Bivalent mRNA booster encoding virus-like particles elicits potent polyclass RBD antibodies in pre-vaccinated mice
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Curated by eLife
eLife Assessment
This report provides useful evidence that EABR mRNA is at least as effective as standard S mRNA vaccines for the SARS-CoV-2 booster vaccine. Although the methodology and the experimental approaches are solid, the inconsistent statistical significance throughout the study presents limitations in interpreting the results. Also, the absence of results showing possible mechanisms underlying the lack of benefit with EABR in the pre-immune makes the findings mostly observational.
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Abstract
mRNA vaccines emerged as a leading vaccine technology during the COVID-19 pandemic. However, their sustained protective efficacies were limited by relatively short-lived antibody responses and the emergence of SARS-CoV-2 variants, necessitating frequent and variant-updated boosters. We recently developed the ESCRT- and ALIX-binding region (EABR) mRNA vaccine platform, which encodes engineered immunogens that induce budding of enveloped virus-like particles (eVLPs) from the plasma membrane, thereby resulting in presentation of immunogens on cell surfaces and eVLPs. Prior studies showed that spike (S)-EABR mRNA-LNP immunizations elicited enhanced neutralizing antibody responses against ancestral and variant SARS-CoV-2 compared with conventional S mRNA-LNP in naïve mice, but the effectiveness of S-EABR mRNA-LNP boosters in the context of pre-existing immunity has not been investigated. Here, we evaluated monovalent Wuhan-Hu-1 (Wu1) and bivalent (Wu1/BA.5) S-EABR mRNA-LNP boosters in mice pre-vaccinated with conventional Wu1 S mRNA-LNP. Compared to conventional S mRNA-LNP boosters, the EABR approach enhanced monovalent and bivalent mRNA-LNP booster-induced neutralizing responses against Omicron subvariants BA.1, BA.5, BQ.1.1, and XBB.1, with bivalent S-EABR mRNA-LNP consistently eliciting the highest titers. Epitope mapping of polyclonal antisera by deep mutational scanning revealed that bivalent S-EABR mRNA-LNP boosted diverse “polyclass” anti-RBD responses, suggesting balanced targeting of multiple RBD epitope classes. In contrast, monovalent S, bivalent S, and monovalent S-EABR mRNA-LNP boosters elicited less diverse polyclonal serum responses primarily targeting immunodominant RBD epitopes. Cryo-EM structures demonstrated that bivalent mRNA immunizations promote S heterotrimer formation, potentially enhancing bivalent S-EABR mRNA-LNP booster-induced antibody breadth and polyclass epitope targeting by activating cross-reactive B cells through intra-S crosslinking. These findings support the future design of bivalent or multivalent S-EABR mRNA-LNP boosters as a promising strategy to confer broader, and therefore potentially more durable, protection against emerging SARS-CoV-2 variants and other rapidly evolving viruses.
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eLife Assessment
This report provides useful evidence that EABR mRNA is at least as effective as standard S mRNA vaccines for the SARS-CoV-2 booster vaccine. Although the methodology and the experimental approaches are solid, the inconsistent statistical significance throughout the study presents limitations in interpreting the results. Also, the absence of results showing possible mechanisms underlying the lack of benefit with EABR in the pre-immune makes the findings mostly observational.
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Reviewer #1 (Public review):
Summary:
This study investigated the immunogenicity of a novel bivalent EABR mRNA vaccine for SARS-CoV-2 that expresses enveloped virus-like particles in pre-immune mice as a model for boosting the population that is already pre-immune to SARS-CoV-2. The study builds on promising data showing a monovalent EABR mRNA vaccine induced substantially higher antibody responses than a standard S mRNA vaccine in naïve mice. In pre-immune mice, the EABR booster increased the breadth and magnitude of the antibody response, but the effects were modest and often not statistically significant.
Strengths:
Evaluating a novel SARS-CoV-2 vaccine that was substantially superior in naive mice in pre-immune mice as a model for its potential in the pre-immune population.
Weaknesses:
(1) Overall, immune responses against Omicron …
Reviewer #1 (Public review):
Summary:
This study investigated the immunogenicity of a novel bivalent EABR mRNA vaccine for SARS-CoV-2 that expresses enveloped virus-like particles in pre-immune mice as a model for boosting the population that is already pre-immune to SARS-CoV-2. The study builds on promising data showing a monovalent EABR mRNA vaccine induced substantially higher antibody responses than a standard S mRNA vaccine in naïve mice. In pre-immune mice, the EABR booster increased the breadth and magnitude of the antibody response, but the effects were modest and often not statistically significant.
Strengths:
Evaluating a novel SARS-CoV-2 vaccine that was substantially superior in naive mice in pre-immune mice as a model for its potential in the pre-immune population.
Weaknesses:
(1) Overall, immune responses against Omicron variants were substantially lower than against the ancestral Wu-1 strain that the mice were primed with. The authors speculate this is evidence of immune imprinting, but don't have the appropriate controls (mice immunized 3 times with just the bivalent EABR vaccine) to discern this. Without this control, it's not clear if the lower immune responses to Omicron are due to immune imprinting (or original antigenic sin) or because the Omicron S immunogen is just inherently more poorly immunogenic than the S protein from the ancestral Wu-1 strain.
(2) The authors reported a statistically significant increase in antibody responses with the bivalent EABR vaccine booster when compared to the monovalent S mRNA vaccine, but consistently failed to show significantly higher responses when compared to the bivalent S mRNA vaccine, suggesting that in pre-immune mice, the EABR vaccine has no apparent advantage over the bivalent S mRNA vaccine which is the current standard. There were, however, some trends indicating the group sizes were insufficiently powered to see a difference. This is mostly glossed over throughout the manuscript. The discussion section needs to better acknowledge these limitations of their studies and the limited benefits of the EABR strategy in pre-immune mice vs the standard bivalent mRNA vaccine.
(3) The discussion would benefit from additional explanation about why they think the EABR S mRNA vaccine was substantially superior in naïve mice vs the standard S mRNA vaccine in their previously published work, but here, there is not much difference in pre-immune mice.
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Reviewer #2 (Public review):
Summary:
In this manuscript, Fan, Cohen, and Dam et al. conducted a follow-up study to their prior work on the ESCRT- and ALIX-binding region (EABR) mRNA vaccine platform that they developed. They tested in mice whether vaccines made in this format will have improved binding/neutralization antibody capacity over conventional antigens when used as a booster. The authors tested this in both monovalent (Wu1 only) or bivalent (Wu1 + BA.5) designs. The authors found that across both monovalent and bivalent designs, the EABR antigens had improved antibody titers than conventional antigens, although they observed dampened titers against Omicron variants, likely due to immune imprinting. Deep mutational scanning experiments suggested that the improvement of the EABR format may be due to a more diversified antibody …
Reviewer #2 (Public review):
Summary:
In this manuscript, Fan, Cohen, and Dam et al. conducted a follow-up study to their prior work on the ESCRT- and ALIX-binding region (EABR) mRNA vaccine platform that they developed. They tested in mice whether vaccines made in this format will have improved binding/neutralization antibody capacity over conventional antigens when used as a booster. The authors tested this in both monovalent (Wu1 only) or bivalent (Wu1 + BA.5) designs. The authors found that across both monovalent and bivalent designs, the EABR antigens had improved antibody titers than conventional antigens, although they observed dampened titers against Omicron variants, likely due to immune imprinting. Deep mutational scanning experiments suggested that the improvement of the EABR format may be due to a more diversified antibody response. Finally, the authors demonstrate that co-expression of multiple spike proteins within a single cell can result in the formation of heterotrimers, which may have potential further usage as an antigen.
Strengths:
(1) The experiments are conducted well and are appropriate to address the questions at hand. Given the significant time that is needed for testing of pre-existing immunity, due to the requirement of pre-vaccinated animals, it is a strength that the authors have conducted a thorough experiment with appropriate groups.
(2) The improvement in titers associated with EABR antigens bodes well for its potential use as a vaccine platform.
Weaknesses:
As noted above, this type of study requires quite a bit of initial time, so the authors cannot be blamed for this, but unfortunately, the vaccine designs that were tested are quite outdated. BA.5 has long been replaced by other variants, and importantly, bivalent vaccines are no longer used. Testing of contemporaneous strains as well as monovalent variant vaccines would be desirable to support the study.
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