Proteotranscriptomic Dissection of Breast Cancer T Cell States Identifies CD103+ Tfh-derived Cytotoxic Cells Linked to Immunotherapy Response

Proteotranscriptomic Dissection of Breast Cancer T Cell States Identifies CD103+ Tfh-derived Cytotoxic Cells Linked to Immunotherapy Response | 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 Proteotranscriptomic Dissection of Breast Cancer T Cell States Identifies CD103+ Tfh-derived Cytotoxic Cells Linked to Immunotherapy Response Alexander Swarbrick, Ghamdan Al-Eryani, Sophie van der Leij, Etienne Masle-Farquhar, and 16 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8394722/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 While cancer immunotherapies have primarily focused on activation of cytotoxic CD8 cells, CD4 T cell activity is also associated with survival and immunotherapeutic response in numerous cancers. We applied integrated single-cell RNA sequencing and multiplexed protein epitope profiling to breast cancer samples to resolve the complexity of immune cell states within the tumor microenvironment. This approach enhanced phenotypic resolution, identifying three distinct states within the CD4 T follicular helper-like (Tfh) cell cluster. A CXCR4high progenitor state gave rise to two differentiated states: an IGFL2high subset resembling conventional Tfh cells and localised to B cell-rich lymphoid aggregates, and a CD103+ subset, exhibiting features of tissue residency, exhaustion, and cytotoxicity, which co-localised with tumor foci. CD103+ Tfh-like cells were found to interact with CXCL10+ macrophages through production of CCL chemokines and CSF1. A higher CD103+ Tfh to IGFL2high Tfh ratio, together with the selective clonal expansion of the CD103+ subset, was strongly associated with improved tumour immunity and superior responses to anti-PD-1 checkpoint blockade, surpassing the predictive value of exhausted CD8 T cells. These findings integrate Tfh and CD4 with cytotoxic potential in breast cancer, offering new insight into anti-tumor immunity and response to checkpoint blockade. Biological sciences/Cancer/Cancer microenvironment Biological sciences/Cancer/Tumour immunology Biological sciences/Molecular biology/Transcription Health sciences/Medical research/Biomarkers Biological sciences/Immunology/Tumour immunology Full Text Additional Declarations Yes there is potential Competing Interest. This work was supported by subsidised reagents provided by Biolegend. BY was an employee of Biolegend at the time this study was conducted. C.M.P is an equity stockholder and consultant of BioClassifier LLC; C.M.P is also listed as an inventor on patent applications for the Breast PAM50 Subtyping assay Supplementary Files SupplementarytableS1CITEpanelADT.xlsx Supplementary_table_S1 SupplementarytableS2patientSampleInfo.xlsx Supplementary_table_S2 SupplementarytableS5Correlationprotein.csv Supplementary_table_S5 SupplementaryTableS8flowantibodies.xlsx Supplementary_Table_S8 SupplementarytableS4Otherstudygenesignatures.csv Supplementary_table_S4 SupplementarytableS6Xeniumpanel.xlsx Supplementary_table_S6 SupplementarytableS3DGEout.xlsx Supplementary_table_S3 CombinedExtendedDataFigures2.pdf Combined Extended Data Figures SupplementarytableS7Tfhsurvivalgenes.xlsx Supplementary_table_S7 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-8394722","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":569577395,"identity":"df0763dc-0f52-4212-9ba6-1024590072eb","order_by":0,"name":"Alexander 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