Inhibiting B-cell-mediated Immunosuppression to Enhance the Immunotherapy Efficacy in Hepatocellular Carcinoma

Inhibiting B-cell-mediated Immunosuppression to Enhance the Immunotherapy Efficacy in Hepatocellular Carcinoma | 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 Inhibiting B-cell-mediated Immunosuppression to Enhance the Immunotherapy Efficacy in Hepatocellular Carcinoma Dan Duda, Xin Liu, Zelong Liu, Tatsuya Kobayashi, Pin-Ji Lei, and 9 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6355345/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 08 Dec, 2025 Read the published version in Nature Communications → Version 1 posted You are reading this latest preprint version Abstract Background : Immunotherapy is efficacious in hepatocellular carcinoma (HCC), but the benefits are limited to a minority of patients. Most HCC patients show resistance to immune checkpoint blockade (ICB). Agonists of the stimulator of interferon genes (STING), potent immune stimulators, showed limited effectiveness. Using preclinical models, we studied the mechanisms of resistance to ICB and STING agonism. Methods : Murine HCA-1 and RIL-175 HCCs were orthotopically grown in mice with underlying liver fibrosis, to mimic the presentation of human HCC. Established tumors were treated with a STING agonist (BMS-986301) or anti-PD1 ICB, and mice were followed to evaluate safety and efficacy, as well as the mechanisms of treatment resistance by RNA sequencing, flow cytometry, and immunofluorescence, B-cell depletion and T-cell immunoglobulin and mucin domain 1 (TIM-1) ICB. Results : Unbiased analyses of transcriptomic data from murine HCC tissues from ICB-treated mice showed an increased abundance of intratumoral CD8+ T cells and B cells. STING agonism alone showed efficacy in the ICB-responsive RIL-175 HCC model but more limited efficacy in the ICB-resistant HCA-1 model. STING agonism increased circulating IL-10 and intratumoral infiltration by B-cells, including TIM-1+ B cells, and promoted the formation of tertiary lymphoid structure (TLS)-like structures, especially in the peritumoral areas. Strikingly, adding B cell depletion to ICB or STING agonism treatment significantly increased survival. Interestingly, unlike ICB, STING agonism also had a pronounced anti-metastatic activity. In addition, the combination of STING agonism and TIM-1 blockade augmented B cell differentiation and antigen presentation in vitro and improved the anti-tumor effects in murine HCC in vivo. This approach decreased the number of TIM-1+ B cells in the tumor and shifted B cells to higher expression of CD86 and MHC class II, enhancing the antigen presentation capability and further boosting the antitumor efficacy of CD8+ cytotoxic T cells. Conclusion : Our findings demonstrate that B cells are associated with ICB- and STING-mediated therapy resistance, and that depleting B-cells or targeting TIM-1 enhances both innate and acquired therapeutic efficacy in HCC. Biological sciences/Cancer/Cancer microenvironment Biological sciences/Cancer/Gastrointestinal cancer/Liver cancer/Hepatocellular carcinoma Hepatocellular carcinoma immunotherapy STING agonist B cell TIM-1 Full Text Additional Declarations Yes there is potential Competing Interest. DGD reports research grants from Bayer, Exelixis, and Surface Oncology, outside the submitted work. The other authors reported no disclosures. Cite Share Download PDF Status: Published Journal Publication published 08 Dec, 2025 Read the published version in Nature Communications → 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-6355345","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":442863071,"identity":"11f57bb7-7795-4fb6-a575-32171f4e5936","order_by":0,"name":"Dan 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The other authors reported no disclosures.","formattedTitle":"Inhibiting B-cell-mediated Immunosuppression to Enhance the Immunotherapy Efficacy in Hepatocellular Carcinoma","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"nature-portfolio","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"","title":"Nature Portfolio","twitterHandle":"","acdcEnabled":false,"dfaEnabled":false,"editorialSystem":"ejp","reportingPortfolio":"","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Hepatocellular carcinoma, immunotherapy, STING agonist, B cell, TIM-1","lastPublishedDoi":"10.21203/rs.3.rs-6355345/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6355345/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground\u003c/strong\u003e: Immunotherapy is efficacious in hepatocellular carcinoma (HCC), but the benefits are limited to a minority of patients. Most HCC patients show resistance to immune checkpoint blockade (ICB). Agonists of the stimulator of interferon genes (STING), potent immune stimulators, showed limited effectiveness. Using preclinical models, we studied the mechanisms of resistance to ICB and STING agonism.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods\u003c/strong\u003e: Murine HCA-1 and RIL-175 HCCs were orthotopically grown in mice with underlying liver fibrosis, to mimic the presentation of human HCC. Established tumors were treated with a STING agonist (BMS-986301) or anti-PD1 ICB, and mice were followed to evaluate safety and efficacy, as well as the mechanisms of treatment resistance by RNA sequencing, flow cytometry, and immunofluorescence, B-cell depletion and T-cell immunoglobulin and mucin domain 1 (TIM-1) ICB.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e: Unbiased analyses of transcriptomic data from murine HCC tissues from ICB-treated mice showed an increased abundance of intratumoral CD8+ T cells and B cells. STING agonism alone showed efficacy in the ICB-responsive RIL-175 HCC model but more limited efficacy in the ICB-resistant HCA-1 model. STING agonism increased circulating IL-10 and intratumoral infiltration by B-cells, including TIM-1+ B cells, and promoted the formation of tertiary lymphoid structure (TLS)-like structures, especially in the peritumoral areas. Strikingly, adding B cell depletion to ICB or STING agonism treatment significantly increased survival. Interestingly, unlike ICB, STING agonism also had a pronounced anti-metastatic activity. In addition, the combination of STING agonism and TIM-1 blockade augmented B cell differentiation and antigen presentation in vitro and improved the anti-tumor effects in murine HCC in vivo. This approach decreased the number of TIM-1+ B cells in the tumor and shifted B cells to higher expression of CD86 and MHC class II, enhancing the antigen presentation capability and further boosting the antitumor efficacy of CD8+ cytotoxic T cells.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion\u003c/strong\u003e: Our findings demonstrate that B cells are associated with ICB- and STING-mediated therapy resistance, and that depleting B-cells or targeting TIM-1 enhances both innate and acquired therapeutic efficacy in HCC.\u003c/p\u003e","manuscriptTitle":"Inhibiting B-cell-mediated Immunosuppression to Enhance the Immunotherapy Efficacy in Hepatocellular Carcinoma","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-16 03:52:38","doi":"10.21203/rs.3.rs-6355345/v1","editorialEvents":[],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"nature-communications","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"NCOMMS","sideBox":"Learn more about [Nature Communications](http://www.nature.com/ncomms/)","snPcode":"","submissionUrl":"https://mts-ncomms.nature.com/","title":"Nature Communications","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature Communications","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"2b71c520-2240-4b97-aa3e-bf4e5ae01cfd","owner":[],"postedDate":"April 16th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":47148852,"name":"Biological sciences/Cancer/Cancer microenvironment"},{"id":47148853,"name":"Biological sciences/Cancer/Gastrointestinal cancer/Liver cancer/Hepatocellular carcinoma"}],"tags":[],"updatedAt":"2025-12-09T08:09:08+00:00","versionOfRecord":{"articleIdentity":"rs-6355345","link":"https://doi.org/10.1038/s41467-025-66581-3","journal":{"identity":"nature-communications","isVorOnly":false,"title":"Nature Communications"},"publishedOn":"2025-12-08 05:00:00","publishedOnDateReadable":"December 8th, 2025"},"versionCreatedAt":"2025-04-16 03:52:38","video":"","vorDoi":"10.1038/s41467-025-66581-3","vorDoiUrl":"https://doi.org/10.1038/s41467-025-66581-3","workflowStages":[]},"version":"v1","identity":"rs-6355345","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6355345","identity":"rs-6355345","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}