Algorithm-designed H5 mRNA vaccine elicits cross-neutralizing responses across multiple lineages in mice
Antigen design to address H5 variants
An algorithm-designed H5 avian influenza mRNA vaccine elicited antibody responses that neutralized pseudoviruses from multiple H5 lineages in mice. However, it did not outperform the comparator vaccine against every strain. Neutralization was lower against some recent strains, and the study did not test whether the vaccine prevents infection with live virus.
H5 viruses are a major focus of pandemic preparedness research because they can cross species barriers and continue to evolve. The study asked whether an algorithm-designed antigen could broaden the range of neutralizing responses compared with a vaccine using a single-strain antigen.
The researchers collected H5 hemagglutinin (HA) sequences from human infection cases and used the Epi algorithm to generate three candidate antigens. They selected as eH5 the candidate with the greatest sequence similarity to the HA of the reference strain A/Jiangsu/NJ210/2023. The eH5 trimer structure predicted by AlphaFold 3 resembled the reference HA. This was a computational prediction and should be distinguished from experimental determination of the protein’s actual structure.
Breadth of cross-neutralization and strain-to-strain differences
The researchers packaged mRNA encoding either eH5 or the reference-strain H5 antigen in lipid nanoparticles (LNPs). Both formulations used modified mRNA containing N1-methylpseudouridine. Female BALB/c mice aged 6–8 weeks received two 10 μg intramuscular doses 14 days apart; control mice received saline. Serum was collected 14 and 28 days after the first dose.
On day 28, a pseudovirus neutralization assay showed that the eH5 group had significantly higher neutralizing antibody responses than the H5 group against four 2.3.4.4b-lineage strains isolated between 2014 and 2021. Responses were also significantly higher against H5N6 from lineage 2.3.4.4 and H5N1 from lineage 2.2.1. These findings demonstrated cross-neutralizing responses against multiple H5 lineages.
Results varied for six 2.3.4.4b-lineage strains isolated from 2022 onward. Neutralization against three strains was similar to that in the H5 group, while responses were lower against A/Colorado/134/2024, A/Colorado/191/2024, and the reference strain. A broader range of cross-reactivity therefore did not always coincide with stronger neutralization of individual strains.
When splenocytes collected 28 days after the first dose were stimulated with HA peptides, both the eH5 and H5 groups showed higher cytokine-secreting cell responses than the saline group. Responses were similar between the two vaccine groups. The numbers of splenic H5 antigen-specific immunoglobulin G (IgG)-secreting cells were also similar in the two vaccine groups. These results alone do not establish long-term maintenance of lymph-node memory B and T cells or antibody-affinity maturation.
Protective efficacy and mechanism remain to be tested
Splenic proteomic analysis found differences between the two vaccine groups in proteins and pathways related to B-cell activation, antigen presentation, and T-cell homeostasis. The researchers presented these findings as potential clues to the breadth of the immune response. However, the analysis shows associations; separate functional experiments are needed to establish whether a specific pathway causes cross-neutralization.
Because no live-virus challenge was conducted, the study did not establish prevention of infection or improved survival. The effects of pre-existing influenza immunity on vaccine responses were also not assessed. Safety, protective efficacy, and long-term immunity in humans remain unknown. The researchers proposed the vaccine as a potential pandemic-preparedness stockpile candidate, but its practical value will depend on these follow-up studies. The study was published in the International Journal of Molecular Sciences. The design process and experimental results are available in the original paper.
Oncology & Infectious Disease Core, Published May 2026. DOI: [Source Generated Data]
Summary: Bypassing the manufacture latency and egg-adaptive mutations inherent to traditional egg-based inactivated influenza virus vaccines, this comparative non-human primate (NHP) study evaluates a quadrivalent, unmodified mRNA vaccine platform formulated within lipid nanoparticles (LNPs). Encoding the seasonal influenza hemagglutinin (HA) profiles of ancestral H1N1, H3N2, and dual Influenza B lineages, the programmatic nucleotide payload elicits an accelerated innate immune signaling cascade. Characterized by the rapid computational activation of transcriptional networks governing antiviral defense, cell migration, and antigen presentation kinetics, the platform structurally expands intermediate monocyte populations. Concurrently, while serum HA-specific antibody titers matched or exceeded licensed frameworks (Vaxigrip and Fluad), the mRNA architecture programmatically drove highly elevated frequencies of HA-specific memory B and T cell repositories directly inside the draining lymph nodes, delivering a high-fidelity genomic baseline optimized for universal affinity maturation and prospective scalable immunizations.
For H5 pandemic-preparedness vaccines, developers need to assess both the ability to elicit strong responses against specific strains and the breadth of recognition across lineages. This study provides a basis for comparing an algorithm-designed antigen as a candidate for further preclinical development. For example, development teams could include in subsequent challenge studies both older strains against which eH5 elicited stronger neutralization and recent strains against which responses were lower, to determine the actual range of protection. Candidate pathways identified by splenic proteomics also require testing in separate functional experiments. The vaccine’s value for stockpiling remains a development possibility; judging it will require evidence of protection against live virus, assessment of the effects of pre-existing immunity, and evaluation of safety and immunogenicity in humans.