Tumor-educated VEGFR-3 lymphatics augment PD-1 resistance in HPV-negative HNSCC

Tumor-educated VEGFR-3 lymphatics augment PD-1 resistance in HPV-negative HNSCC | 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 Tumor-educated VEGFR-3 lymphatics augment PD-1 resistance in HPV-negative HNSCC Hao Li This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8611043/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract The human papillomavirus (HPV) status is closely related to the response to immune checkpoint inhibitors (ICIs) in head and neck squamous cell carcinoma (HNSCC), yet the mechanisms of immunotherapy resistance in HPV-negative disease remain unclear. We integrated single-cell transcriptomics datasets to investigate the underlying driving factors of ICI resistance in HPV-negative HNSCC. The ecosystem of HPV-negative tumors exhibited a lymphocyte-deficient pattern. Further analyses revealed that this lymphocyte exclusion was driven by aberrantly infiltrated lymphatic endothelial cells (LECs) in HPV-negative microenvironments. Mechanistically, LECs release the excess homing cytokine CCL21, which strongly attracts immune cells via the CCL21‒CCR7 axis and eventually excludes them from the HPV-negative microenvironment. We discovered that HPV-negative tumor cells presented relatively high VEGFC expression, which accelerated the growth of LECs via the VEGF-C‒VEGFR-3 axis. Furthermore, VEGFR-3 blockade significantly facilitated ICI therapy in in vivo models by restoring the infiltration of lymphocytes, mainly CD8+ T cells. These results delineate a tumor-LEC-immune axis that deserts antitumoral lymphocytes and highlight VEGFR-3 blockade as a highly desirable target in HPV-negative HNSCC and other ICI-refractory subpopulations. Biological sciences/Cancer/Head and neck cancer Biological sciences/Cancer/Tumour immunology/Immunosurveillance Biological sciences/Cancer/Tumour angiogenesis Full Text Additional Declarations There is NO Competing Interest. Supplementary Files FiguresSupplementary.pdf FigS1，FigS2，FigS3，FigS4.FigS5，FigS6 Cite Share Download PDF Status: Posted 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. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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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-8611043","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":581029025,"identity":"ea759c67-6fe1-42ff-92b3-b43a9e629e6d","order_by":0,"name":"Hao Li","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAxUlEQVRIiWNgGAWjYBACPmYeIFlBihY2sJYzJGlhAGphbCNJCzvvwQ8f59Ulzp+R/PADQ8U9uwb2swcIOIwvWXLmtsOJG26kGUswnClObuDJSyCghcdAmnfbgcQNEjlsQBcmJDNI8BgQ0mL8m3cOyGEkaDGT5m1gTmy4AdFiR5QWyxnHDhtvOPPMWCLhTEICG08Ofi38/GeMb3yoqZOd3w4MsQ8VCfb87Gfwa4EBxwYQmcDAkEh0HNljMEbBKBgFo2AUwAAAysU6cz8QQDUAAAAASUVORK5CYII=","orcid":"","institution":"Sun Yat-sen University Cancer center","correspondingAuthor":true,"prefix":"","firstName":"Hao","middleName":"","lastName":"Li","suffix":""}],"badges":[],"createdAt":"2026-01-15 13:45:18","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8611043/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8611043/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":104406398,"identity":"338a66e6-07b2-4890-94be-681d3e8b5338","added_by":"auto","created_at":"2026-03-11 12:25:56","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":50649621,"visible":true,"origin":"","legend":"","description":"","filename":"TumoreducatedVEGFR3lymphaticsaugmentPD1resistanceinHPVnegativeHNSCC.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8611043/v1_covered_63773b71-ba47-4ce1-8445-dd0f95ead79f.pdf"},{"id":101292682,"identity":"73d835f9-1d5b-4e81-ad1a-d08ba81eb0ad","added_by":"auto","created_at":"2026-01-28 08:27:57","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":6446056,"visible":true,"origin":"","legend":"FigS1\u0026#xFF0C;FigS2\u0026#xFF0C;FigS3\u0026#xFF0C;FigS4.FigS5\u0026#xFF0C;FigS6","description":"","filename":"FiguresSupplementary.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8611043/v1/cf8c6c01b33d1994d91505ee.pdf"}],"financialInterests":"There is \u003cb\u003eNO\u003c/b\u003e Competing Interest.","formattedTitle":"Tumor-educated VEGFR-3 lymphatics augment PD-1 resistance in HPV-negative HNSCC","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-8611043/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8611043/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe human papillomavirus (HPV) status is closely related to the response to immune checkpoint inhibitors (ICIs) in head and neck squamous cell carcinoma (HNSCC), yet the mechanisms of immunotherapy resistance in HPV-negative disease remain unclear. 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