaccine strategies that aim at eliciting broadly protective antibodies to conserved epitopes of the Hemagglutinin show promise for protecting against diverse and drifted influenza viruses. Here a vaccination strategy that focuses antibody responses to conserved epitopes of the H3 Hemagglutinin is described. The approach is based on antigenic silencing of the immunodominant major antigenic sites of an H3 protein from 2014 by replacing them with corresponding sequences of exotic avian Hemagglutinins, yielding “mosaic” Hemagglutinins. In mice, vaccination with inactivated viruses expressing mosaic Hemagglutinins induced highly cross-reactive antibodies against the H3 stalk domain that elicited Fc-mediated effector functions in vitro. In addition, the mosaic viruses elicited head-specific antibodies with neutralizing and hemagglutination-inhibiting activity against recent H3N2 viruses in vitro. Immune sera protected mice from heterologous challenge with viruses carrying H3 proteins from 1968 and 1982, whereas immune sera generated with a seasonal vaccine did not protect. Consequently, the mosaic vaccination approach provides a promising avenue toward a universal influenza virus vaccine. Flu vaccines that target the head domain of the viral Hemagglutinin (HA) surface glycoprotein are highly immunogenic, yet the variable nature of this domain across viral strains and its continuous antigenic drift limit vaccine efficacy. As the HA stalk domain is conserved across influenza viruses, targeting this region is a promising strategy to generate universal flu vaccines. However, eliciting high antibody levels against the HA stalk domain and conserved regions in the head domain remains challenging. Here Peter Palese and colleagues at Mount Sinai develop “mosaic” HAs (mHAs) aimed at eliciting antibodies targeting both the stalk domain and epitopes in the head domain outside of the major antigenic sites. These mHAs are based on antigenic silencing of the immunodominant antigenic sites of an H3 protein, which are exchanged with exotic HA sequences from avian influenza viruses. Vaccination of mice with inactivated viruses expressing mHAs induced both stalk- and head-specific antibodies active in vitro, and transfer of sera from mice immunized with mHAs was better than sera from mice immunized with a seasonal vaccine at protecting mice from heterologous challenge with viruses carrying different H3 proteins, demonstrating the potential of mHAs as universal influenza virus vaccine candidates.
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Antibody Responses toward the Major Antigenic Sites of Influenza B Virus Hemagglutinin in Mice, Ferrets, and Humans.
Journal of Virology, 2018Co-Authors: Weina Sun, Florian Krammer, Davina S. Kang, Allen Zheng, Sean T. H. Liu, Felix Broecker, Viviana Simon, Peter PaleseAbstract:ABSTRACT The influenza B virus Hemagglutinin contains four major antigenic sites (the 120 loop, the 150 loop, the 160 loop, and the 190 helix) within the head domain. These immunodominant antigenic sites are the main targets of neutralizing antibodies and are subject to antigenic drift. Yet little is known about the specific antibody responses toward each site in terms of antibody prevalence and hemagglutination inhibition activity. In this study, we used modified Hemagglutinins of influenza B virus which display only one or none of the major antigenic sites to measure antibody responses toward the classical as well as the noncanonical epitopes in mice, ferrets, and humans. With our novel reagents, we found that both hemagglutination inhibition antibodies and total IgGs were mostly induced by the major antigenic sites. However, in human adults, we observed high hemagglutination inhibition antibody responses toward the noncanonical epitopes. By stratifying the human samples into age groups, we found that the noncanonical antibody responses appeared to increase with age. IMPORTANCE This study dissected the specific antibody responses toward the major antigenic sites and the noncanonical epitopes of influenza B virus Hemagglutinin in animals and humans using novel reagents. These findings will guide the design of the next generation of influenza virus vaccines.
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immunodominance of antigenic site b in the Hemagglutinin of the current h3n2 influenza virus in humans and mice
Journal of Virology, 2018Co-Authors: Felix Broecker, Florian Krammer, Viviana Simon, Peter PaleseAbstract:The Hemagglutinin protein of H3N2 influenza viruses is the major target of neutralizing antibodies induced by infection and vaccination. However, the virus frequently escapes antibody-mediated neutralization due to mutations in the globular head domain. Five topologically distinct antigenic sites in the head domain of H3 Hemagglutinin, A to E, have been previously described by mapping the binding sites of monoclonal antibodies, yet little is known about the contribution of each site to the immunogenicity of modern H3 Hemagglutinins, as measured by hemagglutination inhibition activity, which is known to correlate with protection. To investigate the hierarchy of antibody immunodominance, five Δ1 recombinant influenza viruses expressing Hemagglutinin of the A/Hong Kong/4801/2014 (H3N2) strain with mutations in single antigenic sites were generated. Next, the Δ1 viruses were used to determine the hierarchy of immunodominance by measuring the hemagglutination inhibition reactivity of mouse antisera and plasma from 18 human subjects before and after seasonal influenza vaccination in 2017-2018. In both mice and humans, mutations in antigenic site B caused the most significant decrease in hemagglutination inhibition titers compared to wild-type Hemagglutinin. This study revealed that antigenic site B is immunodominant in the H3N2 influenza virus strain included in the current vaccine preparations. IMPORTANCE Influenza viruses rapidly evade humoral immunity through antigenic drift, making current vaccines poorly effective and antibody-mediated protection short-lived. The majority of neutralizing antibodies target five antigenic sites in the head domain of the Hemagglutinin protein that are also the most sequence-variable regions. A better understanding of the contribution of each antigenic site to the overall antibody response to Hemagglutinin may help in the design of improved influenza virus vaccines.
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age dependence and isotype specificity of influenza virus Hemagglutinin stalk reactive antibodies in humans
Mbio, 2016Co-Authors: Raffael Nachbagauer, Peter Palese, Angela Choi, Ruvim Izikson, Manon M J Cox, Florian KrammerAbstract:ABSTRACT Influenza remains a major global health burden. Seasonal vaccines offer protection but can be rendered less effective when the virus undergoes extensive antigenic drift. Antibodies that target the highly conserved Hemagglutinin stalk can protect against drifted viruses, and vaccine constructs designed to induce such antibodies form the basis for a universal influenza virus vaccine approach. In this study, we analyzed baseline and postvaccination serum samples of children (6 to 59 months), adults (18 to 49 years), and elderly individuals (≥65 years) who participated in clinical trials with a recombinant Hemagglutinin-based vaccine. We found that baseline IgG and IgA antibodies against the H1 stalk domain correlated with the ages of patients. Children generally had very low baseline titers and did not respond well to the vaccine in terms of making stalk-specific antibodies. Adults showed the highest induction of stalk-specific antibodies, but the elderly had the highest absolute antibody titers against the stalk. Importantly, the stalk antibodies measured by enzyme-linked immunosorbent assay (ELISA) showed neutralizing activity in neutralization assays and protected mice in a passive-transfer model in a stalk titer-dependent manner. Finally, we found similar patterns of stalk-specific antibodies directed against the H3 and influenza B virus Hemagglutinins, albeit at lower levels than those measured against the H1 stalk. The relatively high levels of stalk-specific antibodies in the elderly patients may explain the previously reported low influenza virus infection rates in this age group. (This study has been registered at ClinicalTrials.gov under registration no. NCT00336453, NCT00539981, and NCT00395174.) IMPORTANCE The present study provides evidence that titers of broadly neutralizing Hemagglutinin stalk-reactive antibodies increase with age, possibly due to repeated exposure to divergent influenza viruses. These relatively high levels of antistalk titers may be responsible for lower circulation rates of influenza viruses in older individuals. Our findings suggest that the level of antistalk antibodies is a good surrogate marker for protection against influenza virus infection. In addition, the levels of antistalk antibodies might determine the breadth of protection against different drifted strains.
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induction of broadly reactive anti Hemagglutinin stalk antibodies by an h5n1 vaccine in humans
Journal of Virology, 2014Co-Authors: Raffael Nachbagauer, Ariana Hirsh, Peter Palese, Rong Hai, Teddy John Wohlbold, Haakon Sjursen, Rebecca J Cox, Florian KrammerAbstract:ABSTRACT Influenza virus infections are a major public health concern and cause significant morbidity and mortality worldwide. Current vaccines are effective but strain specific due to their focus on the immunodominant globular head domain of the Hemagglutinin (HA). It has been hypothesized that sequential exposure of humans to Hemagglutinins with divergent globular head domains but conserved stalk domains could refocus the immune response to broadly neutralizing epitopes in the stalk. Humans have preexisting immunity against H1 (group 1 Hemagglutinin), and vaccination with H5 HA (also group 1)—which has a divergent globular head domain but a similar stalk domain—represents one such sequential-exposure scenario. To test this hypothesis, we used novel reagents based on chimeric Hemagglutinins to screen sera from an H5N1 clinical trial for induction of stalk-specific antibodies by quantitative enzyme-linked immunosorbent assay (ELISA) and neutralization assays. Importantly, we also investigated the biological activity of these antibodies in a passive transfer in a mouse challenge model. We found that the H5N1 vaccine induced high titers of stalk-reactive antibodies which were biologically active and protective in the passive-transfer experiment. The induced response showed exceptional breadth toward divergent group 1 Hemagglutinins but did not extend to group 2 Hemagglutinins. These data provide evidence for the hypothesis that sequential exposure to Hemagglutinins with divergent globular head domains but conserved stalk domains can refocus the immune response toward the conserved stalk domain. Furthermore, the results support the concept of a chimeric Hemagglutinin universal influenza virus vaccine strategy that is based on the same principle. IMPORTANCE Influenza virus vaccines have to be reformulated and readministered on an annual basis. The development of a universal influenza virus vaccine could abolish the need for this cumbersome and costly process and would also enhance our pandemic preparedness. This study addressed the following questions, which are essential for the development of a Hemagglutinin stalk-based universal influenza virus vaccine. (i) Can stalk-reactive antibodies be boosted by vaccination with divergent HAs that share conserved epitopes? (ii) How long-lived are these vaccine-induced stalk-reactive antibody responses? (iii) What is the breadth of this reactivity? (iv) Are these antibodies functional and protective? Our results further strengthen the concept of induction of stalk-reactive antibodies by sequential exposure to Hemagglutinin immunogens with conserved stalk and divergent head domains. A universal influenza virus vaccine based on the same principles seems possible and might have a significant impact on global human health.
Irina Margine - One of the best experts on this subject based on the ideXlab platform.
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one shot vaccination with an insect cell derived low dose influenza a h7 virus like particle preparation protects mice against h7n9 challenge
Vaccine, 2014Co-Authors: Miriam Klausberger, Monika Wilde, Dieter Palmberger, Irina Margine, Ariana Hirsh, Randy A Albrecht, Adolfo Garciasastre, Reingard Grabherr, Florian KrammerAbstract:Human infections with a novel influenza A H7N9 subtype virus were reported in China recently. The virus caused severe disease with high mortality rates and it raised concerns over its pandemic potential. Here, we assessed in the mouse model protective efficacy of single immunisations with low vaccine doses of insect cell-derived H7 virus-like particles, consisting of Hemagglutinin and matrix protein. Vaccinated mice were fully protected and survived a stringent lethal challenge (100 mLD50) with H7N9, even after a single, unadjuvanted, low vaccine dose (0.03 μg). Serum analysis revealed broad reactivity and hemagglutination inhibition activity across a panel of divergent H7 strains. Moreover, we detected significant levels of cross-reactivity to related group 2 Hemagglutinins. These data demonstrate that virus-like particle vaccines have the potential to induce broadly protective immunity against the novel H7N9 virus and a variety of other H7 strains.
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Hemagglutinin stalk based universal vaccine constructs protect against group 2 influenza a viruses
Journal of Virology, 2013Co-Authors: Irina Margine, Randy A Albrecht, Florian Krammer, Rong Hai, Nicholas S Heaton, Gene Tan, S A Andrews, Jonathan A Runstadler, Patrick C Wilson, Adolfo GarciasastreAbstract:ABSTRACT Current influenza virus vaccines contain H1N1 (phylogenetic group 1 Hemagglutinin), H3N2 (phylogenetic group 2 Hemagglutinin), and influenza B virus components. These vaccines induce good protection against closely matched strains by predominantly eliciting antibodies against the membrane distal globular head domain of their respective viral Hemagglutinins. This domain, however, undergoes rapid antigenic drift, allowing the virus to escape neutralizing antibody responses. The membrane proximal stalk domain of the Hemagglutinin is much more conserved compared to the head domain. In recent years, a growing collection of antibodies that neutralize a broad range of influenza virus strains and subtypes by binding to this domain has been isolated. Here, we demonstrate that a vaccination strategy based on the stalk domain of the H3 Hemagglutinin (group 2) induces in mice broadly neutralizing anti-stalk antibodies that are highly cross-reactive to heterologous H3, H10, H14, H15, and H7 (derived from the novel Chinese H7N9 virus) Hemagglutinins. Furthermore, we demonstrate that these antibodies confer broad protection against influenza viruses expressing various group 2 Hemagglutinins, including an H7 subtype. Through passive transfer experiments, we show that the protection is mediated mainly by neutralizing antibodies against the stalk domain. Our data suggest that, in mice, a vaccine strategy based on the Hemagglutinin stalk domain can protect against viruses expressing divergent group 2 Hemagglutinins.
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h3n2 influenza virus infection induces broadly reactive Hemagglutinin stalk antibodies in humans and mice
Journal of Virology, 2013Co-Authors: Irina Margine, Randy A Albrecht, Adolfo Garciasastre, Peter Palese, Rong Hai, Gerlinde Obermoser, A C Harrod, Jacques Banchereau, Karolina Palucka, John J TreanorAbstract:Broadly neutralizing antibodies directed against the conserved stalk domain of the viral Hemagglutinin have attracted increasing attention in recent years. However, only a limited number of stalk antibodies directed against group 2 influenza Hemagglutinins have been isolated so far. Also, little is known about the general level of induction of these antibodies by influenza virus vaccination or infection. To characterize the anti-stalk humoral response in the mouse model as well as in humans, chimeric Hemagglutinin constructs previously developed in our group were employed in serological assays. Whereas influenza virus infection induced high titers of stalk-reactive antibodies, immunization with inactivated influenza virus vaccines failed to do so in the mouse model. Analysis of serum samples collected from human individuals who were infected by influenza viruses also revealed the induction of stalk-reactive antibodies. Finally, we show that the Hemagglutinin stalk-directed antibodies induced in mice and humans have broad reactivity and neutralizing activity in vitro and in vivo. The results of the study point toward the existence of highly conserved epitopes in the stalk domains of group 2 Hemagglutinins, which can be targeted for the development of a universal influenza virus vaccine in humans.
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a carboxy terminal trimerization domain stabilizes conformational epitopes on the stalk domain of soluble recombinant Hemagglutinin substrates
PLOS ONE, 2012Co-Authors: Florian Krammer, Irina Margine, Gene S Tan, Natalie Pica, Jens C Krause, Peter PaleseAbstract:Recently, a new class of broadly neutralizing anti-influenza virus antibodies that target the stalk domain of the viral Hemagglutinin was discovered. As such, induction, isolation, characterization, and quantification of these novel antibodies has become an area of intense research and great interest. Since most of these antibodies bind to conformational epitopes, the structural integrity of Hemagglutinin substrates for the detection and quantification of these antibodies is of high importance. Here we evaluate the binding of these antibodies to soluble, secreted Hemagglutinins with or without a carboxy-terminal trimerization domain based on the natural trimerization domain of T4 phage fibritin. The lack of such a domain completely abolishes binding to group 1 Hemagglutinins and also affects binding to group 2 Hemagglutinins. Additionally, the presence of a trimerization domain positively influences soluble Hemagglutinin stability during expression and purification. Our findings suggest that a carboxy-terminal trimerization domain is a necessary requirement for the structural integrity of stalk epitopes on recombinant soluble influenza virus Hemagglutinin.
Raffael Nachbagauer - One of the best experts on this subject based on the ideXlab platform.
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A mosaic Hemagglutinin-based influenza virus vaccine candidate protects mice from challenge with divergent H3N2 strains
npj Vaccines, 2019Co-Authors: Felix Broecker, Raffael Nachbagauer, Florian Krammer, Weina Sun, Sean T. H. Liu, Nungruthai Suntronwong, Mark J. Bailey, Peter PaleseAbstract:Current seasonal influenza virus vaccines only provide limited, short-lived protection, and antigenic drift in the Hemagglutinin surface glycoprotein necessitates their annual re-formulation and re-administration. To overcome these limitations, universal vaccine strategies that aim at eliciting broadly protective antibodies to conserved epitopes of the Hemagglutinin show promise for protecting against diverse and drifted influenza viruses. Here a vaccination strategy that focuses antibody responses to conserved epitopes of the H3 Hemagglutinin is described. The approach is based on antigenic silencing of the immunodominant major antigenic sites of an H3 protein from 2014 by replacing them with corresponding sequences of exotic avian Hemagglutinins, yielding “mosaic” Hemagglutinins. In mice, vaccination with inactivated viruses expressing mosaic Hemagglutinins induced highly cross-reactive antibodies against the H3 stalk domain that elicited Fc-mediated effector functions in vitro. In addition, the mosaic viruses elicited head-specific antibodies with neutralizing and hemagglutination-inhibiting activity against recent H3N2 viruses in vitro. Immune sera protected mice from heterologous challenge with viruses carrying H3 proteins from 1968 and 1982, whereas immune sera generated with a seasonal vaccine did not protect. Consequently, the mosaic vaccination approach provides a promising avenue toward a universal influenza virus vaccine. Flu vaccines that target the head domain of the viral Hemagglutinin (HA) surface glycoprotein are highly immunogenic, yet the variable nature of this domain across viral strains and its continuous antigenic drift limit vaccine efficacy. As the HA stalk domain is conserved across influenza viruses, targeting this region is a promising strategy to generate universal flu vaccines. However, eliciting high antibody levels against the HA stalk domain and conserved regions in the head domain remains challenging. Here Peter Palese and colleagues at Mount Sinai develop “mosaic” HAs (mHAs) aimed at eliciting antibodies targeting both the stalk domain and epitopes in the head domain outside of the major antigenic sites. These mHAs are based on antigenic silencing of the immunodominant antigenic sites of an H3 protein, which are exchanged with exotic HA sequences from avian influenza viruses. Vaccination of mice with inactivated viruses expressing mHAs induced both stalk- and head-specific antibodies active in vitro, and transfer of sera from mice immunized with mHAs was better than sera from mice immunized with a seasonal vaccine at protecting mice from heterologous challenge with viruses carrying different H3 proteins, demonstrating the potential of mHAs as universal influenza virus vaccine candidates.
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Universal influenza virus vaccines: what can we learn from the human immune response following exposure to H7 subtype viruses?
Frontiers of Medicine, 2017Co-Authors: Daniel Stadlbauer, Philip Meade, Raffael Nachbagauer, Florian KrammerAbstract:Several universal influenza virus vaccine candidates based on eliciting antibodies against the Hemagglutinin stalk domain are in development. Typically, these vaccines induce responses that target group 1 or group 2 Hemagglutinins with little to no cross-group reactivity and protection. Similarly, the majority of human anti-stalk monoclonal antibodies that have been isolated are directed against group 1 or group 2 Hemagglutinins with very few that bind to Hemagglutinins of both groups. Here we review what is known about the human humoral immune response to vaccination and infection with H7 subtype influenza viruses on a polyclonal and monoclonal level. It seems that unlike vaccination with H5 Hemagglutinin, which induces antibody responses mostly restricted to the group 1 stalk domain, H7 exposure induces both group 2 and cross-group antibody responses. A better understanding of this phenomenon and the underlying mechanisms might help to develop future universal influenza virus vaccine candidates.
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age dependence and isotype specificity of influenza virus Hemagglutinin stalk reactive antibodies in humans
Mbio, 2016Co-Authors: Raffael Nachbagauer, Peter Palese, Angela Choi, Ruvim Izikson, Manon M J Cox, Florian KrammerAbstract:ABSTRACT Influenza remains a major global health burden. Seasonal vaccines offer protection but can be rendered less effective when the virus undergoes extensive antigenic drift. Antibodies that target the highly conserved Hemagglutinin stalk can protect against drifted viruses, and vaccine constructs designed to induce such antibodies form the basis for a universal influenza virus vaccine approach. In this study, we analyzed baseline and postvaccination serum samples of children (6 to 59 months), adults (18 to 49 years), and elderly individuals (≥65 years) who participated in clinical trials with a recombinant Hemagglutinin-based vaccine. We found that baseline IgG and IgA antibodies against the H1 stalk domain correlated with the ages of patients. Children generally had very low baseline titers and did not respond well to the vaccine in terms of making stalk-specific antibodies. Adults showed the highest induction of stalk-specific antibodies, but the elderly had the highest absolute antibody titers against the stalk. Importantly, the stalk antibodies measured by enzyme-linked immunosorbent assay (ELISA) showed neutralizing activity in neutralization assays and protected mice in a passive-transfer model in a stalk titer-dependent manner. Finally, we found similar patterns of stalk-specific antibodies directed against the H3 and influenza B virus Hemagglutinins, albeit at lower levels than those measured against the H1 stalk. The relatively high levels of stalk-specific antibodies in the elderly patients may explain the previously reported low influenza virus infection rates in this age group. (This study has been registered at ClinicalTrials.gov under registration no. NCT00336453, NCT00539981, and NCT00395174.) IMPORTANCE The present study provides evidence that titers of broadly neutralizing Hemagglutinin stalk-reactive antibodies increase with age, possibly due to repeated exposure to divergent influenza viruses. These relatively high levels of antistalk titers may be responsible for lower circulation rates of influenza viruses in older individuals. Our findings suggest that the level of antistalk antibodies is a good surrogate marker for protection against influenza virus infection. In addition, the levels of antistalk antibodies might determine the breadth of protection against different drifted strains.
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induction of broadly reactive anti Hemagglutinin stalk antibodies by an h5n1 vaccine in humans
Journal of Virology, 2014Co-Authors: Raffael Nachbagauer, Ariana Hirsh, Peter Palese, Rong Hai, Teddy John Wohlbold, Haakon Sjursen, Rebecca J Cox, Florian KrammerAbstract:ABSTRACT Influenza virus infections are a major public health concern and cause significant morbidity and mortality worldwide. Current vaccines are effective but strain specific due to their focus on the immunodominant globular head domain of the Hemagglutinin (HA). It has been hypothesized that sequential exposure of humans to Hemagglutinins with divergent globular head domains but conserved stalk domains could refocus the immune response to broadly neutralizing epitopes in the stalk. Humans have preexisting immunity against H1 (group 1 Hemagglutinin), and vaccination with H5 HA (also group 1)—which has a divergent globular head domain but a similar stalk domain—represents one such sequential-exposure scenario. To test this hypothesis, we used novel reagents based on chimeric Hemagglutinins to screen sera from an H5N1 clinical trial for induction of stalk-specific antibodies by quantitative enzyme-linked immunosorbent assay (ELISA) and neutralization assays. Importantly, we also investigated the biological activity of these antibodies in a passive transfer in a mouse challenge model. We found that the H5N1 vaccine induced high titers of stalk-reactive antibodies which were biologically active and protective in the passive-transfer experiment. The induced response showed exceptional breadth toward divergent group 1 Hemagglutinins but did not extend to group 2 Hemagglutinins. These data provide evidence for the hypothesis that sequential exposure to Hemagglutinins with divergent globular head domains but conserved stalk domains can refocus the immune response toward the conserved stalk domain. Furthermore, the results support the concept of a chimeric Hemagglutinin universal influenza virus vaccine strategy that is based on the same principle. IMPORTANCE Influenza virus vaccines have to be reformulated and readministered on an annual basis. The development of a universal influenza virus vaccine could abolish the need for this cumbersome and costly process and would also enhance our pandemic preparedness. This study addressed the following questions, which are essential for the development of a Hemagglutinin stalk-based universal influenza virus vaccine. (i) Can stalk-reactive antibodies be boosted by vaccination with divergent HAs that share conserved epitopes? (ii) How long-lived are these vaccine-induced stalk-reactive antibody responses? (iii) What is the breadth of this reactivity? (iv) Are these antibodies functional and protective? Our results further strengthen the concept of induction of stalk-reactive antibodies by sequential exposure to Hemagglutinin immunogens with conserved stalk and divergent head domains. A universal influenza virus vaccine based on the same principles seems possible and might have a significant impact on global human health.
Adolfo Garciasastre - One of the best experts on this subject based on the ideXlab platform.
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one shot vaccination with an insect cell derived low dose influenza a h7 virus like particle preparation protects mice against h7n9 challenge
Vaccine, 2014Co-Authors: Miriam Klausberger, Monika Wilde, Dieter Palmberger, Irina Margine, Ariana Hirsh, Randy A Albrecht, Adolfo Garciasastre, Reingard Grabherr, Florian KrammerAbstract:Human infections with a novel influenza A H7N9 subtype virus were reported in China recently. The virus caused severe disease with high mortality rates and it raised concerns over its pandemic potential. Here, we assessed in the mouse model protective efficacy of single immunisations with low vaccine doses of insect cell-derived H7 virus-like particles, consisting of Hemagglutinin and matrix protein. Vaccinated mice were fully protected and survived a stringent lethal challenge (100 mLD50) with H7N9, even after a single, unadjuvanted, low vaccine dose (0.03 μg). Serum analysis revealed broad reactivity and hemagglutination inhibition activity across a panel of divergent H7 strains. Moreover, we detected significant levels of cross-reactivity to related group 2 Hemagglutinins. These data demonstrate that virus-like particle vaccines have the potential to induce broadly protective immunity against the novel H7N9 virus and a variety of other H7 strains.
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Hemagglutinin stalk based universal vaccine constructs protect against group 2 influenza a viruses
Journal of Virology, 2013Co-Authors: Irina Margine, Randy A Albrecht, Florian Krammer, Rong Hai, Nicholas S Heaton, Gene Tan, S A Andrews, Jonathan A Runstadler, Patrick C Wilson, Adolfo GarciasastreAbstract:ABSTRACT Current influenza virus vaccines contain H1N1 (phylogenetic group 1 Hemagglutinin), H3N2 (phylogenetic group 2 Hemagglutinin), and influenza B virus components. These vaccines induce good protection against closely matched strains by predominantly eliciting antibodies against the membrane distal globular head domain of their respective viral Hemagglutinins. This domain, however, undergoes rapid antigenic drift, allowing the virus to escape neutralizing antibody responses. The membrane proximal stalk domain of the Hemagglutinin is much more conserved compared to the head domain. In recent years, a growing collection of antibodies that neutralize a broad range of influenza virus strains and subtypes by binding to this domain has been isolated. Here, we demonstrate that a vaccination strategy based on the stalk domain of the H3 Hemagglutinin (group 2) induces in mice broadly neutralizing anti-stalk antibodies that are highly cross-reactive to heterologous H3, H10, H14, H15, and H7 (derived from the novel Chinese H7N9 virus) Hemagglutinins. Furthermore, we demonstrate that these antibodies confer broad protection against influenza viruses expressing various group 2 Hemagglutinins, including an H7 subtype. Through passive transfer experiments, we show that the protection is mediated mainly by neutralizing antibodies against the stalk domain. Our data suggest that, in mice, a vaccine strategy based on the Hemagglutinin stalk domain can protect against viruses expressing divergent group 2 Hemagglutinins.
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h3n2 influenza virus infection induces broadly reactive Hemagglutinin stalk antibodies in humans and mice
Journal of Virology, 2013Co-Authors: Irina Margine, Randy A Albrecht, Adolfo Garciasastre, Peter Palese, Rong Hai, Gerlinde Obermoser, A C Harrod, Jacques Banchereau, Karolina Palucka, John J TreanorAbstract:Broadly neutralizing antibodies directed against the conserved stalk domain of the viral Hemagglutinin have attracted increasing attention in recent years. However, only a limited number of stalk antibodies directed against group 2 influenza Hemagglutinins have been isolated so far. Also, little is known about the general level of induction of these antibodies by influenza virus vaccination or infection. To characterize the anti-stalk humoral response in the mouse model as well as in humans, chimeric Hemagglutinin constructs previously developed in our group were employed in serological assays. Whereas influenza virus infection induced high titers of stalk-reactive antibodies, immunization with inactivated influenza virus vaccines failed to do so in the mouse model. Analysis of serum samples collected from human individuals who were infected by influenza viruses also revealed the induction of stalk-reactive antibodies. Finally, we show that the Hemagglutinin stalk-directed antibodies induced in mice and humans have broad reactivity and neutralizing activity in vitro and in vivo. The results of the study point toward the existence of highly conserved epitopes in the stalk domains of group 2 Hemagglutinins, which can be targeted for the development of a universal influenza virus vaccine in humans.