PONE-D-20-28839

Construction, characterization, and immunization of nanoparticles that display a diverse array of influenza HA trimers

PLOS ONE

Dear Dr. Bjorkman,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

During the revision process, please address the issues related to choice of vaccine delivery route and provide additional insight into the potential responses in the germinal center and at the T cell level.

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Victor C Huber

Academic Editor

PLOS ONE

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Reviewers' comments:

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Comments to the Author

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Reviewer #1: Yes

Reviewer #2: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

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Reviewer #1: This excellent manuscript by Cohen et al. describes nice “Plug and Display” virus like particle (VLP) and nanoparticle (NP) vaccine platforms to present trimeric HA proteins. Both platforms are immunogenic and allow quick generation of mosaic particles displaying HAs of different strains, while the NP platform shows improved yields. The group was able to generate VLPs and NP with 2, 4 and 8 HA valency aiming to target the cross-reactive epitopes on different HA strains. As truthfully reported by the authors, this mosaic strategy did not successfully induce cross-reactive antibodies or B-cells. This, on the other hand, seems to support the hypothesis that an immunodominant hierarchy hinders the induction of cross-reactive antibodies, in which the variable epitopes (typically in the globular head domain) are more accessible than the conserved epitopes in the stem by the BCR, regardless of number of epitopes that are presented at the same time. Nevertheless, this study shows promising vaccine platforms that might be more advantageous when used as an improved seasonal vaccine instead of a universal influenza virus vaccine.

1. The HA-VLP in Figure 7 appears to elicit higher antibody responses. The authors hypothesized that this could due to the self-adjuvant effect of the VLPs. However, Page 14, line 2 “The HAs for the SpyCatcher-mi3 conjugations include the sialic acid binding knockout mutation Y98F”; This gives the impression that HAs conjugated with VLP do not have the Y98F mutation. If that is the case, ELISAs performed using recombinant HA without the Y98F could give different results for the two platforms. The authors should provide more clarification of whether the wt or mutant HA were used in the VLP platform and the ELISAs.

2. Is the SpyCatcher immunogenic? Have the author measured antibodies specific to the Spycatcher or the NP/VLP separately?

3. The immunization route and adjuvant used in Fig 4 and 5 are different from those used in Fig 7. Can the author explain a little bit more why that is?

Reviewer #2: The manuscript by Cohen et al. demonstrates that in two different nanoparticle formulations (AP205 VLPs and mi3) carrying multiple strains of influenza HA, there are no major differences in antibody responses between antigen (HA trimers) presented as mosaic particles or admixed particles. Although the results are "negative" per se, the manuscript provides valuable insights for the field and kudos to the authors for such a through work. However, prior to publications several questions should be addressed and some new data needs to presented.

1. The choice of i.p delivery of the vaccine in mice is interesting but unusual. Why was i.p. chosen as the route? Typically intramuscular, intradermal/subcutaneous, or intranasal routes are chosen.

2. Please provide statistics in the figures. There are discussions of the stats in the text, but at least each figure or figure legend should summarize the statistics for the corresponding data.

3. Do the authors have any data on the germinal center reaction for these groups? It is critical to understand, from an Ab response point of view, whether the GC responses were all similar or whether these formulations failed to elicit strong GC responses. This would provide mechanistic insights better than just looking at Ab responses.

4. It is disappointing that no T cell response data were shown. It would much to the study if lymph nod eor splenic T cell assays are performed and reported.

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Reviewer #1: No

Reviewer #2: No

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Construction, characterization, and immunization of nanoparticles that display a diverse array of influenza HA trimers

PONE-D-20-28839R1

Dear Dr. Bjorkman,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

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Kind regards,

Victor C Huber

Academic Editor

PLOS ONE

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