GPNMB-Mediated Immune Modulation in Triple- Negative Breast Cancer: Transcriptomic Analysis of GPNMB Knockdown in TAM Co-Cultures Reveals Therapeutic Targeting Opportunities

GPNMB-Mediated Immune Modulation in Triple- Negative Breast Cancer: Transcriptomic Analysis of GPNMB Knockdown in TAM Co-Cultures Reveals Therapeutic Targeting Opportunities | 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 GPNMB-Mediated Immune Modulation in Triple- Negative Breast Cancer: Transcriptomic Analysis of GPNMB Knockdown in TAM Co-Cultures Reveals Therapeutic Targeting Opportunities Ranit Ghosh, Arghya Adhikary This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8976503/v3 This work is licensed under a CC BY 4.0 License Status: Posted Version 3 posted You are reading this latest preprint version Show more versions Abstract Triple Negative Breast Cancer (TNBC) can be regarded as one of the most aggressive and therapeutically chal- lenging forms of cancer. It is responsible for 10–15% of all breast cancer cases worldwide along with significantly low success rate in prognosis when compared to hormone receptor positive (ER, PR, HER2) tumors. The immunosup- pressive tumor microenvironment (TME) characterized by abundant tumor associated macrophages (TAMs) with M2-like pro tumoral phenotypes, plays a critical role in TNBC progression and also in therapy failure. Glycoprotein non-metastatic melanoma protein B(GPNMB) has emerged as one of the key immunomodulatory factors with the ability to polarize macrophages toward immunosuppressive states and facilitating immune evasion. This analyti- cal study presents a comprehensive transcriptomic analysis of GPNMB knockdown (siGPNMB1 and siGPNMB2) in co-cultured TNBC cancer cells with THP1 derived macrophages when compared to control conditions. Using RNAseq differential gene expression (DGE) analysis with DESeq2 and pathway enrichment analysis through Gene Set Enrichment Analysis (GSEA), we were able to identify 232 genes which show significant differential expres- sion. GPNMB knockdown resulted in substantial downregulation of pro-tumoral inflammatory programs including IL6-JAK-STAT3 signalling, TGFβ signalling, and epithelial mesenchymal transition (EMT), while upregulating ox- idative phosphorylation and MYC target pathways. Our findings demonstrate that GPNMB functions as a critical orchestrator of macrophage mediated immune suppression in TNBC, making it an attractive therapeutic target for re-educating the TME and enhancing anti tumor immunity. Triple-negative breast cancer (TNBC) Glycoprotein non-metastatic melanoma protein B (GP- NMB) tumor-associated macrophages (TAMs) transcriptomics RNA-seq GSEA immunotherapy Full Text Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 3 posted You are reading this latest preprint version Show more versions 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-8976503","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":604002121,"identity":"55ec201e-835a-4724-b59c-639ae637a173","order_by":0,"name":"Ranit Ghosh","email":"","orcid":"","institution":"Jadavpur University","correspondingAuthor":false,"prefix":"","firstName":"Ranit","middleName":"","lastName":"Ghosh","suffix":""},{"id":604002122,"identity":"eb8d395f-c472-4ba8-bb88-51db2945a76b","order_by":1,"name":"Arghya Adhikary","email":"data:image/png;base64,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","orcid":"","institution":"Jadavpur University","correspondingAuthor":true,"prefix":"","firstName":"Arghya","middleName":"","lastName":"Adhikary","suffix":""}],"badges":[],"createdAt":"2026-02-26 10:09:42","currentVersionCode":3,"declarations":{"humanSubjects":false,"vertebrateSubjects":false,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":false},"doi":"10.21203/rs.3.rs-8976503/v3","doiUrl":"https://doi.org/10.21203/rs.3.rs-8976503/v3","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":104798101,"identity":"9a12fc5d-2790-49f1-ba75-eff115fe9fec","added_by":"auto","created_at":"2026-03-17 09:58:20","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":738376,"visible":true,"origin":"","legend":"","description":"","filename":"GPNMB2.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8976503/v3_covered_8121eb3f-9cfa-419c-9ef5-262f5717b662.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"GPNMB-Mediated Immune Modulation in Triple- Negative Breast Cancer: Transcriptomic Analysis of GPNMB Knockdown in TAM Co-Cultures Reveals Therapeutic Targeting Opportunities","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Jadavpur University","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":"Triple-negative breast cancer (TNBC), Glycoprotein non-metastatic melanoma protein B (GP- NMB), tumor-associated macrophages (TAMs), transcriptomics, RNA-seq, GSEA, immunotherapy","lastPublishedDoi":"10.21203/rs.3.rs-8976503/v3","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8976503/v3","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eTriple Negative Breast Cancer (TNBC) can be regarded as one of the most aggressive and therapeutically chal- lenging forms of cancer. It is responsible for 10\u0026ndash;15% of all breast cancer cases worldwide along with significantly low success rate in prognosis when compared to hormone receptor positive (ER, PR, HER2) tumors. The immunosup- pressive tumor microenvironment (TME) characterized by abundant tumor associated macrophages (TAMs) with M2-like pro tumoral phenotypes, plays a critical role in TNBC progression and also in therapy failure. Glycoprotein non-metastatic melanoma protein B(GPNMB) has emerged as one of the key immunomodulatory factors with the ability to polarize macrophages toward immunosuppressive states and facilitating immune evasion. This analyti- cal study presents a comprehensive transcriptomic analysis of GPNMB knockdown (siGPNMB1 and siGPNMB2) in co-cultured TNBC cancer cells with THP1 derived macrophages when compared to control conditions. 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