Clade 2.3.4.4b but not historical clade 1 HA replicating RNA vaccine protects against bovine H5N1 challenge

Clade 2.3.4.4b but not historical clade 1 HA replicating RNA vaccine protects against bovine H5N1 challenge | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Biological Sciences - Article Clade 2.3.4.4b but not historical clade 1 HA replicating RNA vaccine protects against bovine H5N1 challenge David Hawman, Thomas Tipih, Eddie Hodge, Kimberly Meade-White, and 9 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4946897/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 14 Jan, 2025 Read the published version in Nature Communications → Version 1 posted You are reading this latest preprint version Abstract The ongoing circulation of influenza A H5N1 in the United States has raised concerns of a pandemic caused by highly pathogenic avian influenza. Although the United States has stockpiled and is prepared to produce millions of vaccine doses to address an H5N1 pandemic, the currently circulating influenza A H5N1 contains multiple mutations within the immunodominant head domain of HA compared to the antigens used in stockpiled vaccines. It is unclear if these stockpiled vaccines will need to be updated to match the contemporary H5N1 strains. Here we show that a replicating RNA vaccine expressing the hemagglutinin (HA) of an H5N1 isolated from a US dairy cow confers complete protection against homologous lethal challenge in mice. Strikingly, a repRNA encoding the HA of a pre-pandemic H5 subtype from 2004 (A/Vietnam/1203/2004), only conferred partial protection. Our data highlight the utility of nucleic acid vaccines to be rapidly updated to match emergent viruses of concern while demonstrating that contemporary bovine H5N1 viruses can evade immunity elicited by historical HA antigens. Biological sciences/Microbiology/Vaccines/RNA vaccines Biological sciences/Microbiology/Virology/Influenza virus Figures Figure 1 Figure 2 Figure 3 Full Text Additional Declarations Yes there is potential Competing Interest. JHE, ETS, NW, SP, KH, EH, and KG have equity interest in HDT Bio. JHE is an inventor on patents (US Patent nos. 11,458,209; 11,433,142; 11,752,218; 11,648,321; and 11,654,200) and patent applications (PCT/US22/76787, PCT/US23/60225, and PCT/US2024/010326) pertaining to the LION formulation and repRNA compositions described in the studies. Cite Share Download PDF Status: Published Journal Publication published 14 Jan, 2025 Read the published version in Nature Communications → Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4946897","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Biological Sciences - Article","associatedPublications":[],"authors":[{"id":342832957,"identity":"1af544e6-9d0b-44ac-93b8-83f86de9e0ba","order_by":0,"name":"David Hawman","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA40lEQVRIiWNgGAWjYDCCA0DEAyJB4AODRAJpWhhnEKuFAaaFmYeBgbAWvuNnDA+8Ybgjb3Dt8LPHtjss8hj4Fx+TwKdF8kyOwcE5DM8MN9xOMzfOPSNRzCDxLA2vFoMDaQmHeRgOM267nWAmndsmkdggccbYAK+W88/AWuy33U7/Jm1JlJYbyQdAWhK33c4xk2YEaeHvMXyA1y83Hh84OMfgcPL+2zllkr1ALW0SbIl4tfCdT2z+8KbisO3M2enbJH621SX28x8+cACfFqjzkNhsxMQmGuAnwo5RMApGwSgYUQAA8mhTzh4hOLUAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0001-8233-8176","institution":"Laboratory of Virology, Division of Intramural Research, NIAID/NIH","correspondingAuthor":true,"prefix":"","firstName":"David","middleName":"","lastName":"Hawman","suffix":""},{"id":342832958,"identity":"a96469a3-54f2-40df-a37c-a7be91eb37fd","order_by":1,"name":"Thomas Tipih","email":"","orcid":"","institution":"NIH/NIAD/Rocky Mountain Laboratories","correspondingAuthor":false,"prefix":"","firstName":"Thomas","middleName":"","lastName":"Tipih","suffix":""},{"id":342832959,"identity":"dde849e7-8859-4f05-9bd1-a2797c3703f9","order_by":2,"name":"Eddie Hodge","email":"","orcid":"","institution":"HDT Bio","correspondingAuthor":false,"prefix":"","firstName":"Eddie","middleName":"","lastName":"Hodge","suffix":""},{"id":342832960,"identity":"93c018ee-cd5d-44a9-a430-de5e65dc2f50","order_by":3,"name":"Kimberly Meade-White","email":"","orcid":"","institution":"NIAID/NIH","correspondingAuthor":false,"prefix":"","firstName":"Kimberly","middleName":"","lastName":"Meade-White","suffix":""},{"id":342832961,"identity":"40656b5d-39b3-44da-8fc6-8187198f72c6","order_by":4,"name":"Shanna Leventhal","email":"","orcid":"","institution":"Laboratory of Virology, Division of Intramural Research, NIAID/NIH","correspondingAuthor":false,"prefix":"","firstName":"Shanna","middleName":"","lastName":"Leventhal","suffix":""},{"id":342832962,"identity":"94aafd25-905e-45c0-99d7-a3a4526603fd","order_by":5,"name":"Kiara Hatzakis","email":"","orcid":"","institution":"HDT Bio","correspondingAuthor":false,"prefix":"","firstName":"Kiara","middleName":"","lastName":"Hatzakis","suffix":""},{"id":342832963,"identity":"b4d09c66-520b-4712-b705-dbf4e3cd91cb","order_by":6,"name":"Stephanie Park","email":"","orcid":"","institution":"HDT Bio","correspondingAuthor":false,"prefix":"","firstName":"Stephanie","middleName":"","lastName":"Park","suffix":""},{"id":342832964,"identity":"1101f003-0a3f-46f2-aa06-4093c8080df7","order_by":7,"name":"Karen Gaffney","email":"","orcid":"","institution":"HDT Bio","correspondingAuthor":false,"prefix":"","firstName":"Karen","middleName":"","lastName":"Gaffney","suffix":""},{"id":342832965,"identity":"5d83a9b5-3360-44eb-9ce2-46ec53a06bec","order_by":8,"name":"E. Taylor Stone","email":"","orcid":"","institution":"HDT Bio","correspondingAuthor":false,"prefix":"","firstName":"E.","middleName":"Taylor","lastName":"Stone","suffix":""},{"id":342832966,"identity":"3d3d6c32-53e8-4792-bcd4-a9a7a6287d38","order_by":9,"name":"Nikole Warner","email":"","orcid":"","institution":"HDT Bio","correspondingAuthor":false,"prefix":"","firstName":"Nikole","middleName":"","lastName":"Warner","suffix":""},{"id":342832967,"identity":"ae57bbbc-1cbc-4f14-9943-e6a64c3f3276","order_by":10,"name":"Kyle Rosenke","email":"","orcid":"","institution":"NIH/NIAD/Rocky Mountain Laboratories","correspondingAuthor":false,"prefix":"","firstName":"Kyle","middleName":"","lastName":"Rosenke","suffix":""},{"id":342832968,"identity":"ac08bdc4-e6c5-43be-b307-c2df999db115","order_by":11,"name":"Jesse Erasmus","email":"","orcid":"https://orcid.org/0000-0003-1612-2697","institution":"HDT Bio Corp","correspondingAuthor":false,"prefix":"","firstName":"Jesse","middleName":"","lastName":"Erasmus","suffix":""},{"id":342832969,"identity":"dbb8a9c2-f4d1-427c-9ccb-2c5427e12cad","order_by":12,"name":"Heinz Feldmann","email":"","orcid":"https://orcid.org/0000-0001-9448-8227","institution":"Laboratory of Virology, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health","correspondingAuthor":false,"prefix":"","firstName":"Heinz","middleName":"","lastName":"Feldmann","suffix":""}],"badges":[],"createdAt":"2024-08-20 18:05:23","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4946897/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4946897/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1038/s41467-024-55546-7","type":"published","date":"2025-01-14T05:00:00+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":63315635,"identity":"44b94f0b-d620-4f11-be8d-78c67c474ce5","added_by":"auto","created_at":"2024-08-26 22:21:44","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":753271,"visible":true,"origin":"","legend":"\u003cp\u003eMutations within the A/bovine HA may impact vaccine efficacy. (a) The amino acid sequence homology between A/Vietnam and A/bovine was mapped onto the HA trimer structure (PDB 6CFG). 2 of the protomers are displayed in surface representation, and 1 protomer is displayed in ribbon representation. Cyan indicates regions with differences in sequence and areas colored white indicate conserved sequence homology. (b) Alphafold2 was used to predict the structure of the Bovine HA (orange) and it was overlayed with the known Vietnam HA structure (PDB 6CFG colored in cyan). (c) The quality of the HA model was assessed via the predicted local distance difference test (pLDDT). Regions colored red and orange are too low scoring in confidence to interpret, and regions colored yellow should be interpreted with caution. Blue indicates high confidence in the predicted structure.\u003c/p\u003e","description":"","filename":"Combined1.png","url":"https://assets-eu.researchsquare.com/files/rs-4946897/v1/3e03ab9e7956ed8ea8f56566.png"},{"id":63315634,"identity":"db3ae31d-aa39-4cad-a7ea-81371047b5f9","added_by":"auto","created_at":"2024-08-26 22:21:43","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":2995250,"visible":true,"origin":"","legend":"\u003cp\u003eRepRNA drives expression of HA and NP in vitro. (a-b) BHK-21 cells were transfected with the indicated repRNA and 24 hours later protein expression for HA (a) or NP (b) evaluated by western blot under reducing (R) or non-reducing (NR) conditions. (c) BHK-21 cells were transfected with indicated repRNA and 24 hours later fixed, permeabilized and stained for HA or NP. Scale bars indicate 50µm.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-4946897/v1/15d8a37b288a365446042fd9.png"},{"id":63315636,"identity":"94e1601d-cfdd-48f8-b6ba-65cb78c04a3e","added_by":"auto","created_at":"2024-08-26 22:21:44","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":309693,"visible":true,"origin":"","legend":"\u003cp\u003eRepHA-Bovine is immunogenic and protective against lethal A/bovine challenge in mice. (a) Mice (n = 3-4 per group) were immunized with indicated repRNAs and HA binding antibodies measured against indicated recombinant HAs (a) or NP (b). (c) Viral neutralization (VN) titers of pooled sera (n = 4 per group) were measured against infectious A/Vietnam or A/bovine virus and data shown as technical replicates. (d-e) Mice (n = 6 per group) were challenged with 100,000 TCID50 of A/bovine via the intranasal route and weighed daily (d) and euthanized when they reached \u0026gt;20% weight loss or exhibited signs such as severe lethargy or neurological signs. (a) P values calculated with an ordinary one-way ANOVA with Tukey’s multiple comparisons test. (e) P values calculated with Log-Rank test with Bonferroni correction for multiple comparisons. * P \u0026lt; 0.05, **P\u0026lt; 0.01, *** P \u0026lt; 0.001, **** P \u0026lt; 0.0001.\u003c/p\u003e","description":"","filename":"Combined3.png","url":"https://assets-eu.researchsquare.com/files/rs-4946897/v1/4562a4dc398972311fa8004a.png"},{"id":73839479,"identity":"be97f95e-72b6-4816-857c-1457dcfb1df7","added_by":"auto","created_at":"2025-01-15 08:06:13","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1204027,"visible":true,"origin":"","legend":"Article File","description":"","filename":"HawmanrepRNAH5.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4946897/v1_covered_94eeb36b-66f1-4a72-acac-455ec7e419e1.pdf"}],"financialInterests":"\u003cb\u003eYes\u003c/b\u003e there is potential Competing Interest.\nJHE, ETS, NW, SP, KH, EH, and KG have equity interest in HDT Bio. JHE is an inventor on patents (US Patent nos. 11,458,209; 11,433,142; 11,752,218; 11,648,321; and 11,654,200) and patent applications (PCT/US22/76787, PCT/US23/60225, and PCT/US2024/010326) pertaining to the LION formulation and repRNA compositions described in the studies.","formattedTitle":"Clade 2.3.4.4b but not historical clade 1 HA replicating RNA vaccine protects against bovine H5N1 challenge","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"nature-portfolio","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"","title":"Nature Portfolio","twitterHandle":"","acdcEnabled":false,"dfaEnabled":false,"editorialSystem":"ejp","reportingPortfolio":"","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-4946897/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4946897/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"The ongoing circulation of influenza A H5N1 in the United States has raised concerns of a pandemic caused by highly pathogenic avian influenza. Although the United States has stockpiled and is prepared to produce millions of vaccine doses to address an H5N1 pandemic, the currently circulating influenza A H5N1 contains multiple mutations within the immunodominant head domain of HA compared to the antigens used in stockpiled vaccines. It is unclear if these stockpiled vaccines will need to be updated to match the contemporary H5N1 strains. Here we show that a replicating RNA vaccine expressing the hemagglutinin (HA) of an H5N1 isolated from a US dairy cow confers complete protection against homologous lethal challenge in mice. Strikingly, a repRNA encoding the HA of a pre-pandemic H5 subtype from 2004 (A/Vietnam/1203/2004), only conferred partial protection. Our data highlight the utility of nucleic acid vaccines to be rapidly updated to match emergent viruses of concern while demonstrating that contemporary bovine H5N1 viruses can evade immunity elicited by historical HA antigens.","manuscriptTitle":"Clade 2.3.4.4b but not historical clade 1 HA replicating RNA vaccine protects against bovine H5N1 challenge","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-08-26 22:21:39","doi":"10.21203/rs.3.rs-4946897/v1","editorialEvents":[],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"nature-communications","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"NCOMMS","sideBox":"Learn more about [Nature Communications](http://www.nature.com/ncomms/)","snPcode":"","submissionUrl":"https://mts-ncomms.nature.com/","title":"Nature Communications","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature Communications","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"64dfd650-8e30-44fe-bea3-e26c519e0528","owner":[],"postedDate":"August 26th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":36323513,"name":"Biological sciences/Microbiology/Vaccines/RNA vaccines"},{"id":36323514,"name":"Biological sciences/Microbiology/Virology/Influenza virus"}],"tags":[],"updatedAt":"2025-01-15T08:06:03+00:00","versionOfRecord":{"articleIdentity":"rs-4946897","link":"https://doi.org/10.1038/s41467-024-55546-7","journal":{"identity":"nature-communications","isVorOnly":false,"title":"Nature Communications"},"publishedOn":"2025-01-14 05:00:00","publishedOnDateReadable":"January 14th, 2025"},"versionCreatedAt":"2024-08-26 22:21:39","video":"","vorDoi":"10.1038/s41467-024-55546-7","vorDoiUrl":"https://doi.org/10.1038/s41467-024-55546-7","workflowStages":[]},"version":"v1","identity":"rs-4946897","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4946897","identity":"rs-4946897","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}