Spatially Resolved Epithelial–Immune Heterogeneity Identifies an Endometriosis Subtype with Molecular Features of Endometriosis-Associated Ovarian Cancer

Spatially Resolved Epithelial–Immune Heterogeneity Identifies an Endometriosis Subtype with Molecular Features of Endometriosis-Associated Ovarian Cancer | 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 Help Center Sign In Submit a Preprint Cite Share Download PDF Article Spatially Resolved Epithelial–Immune Heterogeneity Identifies an Endometriosis Subtype with Molecular Features of Endometriosis-Associated Ovarian Cancer Valentina Iacobelli, Luca Mastrantoni, Stefano Alivernini, Giulia Scaglione, and 13 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-10497180/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 9 You are reading this latest preprint version Abstract Evidence suggests that clear cell and endometrioid ovarian carcinoma may arise from endometriosis. We previously identified that some endometriosis lesions exhibit a “cancer-like” PD-1/PD-L1 positive staining similar to endometriosis-associated ovarian cancers. If this condition is associated with oncogenic signaling and immune tolerance is currently unknown. Using spatial transcriptomics, we mapped epithelial cells, T lymphocytes and macrophages in 14 patients including simple endometriosis (n = 2), atypical endometriosis (n = 2), PD-L1-positive “cancer-like” endometriosis (n = 4), endometriosis-associated clear cell (n = 3) and endometrioid ovarian carcinoma (n = 3). We found that epithelial cells in cancer-like endometriosis, despite a conventional histological appearance, display significant enrichment in oncogenic signaling, extracellular matrix remodeling and wound-healing programs. This phenotype is further defined by epithelial NNMT and MUC5B expression, subepithelial neutrophil infiltration, and a shift toward an M2-macrophage-dominant, T-CD8 + cell depleted microenvironment. Proliferative signaling was enriched in endometroid carcinoma and adjacent endometriotic epithelium, while secretory signaling in cancer-like endometriosis, clear cell carcinoma and adjacent endometriotic areas. Our findings support a model of endometriosis-associated tumorigenesis through two different mechanisms: a damage-repair/inflammatory loop mimicking a “window-of-implantation-like” secretory niche or a hormone-driven proliferative signaling, providing a spatial framework for risk stratification. Biological sciences/Cancer Biological sciences/Cell biology Health sciences/Diseases Health sciences/Oncology Full Text Additional Declarations No competing interests reported. Supplementary Files SupplementaryMaterial.docx Cite Share Download PDF Status: Under Review Version 1 posted Reviews received at journal 26 Aug, 2026 Reviews received at journal 24 Aug, 2026 Reviewers agreed at journal 19 Aug, 2026 Reviewers agreed at journal 18 Aug, 2026 Reviewers agreed at journal 16 Aug, 2026 Reviewers invited by journal 15 Aug, 2026 Editor assigned by journal 09 Aug, 2026 Submission checks completed at journal 29 Jul, 2026 First submitted to journal 27 Jul, 2026 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 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-10497180","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":700452468,"identity":"ee080a78-f46a-404d-a50f-da29e244293a","order_by":0,"name":"Valentina Iacobelli","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Valentina","middleName":"","lastName":"Iacobelli","suffix":""},{"id":700452469,"identity":"0e84a2a0-5415-4267-8514-409d624264fc","order_by":1,"name":"Luca Mastrantoni","email":"","orcid":"","institution":"Catholic University of the Sacred Heart","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Luca","middleName":"","lastName":"Mastrantoni","suffix":""},{"id":700452470,"identity":"67eee2aa-cc73-4f79-a17d-90152791867e","order_by":2,"name":"Stefano Alivernini","email":"","orcid":"","institution":"Catholic University of the Sacred Heart","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Stefano","middleName":"","lastName":"Alivernini","suffix":""},{"id":700452471,"identity":"30b5d68c-33fc-4180-9119-fa4cb3f15151","order_by":3,"name":"Giulia Scaglione","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Giulia","middleName":"","lastName":"Scaglione","suffix":""},{"id":700452472,"identity":"1beb3b3c-79f4-4c7f-addc-d11f87b5e610","order_by":4,"name":"Antonio Agostini","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Antonio","middleName":"","lastName":"Agostini","suffix":""},{"id":700452473,"identity":"75d75045-6296-4956-ae39-2fa69921ddd0","order_by":5,"name":"Giulia Mantini","email":"","orcid":"","institution":"Gemelli Science and Technology Park (GSTeP), Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Giulia","middleName":"","lastName":"Mantini","suffix":""},{"id":700452474,"identity":"987db178-77e4-43b8-8b28-dd7be3ea55b3","order_by":6,"name":"Alessia Piermattei","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Alessia","middleName":"","lastName":"Piermattei","suffix":""},{"id":700452475,"identity":"82229770-0002-4a9c-8fd1-40d7162d0ca8","order_by":7,"name":"Federica Persiani","email":"","orcid":"","institution":"Gemelli Science and Technology Park (GSTeP), Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Federica","middleName":"","lastName":"Persiani","suffix":""},{"id":700452476,"identity":"c570b340-8cae-474c-82c3-e58919e72389","order_by":8,"name":"Floriana Camarda","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Floriana","middleName":"","lastName":"Camarda","suffix":""},{"id":700452477,"identity":"4880f27c-96b2-4353-a35a-0eb0f6ef45d6","order_by":9,"name":"Rita Trozzi","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Rita","middleName":"","lastName":"Trozzi","suffix":""},{"id":700452478,"identity":"0ca8b4b6-4c98-4a46-9f12-0e6c8c85ef25","order_by":10,"name":"Francesca Sillano","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Francesca","middleName":"","lastName":"Sillano","suffix":""},{"id":700452479,"identity":"9a439733-46f2-4940-a6c9-b72f0e416c0f","order_by":11,"name":"Maria Antonietta D’Agostino","email":"","orcid":"","institution":"Catholic University of the Sacred Heart","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Maria","middleName":"Antonietta","lastName":"D’Agostino","suffix":""},{"id":700452480,"identity":"eb2d683c-dca4-4fbd-8789-5b81f6156cc2","order_by":12,"name":"Georg Schett","email":"","orcid":"","institution":"Gemelli Science and Technology Park (GSTeP), Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Georg","middleName":"","lastName":"Schett","suffix":""},{"id":700452481,"identity":"b253d964-e7c0-4a82-8b08-599e70774ec9","order_by":13,"name":"Carmine Carbone","email":"","orcid":"","institution":"Fondazione Policlinico Universitario Agostino Gemelli, IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Carmine","middleName":"","lastName":"Carbone","suffix":""},{"id":700452482,"identity":"899faa22-3f15-435b-b1be-9cee1aec97ac","order_by":14,"name":"Giovanni Scambia","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Giovanni","middleName":"","lastName":"Scambia","suffix":""},{"id":700452483,"identity":"db6193b9-e2f0-48be-af1d-5a96b3b2e8cf","order_by":15,"name":"Anna Fagotti","email":"","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Anna","middleName":"","lastName":"Fagotti","suffix":""},{"id":700452484,"identity":"fb5d9ce8-d607-47f3-bc4b-5f543f658731","order_by":16,"name":"Camilla Nero","email":"data:image/png;base64,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","orcid":"","institution":"Fondazione Policlinico Universitario A. Gemelli IRCCS","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Camilla","middleName":"","lastName":"Nero","suffix":""}],"badges":[],"createdAt":"2026-07-27 10:40:07","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-10497180/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-10497180/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":118218060,"identity":"4b2b5b32-6f17-4908-a531-130948afd67f","added_by":"auto","created_at":"2026-08-25 07:34:04","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":7328883,"visible":true,"origin":"","legend":"","description":"","filename":"Manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-10497180/v1_covered_ab420caa-2420-48c2-8a42-a92fa9907c9f.pdf"},{"id":118213939,"identity":"5ebea30e-7093-43df-ae63-5903d105101f","added_by":"auto","created_at":"2026-08-25 06:05:44","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":4741433,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryMaterial.docx","url":"https://assets-eu.researchsquare.com/files/rs-10497180/v1/57f682d00c72e7ec387ca770.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Spatially Resolved Epithelial–Immune Heterogeneity Identifies an Endometriosis Subtype with Molecular Features of Endometriosis-Associated Ovarian Cancer","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"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":"npj-precision-oncology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"npjprecisiononcology","sideBox":"Learn more about [npj Precision Oncology](http://www.nature.com/npjprecisiononcology/)","snPcode":"41698","submissionUrl":"https://submission.springernature.com/new-submission/41698/3","title":"npj Precision Oncology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"NPJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-10497180/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-10497180/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eEvidence suggests that clear cell and endometrioid ovarian carcinoma may arise from endometriosis. We previously identified that some endometriosis lesions exhibit a \u0026ldquo;cancer-like\u0026rdquo; PD-1/PD-L1 positive staining similar to endometriosis-associated ovarian cancers. If this condition is associated with oncogenic signaling and immune tolerance is currently unknown.\u003c/p\u003e \u003cp\u003eUsing spatial transcriptomics, we mapped epithelial cells, T lymphocytes and macrophages in 14 patients including simple endometriosis (n\u0026thinsp;=\u0026thinsp;2), atypical endometriosis (n\u0026thinsp;=\u0026thinsp;2), PD-L1-positive \u0026ldquo;cancer-like\u0026rdquo; endometriosis (n\u0026thinsp;=\u0026thinsp;4), endometriosis-associated clear cell (n\u0026thinsp;=\u0026thinsp;3) and endometrioid ovarian carcinoma (n\u0026thinsp;=\u0026thinsp;3). We found that epithelial cells in cancer-like endometriosis, despite a conventional histological appearance, display significant enrichment in oncogenic signaling, extracellular matrix remodeling and wound-healing programs. This phenotype is further defined by epithelial \u003cem\u003eNNMT\u003c/em\u003e and \u003cem\u003eMUC5B\u003c/em\u003e expression, subepithelial neutrophil infiltration, and a shift toward an M2-macrophage-dominant, T-CD8\u0026thinsp;+\u0026thinsp;cell depleted microenvironment.\u003c/p\u003e \u003cp\u003eProliferative signaling was enriched in endometroid carcinoma and adjacent endometriotic epithelium, while secretory signaling in cancer-like endometriosis, clear cell carcinoma and adjacent endometriotic areas.\u003c/p\u003e \u003cp\u003eOur findings support a model of endometriosis-associated tumorigenesis through two different mechanisms: a damage-repair/inflammatory loop mimicking a \u0026ldquo;window-of-implantation-like\u0026rdquo; secretory niche or a hormone-driven proliferative signaling, providing a spatial framework for risk stratification.\u003c/p\u003e","manuscriptTitle":"Spatially Resolved Epithelial–Immune Heterogeneity Identifies an Endometriosis Subtype with Molecular Features of Endometriosis-Associated Ovarian Cancer","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-08-25 00:55:10","doi":"10.21203/rs.3.rs-10497180/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"editorInvitedReview","content":"","date":"2026-08-26T08:10:12+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-08-24T10:25:24+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"187626399193702419474010517992736822973","date":"2026-08-19T05:22:53+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"24040887931247321056772173954598410496","date":"2026-08-18T06:48:09+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"297671070129276515533240624440701549918","date":"2026-08-16T18:36:53+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-08-15T19:52:05+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-08-09T21:30:20+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-07-29T12:08:44+00:00","index":"","fulltext":""},{"type":"submitted","content":"npj Precision Oncology","date":"2026-07-27T10:26:26+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"npj-precision-oncology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"npjprecisiononcology","sideBox":"Learn more about [npj Precision Oncology](http://www.nature.com/npjprecisiononcology/)","snPcode":"41698","submissionUrl":"https://submission.springernature.com/new-submission/41698/3","title":"npj Precision Oncology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"NPJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ba7e07d4-9de9-436d-bbaf-84aeb4759b7e","owner":[],"postedDate":"August 25th, 2026","published":true,"recentEditorialEvents":[{"type":"editorInvitedReview","content":"","date":"2026-08-26T08:10:12+00:00","index":16,"fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-08-24T10:25:24+00:00","index":15,"fulltext":""},{"type":"reviewerAgreed","content":"187626399193702419474010517992736822973","date":"2026-08-19T05:22:53+00:00","index":14,"fulltext":""},{"type":"reviewerAgreed","content":"24040887931247321056772173954598410496","date":"2026-08-18T06:48:09+00:00","index":13,"fulltext":""},{"type":"reviewerAgreed","content":"297671070129276515533240624440701549918","date":"2026-08-16T18:36:53+00:00","index":12,"fulltext":""},{"type":"reviewersInvited","content":"7","date":"2026-08-15T19:52:05+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-08-09T21:30:20+00:00","index":"","fulltext":""}],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[{"id":74364754,"name":"Biological sciences/Cancer"},{"id":74364755,"name":"Biological sciences/Cell biology"},{"id":74364756,"name":"Health sciences/Diseases"},{"id":74364757,"name":"Health sciences/Oncology"}],"tags":[],"updatedAt":"2026-08-25T00:55:11+00:00","versionOfRecord":[],"versionCreatedAt":"2026-08-25 00:55:10","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-10497180","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-10497180","identity":"rs-10497180","version":["v1"]},"buildId":"-D5TCW68w8eVRRLyjaTIo","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}