Spatial Transcriptomics Reveals Stromal-Partitioned Growth in Primary Colorectal Cancer and Host-Permeated Architecture in Liver Metastases

Spatial Transcriptomics Reveals Stromal-Partitioned Growth in Primary Colorectal Cancer and Host-Permeated Architecture in Liver Metastases | 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 Research Article Spatial Transcriptomics Reveals Stromal-Partitioned Growth in Primary Colorectal Cancer and Host-Permeated Architecture in Liver Metastases Tremayne Singh This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8852518/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 spatial organisation of the tumour microenvironment plays a central role in shaping immune engagement and tumour progression, yet how spatial principles differ between primary colorectal cancer and metastatic disease remains incompletely defined. Spatial transcriptomics (10x Visium) was integrated with cell-type deconvolution and transcriptional pathway enrichment to characterise tumour, stromal, vascular, and immune architecture across primary colorectal tumours (CRC) and colorectal liver metastases (CRLM). Across primary CRC sections, spatial organisation resolved into compartmentalised architectures in which hyper-proliferative tumour regions enriched for E2F and MYC signalling were segregated from immune-reactive compartments by cancer associated fibroblast (CAF) enriched stromal interfaces. These stromal regions were transcriptionally active, exhibiting enrichment of epithelial–mesenchymal transition, coagulation, and vascular programmes. Cytotoxic immune activity was heterogeneous and spatially constrained, localising to discrete tumour-associated niches or peripheral inflammatory zones rather than uniformly penetrating proliferative tumour cores. In contrast, liver metastases displayed diffuse, host-permeated architectures in which tumour programmes were embedded within resident hepatic immune and vascular frameworks. Metastatic growth was characterised by infiltrative expansion accompanied by strong myeloid and regulatory immune signatures, alongside spatially ordered transitions from stromal–vascular programmes into proliferative tumour states within a highly reactive hepatic environment. Together, these findings demonstrate that primary and metastatic colorectal cancer are governed by distinct spatial organisational principles. While primary CRC frequently exhibits stromal-partitioned, exclusion-prone architectures, liver metastases are shaped by infiltrative growth within a tolerogenic and highly reactive host context. This context-dependent spatial biology underscores the need for spatially tailored therapeutic strategies, targeting stromal architecture in primary tumours versus immune tolerance in metastatic disease. Bioinformatics Immunology Cancer Biology Bioinformatics Spatial Transcriptomics Genomics Colorectal Cancer Immunology Full Text Additional Declarations The authors declare no competing interests. Supplementary Files DESummaryTopGenes.csv Supplementary Table 1, Differential expression of clusters GSEAAxisSummary.csv Supplementary Table 2, Gene set enrichment analysis axis summary of clusters GSEASummaryTopPathways.csv Supplementary Table 2, Gene set enrichment analysis of clusters 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. 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-8852518","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":589668178,"identity":"b2580a25-f7bb-404d-8cba-735fe5c529c8","order_by":0,"name":"Tremayne Singh","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABBElEQVRIiWNgGAWjYDADCQbGBiCyATIZGw/gU8mDpiUNpKWBWC1gxYfBHLxa7KUPH93ws+2OnGT/4baHX3ect1vbfhhoS41NNE5b+NLSbva2PTOWlkhsN5Y9czt525lEoJZjabkNuLTw8Jjd4G07nDhPgrFNWrLtdrLZAaAWoAvxaOH/dvNv2+H6efwHQVrOJZudf0hICw/bbaAtCdIMiW2SH9sO2JndIGTLGTaz2zLnDhvOnJHYJs3YlpxgdgNoSwIev7D3MD+7+abssLzE+ePPJH+22dmbnU9/+OBDjQ1OLWDAyAahmYGxlAhWmYBPORj8gWr9AYwogopHwSgYBaNgxAEAARdmjvUv6K8AAAAASUVORK5CYII=","orcid":"","institution":"Liverpool John Moores University","correspondingAuthor":true,"prefix":"","firstName":"Tremayne","middleName":"","lastName":"Singh","suffix":""}],"badges":[],"createdAt":"2026-02-11 13:54:42","currentVersionCode":1,"declarations":{"humanSubjects":false,"vertebrateSubjects":false,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":false},"doi":"10.21203/rs.3.rs-8852518/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8852518/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":102746757,"identity":"b6be102b-c96c-40d7-b747-94e78edaa7be","added_by":"auto","created_at":"2026-02-16 09:01:07","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":15710870,"visible":true,"origin":"","legend":"","description":"","filename":"paper.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8852518/v1_covered_a5e54fc0-c7e3-4864-8872-37449ba3d20f.pdf"},{"id":102517824,"identity":"c4274fe5-fd9b-4e09-9f2c-cd98ce2d38a8","added_by":"auto","created_at":"2026-02-12 14:00:37","extension":"csv","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":123505,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Table 1\u003c/strong\u003e, \u003cem\u003eDifferential expression of clusters\u003c/em\u003e\u003c/p\u003e","description":"","filename":"DESummaryTopGenes.csv","url":"https://assets-eu.researchsquare.com/files/rs-8852518/v1/3f56b5cdea72b18098bcb0e8.csv"},{"id":102517823,"identity":"1cb2893e-ec72-4c33-86ad-c5b67650dc4e","added_by":"auto","created_at":"2026-02-12 14:00:37","extension":"csv","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":36153,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Table 2\u003c/strong\u003e, \u003cem\u003eGene set enrichment analysis axis summary of clusters\u003c/em\u003e\u003c/p\u003e","description":"","filename":"GSEAAxisSummary.csv","url":"https://assets-eu.researchsquare.com/files/rs-8852518/v1/80acd3183c53d4d853e3cd10.csv"},{"id":102517825,"identity":"e5de19d9-981e-4a92-9393-7215011af414","added_by":"auto","created_at":"2026-02-12 14:00:37","extension":"csv","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":246829,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Table 2\u003c/strong\u003e, \u003cem\u003eGene set enrichment analysis of clusters\u003c/em\u003e\u003c/p\u003e","description":"","filename":"GSEASummaryTopPathways.csv","url":"https://assets-eu.researchsquare.com/files/rs-8852518/v1/e1da23a19067f119f95249c8.csv"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003eSpatial Transcriptomics Reveals Stromal-Partitioned Growth in Primary Colorectal Cancer and Host-Permeated Architecture in Liver Metastases\u003c/p\u003e","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Liverpool John Moores University","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":true,"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":"Bioinformatics, Spatial Transcriptomics, Genomics, Colorectal Cancer, Immunology","lastPublishedDoi":"10.21203/rs.3.rs-8852518/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8852518/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe spatial organisation of the tumour microenvironment plays a central role in shaping immune engagement \u0026nbsp;and tumour progression, yet how spatial principles differ between primary colorectal cancer and metastatic \u0026nbsp;disease remains incompletely defined. Spatial transcriptomics (10x Visium) was integrated with cell-type \u0026nbsp;deconvolution and transcriptional pathway enrichment to characterise tumour, stromal, vascular, and immune \u0026nbsp;architecture across primary colorectal tumours (CRC) and colorectal liver metastases (CRLM).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAcross primary CRC sections, spatial organisation resolved into compartmentalised architectures in which \u0026nbsp;hyper-proliferative tumour regions enriched for E2F and MYC signalling were segregated from immune-reactive \u0026nbsp;compartments by cancer associated fibroblast (CAF) enriched stromal interfaces. These stromal regions were \u0026nbsp;transcriptionally active, exhibiting enrichment of epithelial–mesenchymal transition, coagulation, and vascular \u0026nbsp;programmes. Cytotoxic immune activity was heterogeneous and spatially constrained, localising to discrete \u0026nbsp;tumour-associated niches or peripheral inflammatory zones rather than uniformly penetrating proliferative \u0026nbsp;tumour cores.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIn contrast, liver metastases displayed diffuse, host-permeated architectures in which tumour programmes \u0026nbsp;were embedded within resident hepatic immune and vascular frameworks. Metastatic growth was \u0026nbsp;characterised by infiltrative expansion accompanied by strong myeloid and regulatory immune signatures, \u0026nbsp;alongside spatially ordered transitions from stromal–vascular programmes into proliferative tumour states \u0026nbsp;within a highly reactive hepatic environment.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTogether, these findings demonstrate that primary and metastatic colorectal cancer are governed by distinct \u0026nbsp;spatial organisational principles. While primary CRC frequently exhibits stromal-partitioned, exclusion-prone \u0026nbsp;architectures, liver metastases are shaped by infiltrative growth within a tolerogenic and highly reactive host \u0026nbsp;context. This context-dependent spatial biology underscores the need for spatially tailored therapeutic \u0026nbsp;strategies, targeting stromal architecture in primary tumours versus immune tolerance in metastatic disease.\u003c/p\u003e","manuscriptTitle":"Spatial Transcriptomics Reveals Stromal-Partitioned Growth in Primary Colorectal Cancer and Host-Permeated Architecture in Liver Metastases","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-02-12 14:00:27","doi":"10.21203/rs.3.rs-8852518/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","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}}],"origin":"","ownerIdentity":"02bc2a62-29cd-41ee-987c-32f99792c806","owner":[],"postedDate":"February 12th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":62745194,"name":"Bioinformatics"},{"id":62745195,"name":"Immunology"},{"id":62745196,"name":"Cancer Biology"}],"tags":[],"updatedAt":"2026-02-12T14:00:27+00:00","versionOfRecord":[],"versionCreatedAt":"2026-02-12 14:00:27","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8852518","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8852518","identity":"rs-8852518","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}