{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,11,4]],"date-time":"2026-11-04T20:03:15Z","timestamp":1793822595166,"version":"4.6.1"},"reference-count":33,"publisher":"BMJ","issue":"8","license":[{"start":{"date-parts":[[2025,8,18]],"date-time":"2025-08-18T00:00:00Z","timestamp":1755475200000},"content-version":"unspecified","delay-in-days":17,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"DOI":"10.13039\/100006827","name":"Florida Department of Health","doi-asserted-by":"publisher","award":["24B04"],"award-info":[{"award-number":["24B04"]}],"id":[{"id":"10.13039\/100006827","id-type":"DOI","asserted-by":"publisher"}]},{"name":"NIH","award":["P30CA042014"],"award-info":[{"award-number":["P30CA042014"]}]},{"name":"NIH","award":["P30CA076292"],"award-info":[{"award-number":["P30CA076292"]}]}],"content-domain":{"domain":["bmj.com"],"crossmark-restriction":true},"short-container-title":["J Immunother Cancer"],"accepted":{"date-parts":[[2025,7,21]]},"published-print":{"date-parts":[[2025,8]]},"abstract":"<jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>A significant percentage of melanomas are refractory to immune checkpoint inhibitor (ICI) monotherapies and combinations. As there are currently no effective second-line therapies available for ICI-resistant patients, we sought to identify novel checkpoint inhibitor combinations for future clinical evaluation.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>We used responding and resistant SM1, SW1 and B16-10 melanoma models to explore the mechanism of action of anti-programmed cell death protein 1 (PD-1)+lymphocyte activation gene (LAG-3), anti-PD-1+cytotoxic lymphocyte associated protein 4 (CTLA-4), the anti-PD-1+LAG-3+CTLA-4 triplet, the anti-PD-1+T-cell immunoglobin and mucin domain 3 (TIM-3) doublet and the anti-PD-1+LAG-3+TIM-3 triplet. Flow cytometry, tetramer binding assays, T-cell depletion and analysis of the tumor-draining lymph nodes were used to determine the mechanisms of action of the ICI combinations. Single-cell RNA sequencing (scRNA-Seq) data from patients with melanoma was additionally analyzed to determine immune checkpoint expression across T-cell subsets and their association with ICI resistance.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>The PD-1+CTLA-4+LAG-3 triplet combination had good efficacy in the SM1 and B16-F10 melanoma models and led to a more favorable immune microenvironment (increased CD69+CD8+ T cells and gp-100-tumor specific T cells, reduced exhausted T cells) than either doublet. The ICI-resistant SW1 model did not respond to the PD-1+CTLA-4+LAG-3 triplet and was characterized by an accumulation of TIM-3+terminally exhausted CD8+T cells. Targeting of TIM-3 using the anti-PD-1+LAG-3+TIM-3 triplet led to complete tumor regressions in the SW1 and SM1 models. The TIM-3-based triplet decreased exhausted CD8+T cells and regulatory T cells, increased numbers of gp-100 reactive CD8+T cells and increased progenitor-exhausted CD8+T cells (TCF7+Ki67+ TIM-3\u2212) numbers in tumor-draining lymph nodes. scRNA-Seq analysis demonstrated that TIM-3 levels were higher in the CD8+T cells of patients with melanoma who were non-responsive to immunotherapy. PD-1 and TIM-3 expression was strongly correlated in the CD8+T cells from patients with melanoma, providing the rationale for co-targeting these checkpoints.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>Melanoma models characterized by high levels of terminally exhausted CD8+T cells (TIM-3+) can be successfully treated with the anti-PD-1+LAG-3+TIM-3 combination. The observation that high TIM-3 expression in CD8+T cells is associated with ICI resistance suggests the potential utility of anti-TIM-3-based combinations as a second-line treatment strategy in patients with advanced melanoma.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1136\/jitc-2025-012011","type":"journal-article","created":{"date-parts":[[2025,8,18]],"date-time":"2025-08-18T15:25:12Z","timestamp":1755530712000},"page":"e012011","update-policy":"https:\/\/doi.org\/10.1136\/crossmarkpolicy","source":"Crossref","is-referenced-by-count":8,"title":["Identification of anti-TIM-3 based checkpoint inhibitor combinations with activity in immunotherapy refractory melanoma models"],"prefix":"10.1136","volume":"13","clinical-trial-number":[{"clinical-trial-number":"nct03459222","registry":"10.18810\/clinical-trials-gov"}],"author":[{"given":"Manali S","family":"Phadke","sequence":"first","affiliation":[{"name":"Department of Tumor Microenvironment and Metastasis, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jiannong","family":"Li","sequence":"additional","affiliation":[{"name":"Department of Biostatistics and Bioinformatics, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sathya","family":"Sriramareddy","sequence":"additional","affiliation":[{"name":"Department of Tumor Microenvironment and Metastasis, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paulo C","family":"Rodriguez","sequence":"additional","affiliation":[{"name":"Department of Immunology, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3846-6872","authenticated-orcid":false,"given":"Brian","family":"Ruffell","sequence":"additional","affiliation":[{"name":"Department of Immunology, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vincent C","family":"Luca","sequence":"additional","affiliation":[{"name":"Department of Immunology, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Thuy T T","family":"Tran","sequence":"additional","affiliation":[{"name":"Departments of Internal Medicine and Population Health Sciences, University of Utah, Salt Lake City, Utah, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yian Ann","family":"Chen","sequence":"additional","affiliation":[{"name":"Department of Biostatistics and Bioinformatics, University of Utah, Salt Lake City, Utah, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1089-9307","authenticated-orcid":false,"given":"Keiran S M","family":"Smalley","sequence":"additional","affiliation":[{"name":"Department of Tumor Microenvironment and Metastasis, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"},{"name":"Department of Cutaneous Oncology, H Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"239","published-online":{"date-parts":[[2025,8,18]]},"reference":[{"key":"2026052108403069000_13.8.e012011.1","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa1003466"},{"key":"2026052108403069000_13.8.e012011.2","doi-asserted-by":"crossref","DOI":"10.1136\/jitc-2019-000398","article-title":"Defining tumor resistance to PD-1 pathway blockade: recommendations from the first meeting of the SITC Immunotherapy Resistance Taskforce","volume":"8","author":"Kluger","year":"2020","journal-title":"J Immunother Cancer"},{"key":"2026052108403069000_13.8.e012011.3","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2017.01.017"},{"key":"2026052108403069000_13.8.e012011.4","doi-asserted-by":"crossref","DOI":"10.3389\/fphar.2017.00561","article-title":"PD-1 and PD-L1 Checkpoint Signaling Inhibition for Cancer Immunotherapy: Mechanism, Combinations, and Clinical Outcome","volume":"8","author":"Alsaab","year":"2017","journal-title":"Front Pharmacol"},{"key":"2026052108403069000_13.8.e012011.5","doi-asserted-by":"crossref","DOI":"10.1038\/s41467-024-51930-5","article-title":"Structural basis for mouse LAG3 interactions with the MHC class II molecule I-Ab","volume":"15","author":"Ming","year":"2024","journal-title":"Nat Commun"},{"key":"2026052108403069000_13.8.e012011.6","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-022-05368-8"},{"key":"2026052108403069000_13.8.e012011.7","doi-asserted-by":"crossref","DOI":"10.1136\/jitc-2023-007239","article-title":"Differential requirements for CD4+ T cells in the efficacy of the anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 combinations in melanoma flank and brain metastasis models","volume":"11","author":"Phadke","year":"2023","journal-title":"J Immunother Cancer"},{"key":"2026052108403069000_13.8.e012011.8","doi-asserted-by":"crossref","DOI":"10.1136\/jitc-2020-000911","article-title":"Tim-3 finds its place in the cancer immunotherapy landscape","volume":"8","author":"Acharya","year":"2020","journal-title":"J Immunother Cancer"},{"key":"2026052108403069000_13.8.e012011.9","doi-asserted-by":"publisher","DOI":"10.1038\/415536a"},{"key":"2026052108403069000_13.8.e012011.10","doi-asserted-by":"publisher","DOI":"10.1038\/ni987"},{"key":"2026052108403069000_13.8.e012011.11","doi-asserted-by":"publisher","DOI":"10.1084\/jem.20100637"},{"key":"2026052108403069000_13.8.e012011.12","doi-asserted-by":"publisher","DOI":"10.1084\/jem.20100643"},{"key":"2026052108403069000_13.8.e012011.13","doi-asserted-by":"publisher","DOI":"10.1158\/0008-5472.CAN-11-0096"},{"key":"2026052108403069000_13.8.e012011.14","doi-asserted-by":"crossref","DOI":"10.1136\/jitc-2021-003571","article-title":"TIM-3 blockade enhances IL-12-dependent antitumor immunity by promoting CD8+ T cell and XCR1+ dendritic cell spatial co-localization","volume":"10","author":"Gardner","year":"2022","journal-title":"J Immunother Cancer"},{"key":"2026052108403069000_13.8.e012011.15","doi-asserted-by":"publisher","DOI":"10.1016\/j.immuni.2021.04.019"},{"key":"2026052108403069000_13.8.e012011.16","doi-asserted-by":"crossref","DOI":"10.1200\/JCO.2024.42.16_suppl.9504","article-title":"Efficacy and safety of triplet nivolumab, relatlimab, and ipilimumab (NIVO + RELA + IPI) in advanced melanoma: Results from RELATIVITY-048","volume":"42","author":"Ascierto","year":"2024","journal-title":"J Clin Oncol"},{"key":"2026052108403069000_13.8.e012011.17","doi-asserted-by":"publisher","DOI":"10.1158\/1078-0432.CCR-21-1694"},{"key":"2026052108403069000_13.8.e012011.18","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2018.10.038"},{"key":"2026052108403069000_13.8.e012011.19","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2024.06.036"},{"key":"2026052108403069000_13.8.e012011.20","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2024.07.018"},{"key":"2026052108403069000_13.8.e012011.21","doi-asserted-by":"crossref","DOI":"10.1080\/2162402X.2016.1261779","article-title":"Adaptive resistance to anti-PD1 therapy by Tim-3 upregulation is mediated by the PI3K-Akt pathway in head and neck cancer","volume":"6","author":"Shayan","year":"2017","journal-title":"Oncoimmunology"},{"key":"2026052108403069000_13.8.e012011.22","doi-asserted-by":"publisher","DOI":"10.1038\/ni1271"},{"key":"2026052108403069000_13.8.e012011.23","doi-asserted-by":"publisher","DOI":"10.1038\/nm.2871"},{"key":"2026052108403069000_13.8.e012011.24","doi-asserted-by":"publisher","DOI":"10.1038\/nature13848"},{"key":"2026052108403069000_13.8.e012011.25","doi-asserted-by":"publisher","DOI":"10.1038\/nature19330"},{"key":"2026052108403069000_13.8.e012011.26","doi-asserted-by":"publisher","DOI":"10.1126\/science.1229620"},{"key":"2026052108403069000_13.8.e012011.27","doi-asserted-by":"publisher","DOI":"10.1038\/s41590-019-0312-6"},{"key":"2026052108403069000_13.8.e012011.28","doi-asserted-by":"publisher","DOI":"10.1038\/s41590-022-01338-4"},{"key":"2026052108403069000_13.8.e012011.29","doi-asserted-by":"crossref","DOI":"10.1172\/jci.insight.124507","article-title":"Tumor-draining lymph nodes are pivotal in PD-1\/PD-L1 checkpoint therapy","volume":"3","author":"Fransen","year":"2018","journal-title":"JCI Insight"},{"key":"2026052108403069000_13.8.e012011.30","doi-asserted-by":"publisher","DOI":"10.1016\/j.ccell.2020.09.001"},{"key":"2026052108403069000_13.8.e012011.31","doi-asserted-by":"publisher","DOI":"10.1158\/1078-0432.CCR-18-1038"},{"key":"2026052108403069000_13.8.e012011.32","doi-asserted-by":"crossref","DOI":"10.1038\/ncomms10501","article-title":"Adaptive resistance to therapeutic PD-1 blockade is associated with upregulation of alternative immune checkpoints","volume":"7","author":"Koyama","year":"2016","journal-title":"Nat Commun"},{"key":"2026052108403069000_13.8.e012011.33","doi-asserted-by":"crossref","DOI":"10.3389\/fimmu.2021.654749","article-title":"Targeting STAT3 Abrogates Tim-3 Upregulation of Adaptive Resistance to PD-1 Blockade on Regulatory T Cells of Melanoma","volume":"12","author":"Huang","year":"2021","journal-title":"Front Immunol"}],"container-title":["Journal for ImmunoTherapy of Cancer"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.1136\/jitc-2025-012011","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,5,21]],"date-time":"2026-05-21T15:40:51Z","timestamp":1779378051000},"score":1,"resource":{"primary":{"URL":"https:\/\/jitc.bmj.com\/lookup\/doi\/10.1136\/jitc-2025-012011"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,8]]},"references-count":33,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2025,8,18]]},"published-print":{"date-parts":[[2025,8]]}},"alternative-id":["10.1136\/jitc-2025-012011"],"URL":"https:\/\/doi.org\/10.1136\/jitc-2025-012011","relation":{},"ISSN":["2051-1426"],"issn-type":[{"value":"2051-1426","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,8]]}}}