Metavac®-RSV mucosal bivalent vaccine candidate protects cotton rats against RSV and HMPV and its production process in serum-free Vero cells is scalable in a 2L bioreactor

Metavac®-RSV mucosal bivalent vaccine candidate protects cotton rats against RSV and HMPV and its production process in serum-free Vero cells is scalable in a 2L bioreactor | 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 Article Metavac®-RSV mucosal bivalent vaccine candidate protects cotton rats against RSV and HMPV and its production process in serum-free Vero cells is scalable in a 2L bioreactor Caroline CHUPIN, Pauline BRUN, Marjorie RAY, Chloé MIALON, Maëlle REITANO, and 12 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6217777/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 22 Aug, 2025 Read the published version in npj Vaccines → Version 1 posted 12 You are reading this latest preprint version Abstract Respiratory Syncytial Virus (RSV) and Human Metapneumovirus (HMPV) are the main etiologic agents of viral bronchiolitis and pneumonia in children and elderly. As Live-Attenuated Vaccines (LAV) can stimulate robust mucosal and cellular responses, we previously engineered an HMPV-based bivalent LAV Metavac®-RSV and reported its capacity to protect mice against HMPV and RSV challenges after intranasal delivery and to induce strong neutralizing antibody responses. To progress towards clinical development, we identified a GMP grade Vero cell platform as permissive and efficient to produce high yields of functional Metavac®-RSV. Deep sequencing and immunostaining analysis showed that Metavac®-RSV maintains genetic stability and RSV and HMPV F antigens expression after several passages in Vero cells. Furthermore, Metavac®-RSV produced in Vero cells protected cotton rats against both HMPV and RSV challenges, reducing significantly viral replication in the respiratory airways and inducing high titers of neutralizing antibodies production. Finally, we identified process parameters to scale-up the production process of infectious Metavac®-RSV using Vero cells cultivated on microcarriers in 2L single-use stirred-tank bioreactor. Altogether, these results underscore the potential of Metavac®-RSV as a promising LAV candidate for mucosal administration that could bring sterilizing immunity against pneumovirus infections, with a scalable upstream production process amenable to industrial manufacturing. Biological sciences/Immunology/Vaccines Biological sciences/Microbiology/Vaccines/Live attenuated vaccines Biological sciences/Immunology/Infectious diseases RSV HMPV chimeric virus bivalent vaccine Live Attenuated Vaccine vectored vaccine reverse genetics gene insertion respiratory viruses pneumovirus bronchiolitis scale-up bioprocess Full Text Additional Declarations Competing interest reported. Manuel Rosa-Calatrava, Guy Boivin, Julia Dubois, and Marie-Eve Hamelin are co-founders and shareholders of Vaxxel SAS. Caroline Chupin is currently employee of Vaxxel SAS and Marjorie Ray was employee of Vaxxel SAS. The other authors declare no competing interests. The authors declare the following patent : EP22305240.2 – PCT/EP2023/055221 concerning Vaccine composition against two respiratory viruses (Inventors: Daniela Ogonczyk-Makowska, Jean-François Eléouët, Guy Boivin, Julia Dubois and Manuel Rosa-Calatrava ; Applicants : Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS), Université Claude Bernard Lyon 1 (UCBL) Ecole Normale Supérieure de Lyon (ENS Lyon), Institut National de Recherche pour l'Agriculture, l'Alimentation et l'Environnement (INRAE) and Vaxxel SAS). Supplementary Files Supplementaryfigures.docx Cite Share Download PDF Status: Published Journal Publication published 22 Aug, 2025 Read the published version in npj Vaccines → Version 1 posted Editorial decision: Revision requested 11 Jun, 2025 Reviews received at journal 27 May, 2025 Reviews received at journal 11 May, 2025 Reviewers agreed at journal 30 Apr, 2025 Reviewers agreed at journal 28 Apr, 2025 Reviews received at journal 23 Apr, 2025 Reviewers agreed at journal 02 Apr, 2025 Reviewers agreed at journal 02 Apr, 2025 Reviewers invited by journal 26 Mar, 2025 Editor assigned by journal 26 Mar, 2025 Submission checks completed at journal 17 Mar, 2025 First submitted to journal 13 Mar, 2025 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. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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-6217777","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":441291435,"identity":"e89d3e91-c168-4fad-a3a3-790870ed50ac","order_by":0,"name":"Caroline CHUPIN","email":"","orcid":"","institution":"CIRI, Centre International de Recherche en Infectiologie, Team VirPath, Univ Lyon, CNRS, UMR5308, ENS de Lyon","correspondingAuthor":false,"prefix":"","firstName":"Caroline","middleName":"","lastName":"CHUPIN","suffix":""},{"id":441291436,"identity":"bbabdd5f-547c-41f0-88ac-48c1885bee8c","order_by":1,"name":"Pauline BRUN","email":"","orcid":"","institution":"International Research Laboratory RESPIVIR France - 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Manuel Rosa-Calatrava, Guy Boivin, Julia Dubois, and Marie-Eve Hamelin are co-founders and shareholders of Vaxxel SAS. Caroline Chupin is currently employee of Vaxxel SAS and Marjorie Ray was employee of Vaxxel SAS. The other authors declare no competing interests.\nThe authors declare the following patent : EP22305240.2 – PCT/EP2023/055221 concerning Vaccine composition against two respiratory viruses (Inventors: Daniela Ogonczyk-Makowska, Jean-François Eléouët, Guy Boivin, Julia Dubois and Manuel Rosa-Calatrava ; Applicants : Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS), Université Claude Bernard Lyon 1 (UCBL) Ecole Normale Supérieure de Lyon (ENS Lyon), Institut National de Recherche pour l'Agriculture, l'Alimentation et l'Environnement (INRAE) and Vaxxel SAS).","formattedTitle":"Metavac®-RSV mucosal bivalent vaccine candidate protects cotton rats against RSV and HMPV and its production process in serum-free Vero cells is scalable in a 2L bioreactor","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"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":"npj-vaccines","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"npjvaccines","sideBox":"Learn more about [npj Vaccines](http://www.nature.com/npjvaccines/)","snPcode":"41541","submissionUrl":"https://submission.springernature.com/new-submission/41541/3?","title":"npj Vaccines","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"NPJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"RSV, HMPV, chimeric virus, bivalent vaccine, Live Attenuated Vaccine, vectored vaccine, reverse genetics, gene insertion, respiratory viruses, pneumovirus, bronchiolitis, scale-up, bioprocess","lastPublishedDoi":"10.21203/rs.3.rs-6217777/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6217777/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Respiratory Syncytial Virus (RSV) and Human Metapneumovirus (HMPV) are the main etiologic agents of viral bronchiolitis and pneumonia in children and elderly. As Live-Attenuated Vaccines (LAV) can stimulate robust mucosal and cellular responses, we previously engineered an HMPV-based bivalent LAV Metavac®-RSV and reported its capacity to protect mice against HMPV and RSV challenges after intranasal delivery and to induce strong neutralizing antibody responses. To progress towards clinical development, we identified a GMP grade Vero cell platform as permissive and efficient to produce high yields of functional Metavac®-RSV. Deep sequencing and immunostaining analysis showed that Metavac®-RSV maintains genetic stability and RSV and HMPV F antigens expression after several passages in Vero cells. Furthermore, Metavac®-RSV produced in Vero cells protected cotton rats against both HMPV and RSV challenges, reducing significantly viral replication in the respiratory airways and inducing high titers of neutralizing antibodies production. Finally, we identified process parameters to scale-up the production process of infectious Metavac®-RSV using Vero cells cultivated on microcarriers in 2L single-use stirred-tank bioreactor. 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